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Modulation of cellular excitability in neocortex: Muscarinic receptor and second messenger-mediated actions of acetylcholine
Abstract
Muscarinic-type acetylcholine (ACh) receptor are involved in a variety of cortical functions. ACh "activates" neocortex; simultaneously modifying spontaneous subthreshold activity, intrinsic neuronal oscillations and spike discharge modes, and responsiveness to fast (putative glutamatergic) synaptic inputs. However, beyond the general involvement of muscarinic receptors, a mechanistic understanding of integrated cholinergic actions, and interactions with non-cholinergic transmission, is lacking. We have addressed this problem using intracellular recordings from the in vitro auditory neocortex. First, we investigated cholinergic modification of responses to the excitatory amino acid glutamate. ACh, or the muscarinic agonist methacholine, produced a lasting enhancement of glutamate-mediated membrane depolarizations. Muscarinic receptors of the M1 and/or M3 subtype, rather than M2 or nicotinic receptors, mediated this enhancement. Subsequently, we investigated whether second messenger systems contribute to observed muscarinic actions. Activation of protein kinase C with phorbol 12,13-dibutyrate (4 beta-PDBu), enhanced neuronal responses to glutamate. The effect of 4 beta-PDBu was attenuated by the kinase antagonist H7. Finally, we attempted to identify postsynaptic actions of endogenous ACh. Tetanic stimulation of cholinergic afferents elicited voltage-dependent effects, including reduced spike frequency adaptation and reduced slow afterhyperpolarization (sAHP) elicited by transmembrane depolarizing stimuli. These effects were mimicked by methacholine, enhanced by eserine, and antagonized by muscarinic receptor antagonists. These data suggest that cholinergic modulation in neocortex likely involves the integrated actions of diverse mechanisms, primarily gated by muscarinic receptors, and at least partly involving second messenger systems.
... That the 40-Hz oscillations depend on reciprocal interactions between the thalamus and cerebral cortex has been further demonstrated by intracellular and extracellular recordings in the cat brain [18]. Cholinergic transmission presumably has a central, although heterogeneous, in¯uence in modulating these thalamo-cortical and cortical auditoryneural connections, as suggested by human EEG [3] or MEG measurements [11], in vitro recordings from cortical slices [4], and in vivo animal studies [14]. While the cholinergic in¯uence on transient human 40- Hz evoked response has not yet been studied, in vitro studies on hippocampal slices have indicated that 40-Hz oscillations are induced by cholinergic agonist carbachol and abolished by the addition of the muscarinic M1 receptor antagonist pirenzepine [6]. ...
... Based on the cellular-level studies [2] [4] [14] indicating that muscarinic activation produces mainly excitatory effects in the auditory cortex, one might have expected that muscarinic antagonist, such as scopolamine, would have reduced the 40-Hz responses instead of augmenting them. Cholinergic modulation can, however, have either excitatory or inhibitory effects depending on the type of the postsynaptic neurone [16]. ...
... For example , the inhibitory GABA A -agonist temazepam reduced auditory g -oscillations in humans [10], whereas another drug inhibiting CNS function, the NMDA-antagonist ketamine augmented them [17]. Given that muscarinic receptors modulate glutamatergic transmission [4], the present augmentation of 40-Hz oscillations by scopolamine might therefore be mediated by a common mechanism with the ketamine-induced augmentation. Taken together, the apparent heterogeneity of previous 40-Hz results and cholinergic effects on brain function clearly warrants more detailed studies on the transmitter systems underlying these oscilla- tions. ...
The influence of neocortical muscarinic transmission on auditory-evoked 40-Hz magnetic response was studied in 13 healthy subjects in a double-blind randomized cross-over design. Either a centrally (scopolamine hydrobromide, 0.3 mg, i.v.) or a peripherally (glycopyrrolate, 0.2 mg, i.v.) acting antagonist of muscarinic transmission was administered during two sessions of magnetoencephalographic recording of 40-Hz response elicited by monaural tones. Scopolamine significantly (P<0.01) augmented the 40-Hz magnetic response over the hemispheres ipsi- and contralateral to the ear stimulated. This finding suggests muscarinic modulation of the auditory evoked transient 40-Hz response.
... The modulatory role of ACh in neocortical information processing has been implicated in many functions including arousal, attention, learning, and memory (Ashe and Weinberger 1991;Metherate et al. 1992;Nabeshima 1993). Of particular interest are findings that ACh, as well as other cholinergic agonists, modulate glutamatergic transmission by either presynaptic or postsynaptic mechanisms, or both (Yamamoto and Kawai 1967;Hounsgaard 1978;Segal 1989;Markram and Segal 1990;Lin and Phillis 1991;Krnjevic 1993;Cox et al. 1994;Kang 1995). These actions require specific subtypes of the muscarinic receptor with subsequent stimulation of intracellular second messenger systems (for review see Hosey 1992;Caulfield 1993). ...
... These actions require specific subtypes of the muscarinic receptor with subsequent stimulation of intracellular second messenger systems (for review see Hosey 1992;Caulfield 1993). In rat auditory cortex, long-lasting postsynaptic enhancement of glutamate depolarizations is mediated by the M 1 and/or M 3 , rather than the M 2 , muscarinic receptor subtype (Cox et al. 1994). This muscarinic action of ACh is essentially mimicked by activation of protein kinase C by phorbol 12,13dibutyrate (4-β-PDBu) and is attenuated by the kinase antagonist 1-(5-isoquinolinesulfonyl)-2-methylpiperazine hydrochloride (H7). ...
... The methods used for brain slice preparation and maintenance are similar to those described previously (Cox et al. , 1994Metherate and Ashe 1994). Male Sprague-Dawley rats (3-5 weeks old and weighing approximately 80-120 g) were anesthetized with pentobarbital sodium (Nembutal, 25 mg/kg). ...
The present study examines the ability of muscarinic receptor activation to modulate glutamatergic responses in the in vitro rat auditory cortex. Whole-cell patch-clamp recordings were obtained from layer II-III pyramidal neurons and responses elicited by either stimulation of deep gray matter or iontophoretic application of glutamate receptor agonists. Iontophoresis of the muscarinic agonist acetyl-beta-methylcholine (MCh) produced an atropine-sensitive reduction in the amplitude of glutamate-induced membrane depolarizations that was followed by a long-lasting (at least 20 min) response enhancement. Glutamate depolarizations were enhanced by MCh when elicited in the presence of alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA)/kainate receptor antagonists 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) or 2,3-dihydroxy-6-nitro-7-sulfamoyl, benzo(F)quinoxaline (NBQX) but not the NMDA antagonists D-2-amino-5-phosphonovaleric acid (APV) or MK-801 hydrogen maleate. The magnitude of enhancement was voltage-dependent with the percentage increase greater at more depolarized membrane potentials. An involvement of NMDA receptors in these MCh-mediated effects was tested by using AMPA/kainate receptor antagonists to isolate the NMDA-mediated slow excitatory postsynaptic potential (EPSP) from other synaptic potentials. The slow EPSP and iontophoretic responses to NMDA were similarly modified by MCh, i.e., both being reduced during and enhanced (15-55 min) following MCh application. Cholinergic modulation of NMDA responses involves the engagement of G proteins, as enhancement was prevented by intracellular infusion with the nonhydrolyzable GDP analog guanosine-5'-O-(2-thiodiphosphate) trilithium salt (GDPbetaS). GDPbetaS was without effect on the early MCh-induced response suppression. Our results suggest that acetylcholine, acting at muscarinic receptors, produces a long-lasting enhancement of NMDA-mediated neurotransmission in auditory cortex, and that this modulatory effect is dependent upon a G protein-mediated event.
... Stress effects on higher brain functions involve neuronal activation of several neurotransmitter systems, including glutamate [23], dopamine [24] and neuromodulation of both by acetylcholine (ACh) [25][26][27][28][29][30][31][32][33][34]. However, although SB is readily induced by dopamine agonists, and is modulated by stressful experiences [35,36], neither psychiatric patients nor animals with cage stereotypies showed excessive dopaminergic neurotransmission. ...
... Identifying the neurotransmission pathways involved in SB is also challenging, as receptors for the stress-response organizing agent CRF are present on glutamatergic, dopaminergic, serotonergic, noradrenergic and cholinergic neurons, indicating complex neuro-modulatory processes. Specifically, intraventricular CRF administration increases ACh release from cholinergic terminals in several forebrain regions controlling motor behavior [26], thereby potentiating principal neuron reactions to incoming messages via nicotinic receptor activation [25]. This indicates that cholinergic signals and their modification by miRNA regulators may upregulate motor functions under stress. ...
Stereotypic behavior (SB) is common in emotional stress-involved psychiatric disorders and is often attributed to glutamatergic impairments, but the underlying molecular mechanisms are unknown. Given the neuro-modulatory role of acetylcholine, we sought behavioral-transcriptomic links in SB using TgR transgenic mice with impaired cholinergic transmission due to over-expression of the stress-inducible soluble ‘readthrough’ acetylcholinesterase-R splice variant AChE-R. TgR mice showed impaired organization of behavior, performance errors in a serial maze test, escape-like locomotion, intensified reaction to pilocarpine and reduced rearing in unfamiliar situations. Small-RNA sequencing revealed 36 differentially expressed (DE) microRNAs in TgR mice hippocampi, 8 of which target more than 5 cholinergic transcripts. Moreover, compared to FVB/N mice, TgR prefrontal cortices displayed individually variable changes in over 400 DE mRNA transcripts, primarily acetylcholine and glutamate-related. Furthermore, TgR brains presented c-fos over-expression in motor behavior-regulating brain regions and immune-labeled AChE-R excess in the basal ganglia, limbic brain nuclei and the brain stem, indicating a link with the observed behavioral phenotypes. Our findings demonstrate association of stress-induced SB to previously unknown microRNA-mediated perturbations of cholinergic/glutamatergic networks and underscore new therapeutic strategies for correcting stereotypic behaviors.
... In vivo studies have shown that activation of mChR receptors enhance cortical responses to sensory inputs (Metherate et al., 1988;McKenna et al., 1989;Chen and Yan, 2007). In general, this activation increases post-synaptic membrane resistance (due to decreased potassium conductances) and leads to an increase in post-synaptic excitability via a slow EPSP and a decrease in the after hyperpolarization potential (Metherate et al., 1992;Cox et al., 1994). However, parts of these effects can also be evoked by muscarinic receptors located on particular types of interneurons (Disney and Aoki, 2008): it has been shown that activation of muscarinic receptors can increase neuronal responses by decreasing the release of GABA from interneurons in layers II/III (Salgado et al., 2007) and layer V (Kruglikov and Rudy, 2008). ...
... Beta and alpha2 receptors can both contribute to control the post-synaptic excitability of cortical cells Edeline, 1997, 1999;Salgado et al., 2011). (B) Muscarinic receptors increase the post-synaptic excitability of pyramidal cells (Metherate et al., 1992;Cox et al., 1994) but also of some types of interneurons (Disney and Aoki, 2008) and can decrease the release of GABA by Fast-Spiking interneurons (Kruglikov and Rudy, 2008). Nicotinic receptors can act presynaptically on the thalamo-cortical and can increase the excitability of several types of GABAergic interneurons (Disney et al., 2007;review in Metherate, 2004review in Metherate, , 2011. ...
Over the last two decades, a vast literature has described the influence of neuromodulatory systems on the responses of sensory cortex neurons (review in Gu, 2002; Edeline, 2003; Weinberger, 2003; Metherate, 2004, 2011). At the single cell level, facilitation of evoked responses, increases in signal-to-noise ratio, and improved functional properties of sensory cortex neurons have been reported in the visual, auditory, and somatosensory modality. At the map level, massive cortical reorganizations have been described when repeated activation of a neuromodulatory system are associated with a particular sensory stimulus. In reviewing our knowledge concerning the way the noradrenergic and cholinergic system control sensory cortices, I will point out that the differences between the protocols used to reveal these effects most likely reflect different assumptions concerning the role of the neuromodulators. More importantly, a gap still exists between the descriptions of neuromodulatory effects and the concepts that are currently applied to decipher the neural code operating in sensory cortices. Key examples that bring this gap into focus are the concept of cell assemblies and the role played by the spike timing precision (i.e., by the temporal organization of spike trains at the millisecond time-scale) which are now recognized as essential in sensory physiology but are rarely considered in experiments describing the role of neuromodulators in sensory cortices. Thus, I will suggest that several lines of research, particularly in the field of computational neurosciences, should help us to go beyond traditional approaches and, ultimately, to understand how neuromodulators impact on the cortical mechanisms underlying our perceptual abilities.
... It has also been demonstrated that activation of cholinergic receptors may influence the potential for the induction of activity-dependent long-lasting changes of synaptic transmission within the hippocampus (Ito et al. 1988, Williams and Johnston 1988, Hirotsu et al. 1989, Burgard and Sarvey 1990, Maeda et al. 1994) and visual cortex (Brocher et al. 1992 ). In the auditory cortex, cholinergic agonists acting on muscarinic receptors produce a lasting enhancement of glutamate-mediated membrane depolarizations (Cox et al. 1994 ). It has recently been shown that synaptic transmission within layers 111111 horizontal connections of rat motor cortex is mediated by non-NMDA and NMDA subtypes of glutamate receptors (Aroniadou and Keller 1993, Hess et al. 1994). ...
... The results obtained in the course of this study indicate that endogenous and exogenous cholinergic agonists may transiently depress excitatory synaptic transmission within layer 111111 horizontal connections of rat motor cortex in the in vitro slice preparations. The action of endogenous acetylcholjne on neuronal membrane properties has previously been observed in slices isolated from auditory (Metherate et al. 1992, Cox et al. 1994) and somatosensory cortex (Benardo 1993 ). The influence of muscarinic cholinergic activation on neocortical synaptic transmission, where investigated, has been shown to result in a reduction of the synaptic responses. ...
The influence of compounds interacting with cholinergic systems on field potentials evoked in layer II/III horizontal connections was investigated in rat motor cortex in vitro. The cholinesterase inhibitor eserine (10 microM) decreased field responses by 20 +/- 2%. This effect could be prevented by preincubation with atropine (10 microM). Application of 5 microM carbachol resulted in reduction of the responses by 30 +/- 1%. These reductions were reversible, repeatable and independent of stimulus intensity; they could be blocked by the M1 muscarinic receptor antagonist pirenzepine (3 microM) but not by the M2 muscarinic receptor antagonist gallamine (10 microM). During carbachol application, paired-pulse facilitation (40 ms interpulse interval) was increased. The results indicate that endogenous acetylcholine may modulate excitatory synaptic transmission in horizontal connections of rat motor cortex, most likely by acting upon M1 receptors located presynaptically on glutamatergic terminals, and may contribute both to information processing and synaptic plasticity within the motor cortex.
... ACh released in somatosensory cortex acts upon several muscarinic receptors (cf. Dykes 1990; Cox et al. 1994); one effect is to close a potassium channel so that the action potential is broadened, allowing the NMDA channel time to open and triggering the cascade of events that leads to LTP. Another muscarinic receptor is a channel that directly permits calcium entry to activate a similar cascade leading to enhanced neuronal responsiveness (Metherate and Ashe 1993b; Benardo and Prince 1982; Madison et al. 1987; McCormick and Prince 1985; Cox et al. 1994; Haj-Dahmane and Andrade 1996; Fraser and MacVicar 1996). ...
... Dykes 1990; Cox et al. 1994); one effect is to close a potassium channel so that the action potential is broadened, allowing the NMDA channel time to open and triggering the cascade of events that leads to LTP. Another muscarinic receptor is a channel that directly permits calcium entry to activate a similar cascade leading to enhanced neuronal responsiveness (Metherate and Ashe 1993b; Benardo and Prince 1982; Madison et al. 1987; McCormick and Prince 1985; Cox et al. 1994; Haj-Dahmane and Andrade 1996; Fraser and MacVicar 1996). We refer to these long-lasting effects of ACh as long-term cholinergic enhancement (LTCE). ...
gamma-Aminobutyric acid containing neurons in the somatosensory cortex are the major controlling element determining receptive field size and location. This control of the excitability of cortical neurons can be modulated by activity arising in the basal forebrain. A hypothesis is developed stating that both cholinergic and gamma-aminobutyric acid containing projections from the basal forebrain play important roles in the production of a state in cortex permitting neuronal plasticity to occur. This proposed mechanism involving a simultaneous reduction of inhibition and increased release of acetylcholine allows sensory signals to induce long-term changes in the location and responsiveness of cutaneous receptive fields, thereby changing the somatotopic organization of primary somatosensory cortex.
... In terms of chemoarchitecture, the distribution of receptors and enzymes underlying cholinergic transmission in particular differs across the two fields (Hutsler and Gazzaniga, 1996;Zilles et al., 2002). This is relevant as the cholinergic system influences cellular excitability (Cox et al., 1994;Hsieh et al., 2000) as well as the stimulus coding capacity of a neural population (Minces et al., 2017;Schmitz and Duncan, 2018), and disrupting it pharmacologically can reduce alpha power (Osipova et al., 2003;Bauer et al., 2012;Eckart et al., 2016). ...
The functional organization of human auditory cortex can be probed by characterizing responses to various classes of sound at different anatomical locations. Along with histological studies this approach has revealed a primary field in posteromedial Heschl's gyrus (HG) with pronounced induced high-frequency (70-150 Hz) activity and short-latency responses that phase-lock to rapid transient sounds. Low-frequency neural oscillations are also relevant to stimulus processing and information flow, however their distribution within auditory cortex has not been established. Alpha activity (7-14 Hz) in particular has been associated with processes that may differentially engage earlier versus later levels of the cortical hierarchy, including functional inhibition and the communication of sensory predictions. These theories derive largely from the study of occipitoparietal sources readily detectable in scalp electroencephalography. To characterize the anatomical basis and functional significance of less accessible temporal-lobe alpha activity we analyzed responses to sentences in seven human adults (four female) with epilepsy who had been implanted with electrodes in superior temporal cortex. In contrast to primary cortex in posteromedial HG, a non-primary field in anterolateral HG was characterized by high spontaneous alpha activity that was strongly suppressed during auditory stimulation. Alpha-power suppression decreased with distance from anterolateral HG throughout superior temporal cortex, and was more pronounced for clear compared to degraded speech. This suppression could not be accounted for solely by a change in the slope of the power spectrum. The differential manifestation and stimulus-sensitivity of alpha oscillations across auditory fields should be accounted for in theories of their generation and function.SIGNIFICANCE STATEMENTTo understand how auditory cortex is organized in support of perception, we recorded from patients implanted with electrodes for clinical reasons. This allowed measurement of activity in brain regions at different levels of sensory processing. Oscillations in the alpha range (7-14 Hz) have been associated with functions including sensory prediction and inhibition of regions handling irrelevant information, but their distribution within auditory cortex is not known. A key finding was that these oscillations dominated in one particular non-primary field, anterolateral Heschl's gyrus, and were suppressed when subjects listened to sentences. These results build on our knowledge of the functional organization of auditory cortex and provide anatomical constraints on theories of the generation and function of alpha oscillations.
... cholinergic transmission in particular differs across the two fields (Hutsler and Gazzaniga, 1996;Zilles et al., 2002). This is relevant as the cholinergic system influences cellular excitability (Cox et al., 1994;Hsieh et al., 2000) as well as the stimulus coding capacity of a neural population (Minces et al., 2017;Schmitz and Duncan, 2018), and disrupting it pharmacologically can reduce alpha power (Osipova et al., 2003;Bauer et al., 2012;Eckart et al., 2016). ...
The functional organization of human auditory cortex can be probed by characterizing responses to various classes of sound at different anatomical locations. Along with histological studies this approach has revealed a primary field in posteromedial Heschl’s gyrus (HG) with pronounced induced high-frequency (70-150 Hz) activity and short-latency responses that phase-lock to rapid transient sounds. Low-frequency neural oscillations are also relevant to stimulus processing and information flow, however their distribution within auditory cortex has not been established. Alpha activity (7-14 Hz) in particular has been associated with processes that may differentially engage earlier versus later levels of the cortical hierarchy, including functional inhibition and the communication of sensory predictions. These theories derive largely from the study of occipitoparietal sources readily detectable in scalp electroencephalography. To characterize the anatomical basis and functional significance of less accessible temporal-lobe alpha activity we analyzed responses to sentences in seven human adults (four female) with epilepsy who had been implanted with electrodes in superior temporal cortex. In contrast to primary cortex in posteromedial HG, a non-primary field in anterolateral HG was characterized by high spontaneous alpha activity that was strongly suppressed during auditory stimulation. Alpha-power suppression decreased with distance from anterolateral HG throughout superior temporal cortex, and was more pronounced for clear compared to degraded speech. This suppression could not be accounted for solely by a change in the slope of the power spectrum. The differential manifestation and stimulus-sensitivity of alpha oscillations across auditory fields should be accounted for in theories of their generation and function.
Significance Statement
To understand how auditory cortex is organized in support of perception, we recorded from patients implanted with electrodes for clinical reasons. This allowed measurement of activity in brain regions at different levels of sensory processing. Oscillations in the alpha range (7-14 Hz) have been associated with functions including sensory prediction and inhibition of regions handling irrelevant information, but their distribution within auditory cortex is not known. A key finding was that these oscillations dominated in one particular non-primary field, anterolateral Heschl’s gyrus, and were suppressed when subjects listened to sentences. These results build on our knowledge of the functional organization of auditory cortex and provide anatomical constraints on theories of the generation and function of alpha oscillations.
... We have recently reported that general cholinergic stimulation with iontophoresis of carbachol excited a greater proportion of DLPFC neurons (49%) than our results with M1R stimulation here, also reducing neuronal rule selectivity (Major et al., 2018). Since ACh actions in cortex are thought to be primarily mediated by muscarinic receptors (Cox et al., 1994), our previous results with carbachol considered with our results with M1R here suggest that other muscarinic receptor subtypes may have a more defining role in activity for rules and task set. M2Rs are also expressed in DLPFC and, in addition to presynaptic expression suggesting a role in modulating neurotransmitter release, are expressed in pyramidal neurons mostly in spines of apical dendrites apposed to glutamatergic synapses (Mrzljak et al., 1993(Mrzljak et al., , 1998. ...
Acetylcholine release in the prefrontal cortex (PFC), acting through muscarinic receptors, has an essential role in regulating flexible behavior and working memory (WM). General muscarinic receptor blockade disrupts PFC WM representations, while selective stimulation of muscarinic receptor subtypes is of great interest for the treatment of cognitive dysfunction in Alzheimer's disease. Here, we tested selective stimulation and blockade of muscarinic M1 receptors (M1Rs) in macaque PFC, during performance of a cognitive control task in which rules maintained in WM specified saccadic responses. We hypothesized that M1R blockade and stimulation would disrupt and enhance rule representation in WM, respectively. Unexpectedly, M1R blockade did not consistently affect PFC neuronal rule selectivity. Moreover, M1R stimulation suppressed PFC activity, and at higher doses, degraded rule representations. Our results suggest that, in primates, the deleterious effects of general muscarinic blockade on PFC WM activity are not mediated by M1Rs, while their overstimulation deteriorates PFC rule maintenance. Vijayraghavan et al. report that muscarinic M1 receptor blockade did not reproduce the effects of general muscarinic antagonism on maintenance of behavioral rules in prefrontal cortical neurons. M1 receptor stimulation consistently inhibited prefrontal neurons and overstimulation disrupted maintenance of rules.
... Cette interprétation est tout à fait en harmonie avec les résultats de différentes études électrophysiologiques réalisées in vivo dans la région du noyau basal [72,112,500,501] qui montrent que certains neurones sont plus actifs pendant l'éveil que pendant le sommeil lent et que ces neurones (présumés cholinergiques) augmentent leur décharge tonique suite à la stimulation de la formation réticulée [501] ou en association avec certains comportements d'éveil [72]. De cette augmentation de décharge tonique des neurones cholinergiques devrait résulter une augmentation de la libération d'acétylcholine corticale telle qu'on l'observe durant l'éveil ou suite à la stimulation (électrique ou chimique) de la formation réticulée ou du noyau basal [76,80,245,258,407,408]. L'acétylcholine ainsi libérée à partir du noyau basal pourrait contribuer à l'activation corticale, soit directement en dépolarisant les neurones corticaux [93,341,524], soit indirectement en inhibant la genèse des fuseaux par une action au niveau du noyau réticulé thalamique [322,386,488]. En imposant ainsi leur influence aux cellules corticales, les neurones cholinergiques du noyau basal pourraient contribuer à la genèse et/ou au maintien d'une activité corticale à haute fréquence (bêta, 18-30 Hz ou gamma, >40 Hz ) telle qu'elle est observée à l'EEG pendant l'éveil naturel ou suite à l'activation du noyau basal [307,341] ou de la formation réticulée mésencéphalique [358]. ...
Le projet de cette thèse a été d'étudier, en combinant immunohistochimie, électrophysiologie et pharmacologie, les propriétés intrinsèques et la neuromodulation des neurones cholinergiques du noyau basal. Ce noyau, localisé dans le télencéphale basal, fournit l'essentiel de l'innervation cholinergique du cortex cérébral et représente donc une composante essentielle des systèmes d'activation corticale.
... Plastische Veränderungen der Neurone im AC können über elektrische Stimulation des NB oder die Applikation cholinerger Agonisten hervorgerufen werden (Metherate und Ashe, 1991;Bjordahl et al., 1998;Kilgard und Merzenich, 1998;Weinberger und Bakin, 1998;Weinberger, 2003;Ma und Suga, 2003). Zudem kommt es zu einer Erhöhung der spontanen Feuerrate, wenn man cholinerge Agonisten in-vitro appliziert (Cox, et al. 1994;Shulz et al., 1997). Die Wirkung des Acetylcholins im Cortex wird hauptsächlich über die muscarinergen M 1 -Rezeptoren vermittelt, die vermutlich die Leitfähigkeit für Kalium in corticalen Neuronen herabsetzen (McCormick und Prince, 1985). ...
... It is worth pointing out here that the neuromodulator acetylcholine can also induce the desynchronized, high frequency cortical states associated with wakefulness and alertness. Cholinergic projections from the nucleus basalis of the basal forebrain are thought to mediate this effect (Buzsaki et al., 1988;Buzsaki and Gage, 1989;Metherate et al., 1992;Cox et al., 1994;Jones, 2008). ...
... During episodic memory, acetylcholine may facilitate glutamate activity by coordinating states of acquisition and recall in the cortex and the HPC (Cox et al., 1994). Cholinergic projections modulates the glutamatergic pathway into the HPC and is responsible for encoding new information into different hippocampal subregions (Yun et al., 2000). ...
... However, the molecular mechanisms by which muscarinic receptors may affect cortical functions are not completely understood. Electrophysiological studies have demon-strated that one important role of cortical muscarinic receptors is the modulation of cell responsiveness to incoming inputs (Cox et al., 1994). Thus, a central issue is to elucidate how in the cortex the muscarinic signaling interacts with and modulates other neurotransmitter stimuli, particularly those involved in learning and memory. ...
: In the present study, we investigated the involvement of βγ subunits of Gq/11 in the muscarinic M1 receptor-induced potentiation of corticotropin-releasing hormone (CRH)-stimulated adenylyl cyclase activity in membranes of rat frontal cortex. Tissue exposure to either one of two βγ scavengers, the QEHA fragment type II adenylyl cyclase and the GDP-bound form of the α subunit of transducin, inhibited the muscarinic M1 facilitatory effect. Moreover, like acetylcholine (ACh), exogenously added βγ subunits of transducin potentiated the CRH-stimulated adenylyl cyclase activity, and this effect was not additive with that elicited by ACh. Western blot analysis indicated the expression in frontal cortex of both type II and type IV adenylyl cyclases, two isoforms stimulated by βγ subunits in synergism with activated Gs. The M1 receptor-induced enhancement of the adenylyl cyclase response to CRH was counteracted by the Gq/11 antagonist GpAnt-2A but not by GpAnt-2, a preferential Gi/o antagonist. In addition, the muscarinic facilitatory effect was inhibited by membrane preincubation with antiserum directed against the C terminus of the α subunit of Gq/11, whereas the same treatment with antiserum against either Gi1/2 or Go was without effect. These data indicate that in membranes of rat frontal cortex, activation of muscarinic M1 receptors potentiates CRH-stimulated adenylyl cyclase activity through βγ subunits of Gq/11.
... The interaction of VX with presynaptic muscarinic receptors is undoubtedly a more specific mechanism by which synaptic transmission can be altered. However, it has been reported that the muscarinic receptor subtypes mostly associated with modulation of glutamate release in the CNS are the M1 and M3 (Cox et al., 1994). Thus, even if VX was capable of interacting with M1/M3 receptors, no signigicant changes in glutamate release would have been detected in our preparation, because these receptor subtypes do not appear to be the predominant muscarinic receptor subtypes present in hippocampal neurons (McKinney et al., 1993). ...
This project was a major effort to determine the effects of low-level exposure to the nerve agents sarin, soman, and VX in the mammalian CNS. Actions on synaptic transmission and neuronal cell death were assessed. Studies gave us clues to the mechanism of action of the agents, particularly in regard to cognitive function in humans that could be exposed to nerve agents in a chemical warfare attack. Reversible cholinesterase (ChE) inhibitors were tested for their ability to counteract nerve agent effects. Galantamine, a reversible ChE inhibitor with nicotinic allosteric potentiating actions currently used to treat mild to moderate Alzheimer's disease, was found to be effective in counteracting nerve agent toxicity. In vitro studies were therefore employed to determine the mechanism of action of galantamine. In vivo studies in guinea pigs were ultimately begun to assess the effectiveness of a pre- and/or post-treatment regimen of galantamine in protecting against multiple LD50 challenges with nerve agents. We compared the effectiveness of pre- and post-treatment with galantamine with other reversible ChE inhibitors in preventing lethality of the nerve agents and examined potential mechanisms underlying the effectiveness of the best treatment.
... The role of ACh in these processes has been analysed at the biophysical and functional levels in different species and cortical sensory areas (McCormick, 1992a). The iontophoretic application of ACh or the stimulation of the basal forebrain result in a muscarinicmediated modulation of firing modes (McCormick and Prince, 1986;Metherate el al., 1992;Cox et al., 1994). Studies of the modulation by ACh of receptive field properties of sensory cortical neurons for different modalities (Sillito and Kemp, 1983;Sat0 et al., 1987;Lamour et al., 1988: Metherate et al., 1988Bassant et al., 1990) generally show a facilitatory influence of muscarinic agonists on cortical neurons, with varying proportions of cells inhibited by ACh. ...
During normal brain operations, cortical neurons are subjected to continuous cholinergic modulations. In vitro studies have indicated that, in addition to affecting general cellular excitability, acetylcholine also modulates synaptic transmission. Whether these cholinergic mechanisms lead to a modulation of functional connectivity in vivo is not yet known. Herein, the effects were studied of an iontophoretic application of acetylcholine and of the muscarinic agonist, carbachol, on the ongoing activity and co-activity of neurons simultaneously recorded in the auditory cortex of the anaesthetized guinea-pig. Iontophoresis of cholinergic agonists mainly affected the spontaneous firing rates of auditory neurons, affected autocorrelations less (in most cases their central peak areas were reduced), and rarely affected cross-correlations. These findings are consistent with cholinergic agonists primarily affecting the excitability of cortical neurons rather than the strength of cortical connections. However, when changes of cross-correlations occurred, they were usually not correlated with concomitant changes in average firing rates nor with changes in autocorrelations, which suggests a secondary cholinergic effect on specific cortico-cortical or thalamo-cortical connections.
... Blocking M-type K + channels with 10 mM XE-991 does not affect the sAHP in IL neurons (Santini and Porter, 2010), therefore muscarine also blocks the calcium-dependent potassium channels underlying the sAHP. Consistent with this, modulation of the sAHP by muscarinic receptors has been previously shown in other brain regions (Cox et al, 1994;Zhang et al, 1996;Bond et al, 1999;Egorov et al, 1999;Scroggs et al, 2001;Krause et al, 2002;Sah and Faber, 2002;Ghamari-Langroudi and Bourque, 2004). ...
There is considerable interest in identifying pharmacological compounds that could be used to facilitate fear extinction. Recently, we showed that the modulation of M-type K(+) channels regulates the intrinsic excitability of infralimbic (IL) neurons and fear expression. As muscarinic acetylcholine receptors inhibit M-type K(+) channels, cholinergic inputs to IL may have an important role in controlling IL excitability and, thereby, fear expression and extinction. To test this model, we combined whole-cell patch-clamp electrophysiology and auditory fear conditioning. In prefrontal brain slices, muscarine enhanced the intrinsic excitability of IL neurons by reducing the M-current and the slow afterhyperpolarization, resulting in an increased number of spikes with shorter inter-spike intervals. Next, we examined the role of endogenous activation of muscarinic receptors in fear extinction. Systemic injected scopolamine (Scop) (muscarinic receptor antagonist) before or immediately after extinction training impaired recall of extinction 24-h later, suggesting that muscarinic receptors are critically involved in consolidation of extinction memory. Similarly, infusion of Scop into IL before extinction training also impaired recall of extinction 24-h later. Finally, we demonstrated that systemic injections of the muscarinic agonist, cevimeline (Cev), given before or immediately after extinction training facilitated recall of extinction the following day. Taken together, these findings suggest that cholinergic inputs to IL have a critical role in modulating consolidation of fear extinction and that muscarinic agonists such as Cev might be useful for facilitating extinction memory in patients suffering from anxiety disorders.
... Muscarinic stimulation can increase postsynaptic membrane resistance, and, thereby, responses to afferent inputs, due to decreased conductance of several K + channels (Halliwell and Adams, 1982; Krnjevic et al., 1971; Madison et al., 1987; McCormick and Prince, 1986). Accordingly, activation of cholinergic synapses in auditory cortex increases postsynaptic excitability via a slow EPSP associated with increased membrane resistance and decreased afterhyperpolarization potentials (AHPs) (Cox et al., 1994; Metherate et al., 1992). Stimulation of cortically-projecting neurons from the nucleus basalis enhances afferent responses in A1 evoked by thalamic (Metherate and Ashe, 1993; Metherate et al., 1992) or acoustic stimulation (Chen and Yan, 2007; Edeline et al., 1994; Zhang et al., 2005), in part by enhancing tone-evoked EPSPs and simultaneously reducing tone-evoked inhibition (Froemke et al., 2007; Metherate and Ashe, 1993; Metherate et al., 1992). ...
Although it is known that primary auditory cortex (A1) contributes to the processing and perception of sound, its precise functions and the underlying mechanisms are not well understood. Recent studies point to a remarkably broad spectral range of largely subthreshold inputs to individual neurons in A1--seemingly encompassing, in some cases, the entire audible spectrum--as evidence for potential, and potentially unique, cortical functions. We have proposed a general mechanism for spectral integration by which information converges on neurons in A1 via a combination of thalamocortical pathways and intracortical long-distance, "horizontal", pathways. Here, this proposal is briefly reviewed and updated with results from multiple laboratories. Since spectral integration in A1 is dynamically regulated, we also show how one regulatory mechanism--modulation by the neurotransmitter acetylcholine (ACh)--could act within the hypothesized framework to alter integration in single neurons. The results of these studies promote a cellular understanding of information processing in A1.
... studies conducted in brain slices or in cultured cells showed much more heterogeneous and not univocal results. for instance, cox et al. (30) using single cell recordings from in vitro auditory cortex observed an increase of post-synaptic response elicited in neocortical neurons by puff application of glutamate in the presence of ach; the authors showed that this enhancement was mediated by machrs. a similar result was obtained by calabresi et al. (17) in the striatum showing selective nmda enhancement after muscarine application. ...
This review is focused on the basal forebrain (BFB) cholinergic system, cholinergic receptors and cholinoceptive target areas such as the neocortex, all of which are intimately involved in high cognitive functions and synaptic plasticity. The neurons of the BFB synthesize acetylcholine (ACh) whose action is mediated by two subclasses of receptors, namely nicotinic and muscarinic receptors. Using the visual system as a model, the aim here is to integrate and discuss the current knowledge on anatomy, ontogeny and function of the BFB cholinergic system. This signaling system represents the anatomo-functional basis of ACh action on neuronal network, neuronal plasticity and cognitive functions. Cholinergic system role on higher brain functions has received increasing attention since the first observation of A. Alzheimer (1907) reporting dramatic changes of the BFB cholinergic neuro-anatomy in one of the most devastating neUrodegenerative diseases of adult brain, i.e. Alzheirner's disease (AD). In addition to this observation, later work demonstrated its participation in deep re-arrangements of brain connectivity such as the regulation of neuronal plasticity during maturation of cortical sensory maps, in adult and aged brain.
... The role of ACh in these processes has been analysed at the biophysical and functional levels in different species and cortical sensory areas (McCormick, 1992a). The iontophoretic application of ACh or the stimulation of the basal forebrain result in a muscarinicmediated modulation of firing modes (McCormick and Prince, 1986; Metherate el al., 1992; Cox et al., 1994). Studies of the modulation by ACh of receptive field properties of sensory cortical neurons for different modalities (Sillito and Kemp, 1983; Sat0 et al., 1987; Lamour et al., 1988: Metherate et al., 1988 Bassant et al., 1990) generally show a facilitatory influence of muscarinic agonists on cortical neurons, with varying proportions of cells inhibited by ACh. ...
During normal brain operations, cortical neurons are subjected to continuous cholinergic modulations. In vitro studies have indicated that, in addition to affecting general cellular excitability, acetylcholine also modulates synaptic transmission. Whether these cholinergic mechanisms lead to a modulation of functional connectivity in vivo is not yet known. Herein, the effects were studied of an iontophoretic application of acetylcholine and of the muscarinic agonist, carbachol, on the ongoing activity and co-activity of neurons simultaneously recorded in the auditory cortex of the anaesthetized guinea-pig. Iontophoresis of cholinergic agonists mainly affected the spontaneous firing rates of auditory neurons, affected autocorrelations less (in most cases their central peak areas were reduced), and rarely affected cross-correlations. These findings are consistent with cholinergic agonists primarily affecting the excitability of cortical neurons rather than the strength of cortical connections. However, when changes of cross-correlations occurred, they were usually not correlated with concomitant changes in average firing rates nor with changes in autocorrelations, which suggests a secondary cholinergic effect on specific cortico-cortical or thalamo-cortical connections.
... Appealing candidates are cholinergic receptors in cortex, because transient impairment of cholinergic function in the insular cortex disrupts the encoding of information on novel tastes (Naor and Dudai, 1996), and microinjection of carbachol into the insular cortex enhances tyrosine phosphorylation of NR2B ( Rosenblum et al., 1996a). A plausible model is thus that activation of the cholinergic system by contextual saliency leads to acetylcholine receptor-induced increase in Ca 2 influx in cortical neurons (Cox et al., 1994;Lev et al., 1995), resulting in activation of protein kinase(s) such as members of the src family mentioned above ( Kohr and Seeburg, 1996) or the fak family ( Lev et al., 1995), and culminating in enhanced phosphorylation of a set of substrates, including NR2B. This might be a molecular manifestation of the cross-talk of the glutamatergic and cholinergic systems in cortex, a cross-talk that is proposed to play a prominent function in learning and attention (Aigner, 1995). ...
We demonstrate that the NMDA receptor is involved in taste learning in the insular cortex of the behaving rat and describe two facets of this involvement. Blockage of the NMDA receptor in the insular cortex by the reversible antagonist APV during training in a conditioned taste aversion (CTA) paradigm impaired CTA memory, whereas blockage of the NMDA receptor in an adjacent cortex or before a retrieval test had no effect. When rats sampled an unfamiliar taste and hence learned about it, either incidentally or in the context of CTA training, the tyrosine phosphorylation of the NMDA receptor subunit 2B (NR2B) in the insular cortex was specifically increased. The level of tyrosine phosphorylation on NR2B was a function of the novelty of the taste stimulus and the quantity of the taste substance consumed, properties that also determined the efficacy of the taste stimulus as a conditioned stimulus in CTA; however, blockage of the NMDA receptor by APV during training did not prevent tyrosine phosphorylation of NR2B. We suggest that tyrosine phosphorylation of NR2B subserves encoding of saliency in the insular cortex during the first hours after an unfamiliar taste is sampled and that this encoding is independent of another, necessary role of NMDA receptors in triggering experience-dependent modifications in the insular cortex during taste learning. Because a substantial fraction of the NR2B protein in the insular cortex seems to be expressed in interneurons, saliency and the tyrosine phosphorylation of NR2B correlated with it may modulate inhibition in cortex.
Kainate receptors (KARs), together with AMPA and NMDA, are typically described as ionotropic glutamate receptors. The functions of KARs have begun to be elucidated only in the last decade. While some the actions of KARs are classically ionotropic, surprisingly, others seem to involve the activation of second messenger cascades and invoke metabotropic roles for this type of glutamate receptor. In this chapter, we describe these metabotropic actions of KARs in relation to the putative signalling cascades involved. Although, it is still a mystery how KARs activate G-proteins to stimulate second messenger cascades, intriguingly in very recent studies, specific subunits of KARs have been demonstrated to associate with G-proteins. Altogether, the body of evidence supports the hypothesis that, together with the canonical ionotropic operation, KARs expedite long-lasting signalling by novel metabotropic modes of action. © 2011 Nova Science Publishers, Inc. All rights reserved.
This volume critically examines the functional actions of the kainate‑type glutamate receptors (KARs). Following on from the larger body of work on the NMDA‑ and AMPA-type ionotropic glutamate receptors (GluRs), studies with KARs have consistently thrown up exceptions to general rules about synaptic modulation. Contributors herein provide an insight to the idiosyncracies that now almost typify the KAR field. The fascinating insights provided in this volume serve to encourage searching mechanistic questions.
Learning to associate a stimulus with reinforcement causes plasticity in primary sensory cortex. Neural activity caused by the associated stimulus is paired with reinforcement, but population analyses have not found a selective increase in response to that stimulus. Responses to other stimuli increase as much as, or more than, responses to the associated stimulus. Here, we applied population analysis at a new time point, and additionally evaluated whether cholinergic receptor blockers interacted with the plastic changes in cortex. Three days of tone identification behavior caused responsiveness to increase broadly across primary auditory cortex, and target responses strengthened less than overall responsiveness. In pharmacology studies, behaviorally impairing doses of selective acetylcholine receptor blockers were administered during behavior. Neural responses were evaluated on the following day while the blockers were absent. The muscarinic group, blocked by scopolamine, showed lower responsiveness, and an increased response to the tone identification target that exceeded both the three day control group and task-naïve controls. Also, a selective increase in the late phase of the response to the tone identification stimulus emerged. Nicotinic receptor antagonism, with mecamylamine, more modestly lowered responses the following day, and lowered target responses more than overall responses. Control acute studies demonstrated the muscarinic block did not acutely alter response rates, but the nicotinic block did. These results lead to the hypothesis that the decrease in the proportion of the population spiking response that is selective for the target may be explained by the balance between effects modulated by muscarinic and nicotinic receptors. This article is protected by copyright. All rights reserved.
NMRI mice were trained in a one-trial inhibitory avoidance task. They were injected immediately after training with the muscarinic cholinergic agonist oxotremorine, the muscarinic cholinergic antagonist atropine, the N-methyl- D-aspartate (NMDA) noncompetitive antagonist MK-801, or with a combination of MK-801 and one of the cholinergic agents. Oxotremorine improved, while atropine and MK-801 impaired, memory retention. In addition, oxotremorine attenuated, while atropine enhanced, the effect of MK-801. The results show the existence of a glutamatergic-cholinergic interaction in modulating memory consolidation of mice.
The phase response curve (PRC) reflects the dynamics of the interplay between diverse intrinsic conductances that lead to spike generation. PRCs measure the spike time shift caused by perturbations of the membrane potential as a function of the phase of the spike cycle of a neuron. A purely positive PRC is a signature of type I (saddle-node) dynamics while type II (subcritical Hopf dynamics) yield a biphasic PRC with both negative and positive lobes. Previous computational work hypothesized that cholinergic modulation of M-type potassium current can switch a neuron with type II dynamics to type I dynamics. We recorded from layer 2/3 pyramidal neurons in cortical slices, and found that cholinergic action, consistent with downregulation of slow voltage-dependent potassium currents such as the M-current, indeed changed the PRC from type II to type I. We then explored the potential specific K-current-dependent mechanisms for this switch using a series of computational models. In all of these models, we show that a decrease in spike-frequency adaptation due to downregulation of the M-current is associated with the switch in PRC type. Interestingly spike-dependent I-AHP is downregulated at lower Ach concentrations than the M-current. Our simulations showed that type II nature of the PRC is amplified by low Ach level, while the PRC became type I at high Ach concentrations. We further explored the spatial aspects of Ach modulation in a compartmental model. This work suggests that cholinergic modulation of slow potassium currents may shape neuronal responding between “resonator” to “integrator.”
Several theories consider the brain to be a network of neurons that process perception with simple activation functions. Real neurons, however, are far more intricate.Through reviews of literature and results from original experiments, Neural Cell Behavior and Fuzzy Logic offers a comprehensive look at these complex systems, supplying trustworthy evidence that neurons can predict the consequences of input signals and transiently change their own excitability to suit. The book also examines how fuzzy logic, the computing of perceptions, can be used to provide a theoretical description of real neuron behavior, and as a model for the "logic" the brain uses to describe environments and make decisions. This book includes sections for general and advanced readers, and will be particularly useful to neuroscience students, academics and researchers as well as to mathematicians and theoretical physicists. © 2008 Springer Science+Business Media, LLC. All rights reserved.
This chapter reviews cholinergic anatomy, neurochemistry, and physiology related to frontal lobe function. The chapter discusses the contribution of cholinergic dysfunction to Alzheimer's disease (AD) and other disorders associated with cognitive deterioration, as well as the use of drugs to enhance cholinergic activity under these conditions. Finally, imaging strategies, such as functional MRI (fMRI) and positron emission tomography (PET), which are playing an increasingly important role in investigations of the human ACh system, are reviewed. These techniques can be used to investigate effects of cholinergic agonists and antagonists on cognitive performance in healthy and compromised individuals and can assist in disease detection and monitoring of progression, treatment, and clinical outcome. Throughout the chapter, important findings related to the brain cholinergic system are presented and directions for future research are also highlighted.
Previously, we reported that activation of muscarinic receptors modulates N-methyl-D-aspartate (NMDA) receptor-mediated synaptic transmission in auditory neocortex [Aramakis et al. (1997a) Exp Brain Res 113:484–496]. Here, we describe the muscarinic subtypes responsible for these modulatory effects, and a role for G-proteins and intracellular messengers. The muscarinic agonist oxotremorine-M (oxo-M), at 25–100 μM, produced a long-lasting enhancement of NMDA-induced membrane depolarizations. We examined the postsynaptic G-protein dependence of the modulatory effects of oxo-M with the use of the G-protein activator GTPγS and the nonhydrolyzable GDP analog GDPβS. Intracellular infusion of GTPγS mimicked the facilitating actions of oxo-M. After obtaining the whole-cell recording configuration, there was a gradual, time-dependent increase of the NMDA receptor-mediated slow-EPSP, and of iontophoretic NMDA-induced membrane depolarizations. In contrast, intracellular infusion of either GDPβS or the IP3 receptor antagonist heparin prevented oxo-M mediated enhancement of NMDA depolarizations. The muscarinic receptor involved in enhancement of NMDA iontophoretic responses is likely the M1 receptor, because the increase was prevented by pirenzepine, but not the M2 antagonists methoctramine or AF-DX 116. Oxo-M also reduced the amplitude of the pharmacologically isolated slow-EPSP, and this effect was blocked by M2 antagonists. Thus, muscarinic-mediated enhancement of NMDA responses involves activation of M1 receptors, leading to the engagement of a postsynaptic G-protein and subsequent IP3 receptor activity. Synapse 32:262–275, 1999. © 1999 Wiley-Liss, Inc.
Attention involves several functions such as alertness, shift, stabilization, and distractor suppression. Alertness is a global mental state characterized by increased motivation and lowered thresholds for encoding new information. Attention shift allows either transiting from a passive inattentive to an active focused state, or refocusing perceptual resources from a previously targeted perceptual object to a more salient one. During attention stabilization perceptual resources are kept concentrated onto a particular target in a manner that the neural activity evoked by selected stimuli is momentarily enhanced. Suppression is an active process by which neural activity evoked by task irrelevant stimuli is diminished. These functions are impaired in several neuropsychiatric conditions. We review clinical and neurophysiological data in humans and laboratory animals suggesting that acetylcholine and dopamine interact in the neocortex to produce purposeful attention.
It is proposed that a more satisfactory theory of attention needs to integrate both tonic and phasic effects produced by acetylcholine and dopamine. In the model here proposed, nicotinic receptors are thought to play a pivotal role in the enhancement of neural activity evoked by task relevant stimuli. Muscarinic receptors are proposed to be involved in alertness, and dopaminergic receptors in the temporary representation of intermediate goals. A combination of signals triggered by muscarinic and dopaminergic receptor coactivation may facilitate the suppression of neural activity evoked by task irrelevant stimuli. A better understanding of the interplay between dopamine and acetylcholine in attention modulation may help to develop better psychopharmacological interventions for neuropsychiatric conditions in which attention is impaired.
What does the auditory cortex do? Most would agree that it processes auditory information, but few would assert that we understand
just what computations are performed by auditory cortical neurons. If we describe computation as the transformation of information
from one representation to another, then which transformations are accomplished by the auditory cortex remains an open question
at the heart of the discipline.
Both acetylcholine and glutamate are now thought to play important roles in memory. Recent evidence suggests that the interaction of these two neurotransmitters may be important for some forms of memory, and that acetylcholine, in particular, may function to facilitate glutamate activity by coordinating states of acquisition and recall in the cortex and hippocampus.
Kainate receptors (KAR) form a class of glutamate receptors that have been implicated in epilepsy, stroke, Alzheimer's and neuropathic pain.1 KAR subtypes are known to be segregated to specific locations within neurons and play significant roles in synaptic transmission and plasticity.2 Increasing evidence suggests a the role for ubiqutination in regulating the number of synaptic neurotransmitter receptors.3-5 The ubiquitin pathway consists of activation (E1), conjugation (E2) and ligation (E3). Cullins form the largest family of E3 ligase complexes. We have recently shown that the BTB/Kelch domain proteins, actinfilin and mayven, bind both Cul3 and specific KAR subtypes (GluR6 and GluR5-2b) to target these KARs for ubiquitination and degradation.5 In this chapter we will review how these interactions occur, what they mean for the stability of KARs and their associated proteins and how, in turn, they may affect synaptic functions in the central nervous system.
Most of our knowledge of the synaptic function of kainate receptors stems from a detailed analysis of synaptic transmission between dentate granule cells and CA3 pyramidal neurons, where kainate receptors mediate a slow excitatory current with integrative properties ideally suited for repetitive neuronal firing. Besides this well characterized ionotropic effect of kainate receptors, they can also enhance neuronal excitability by inhibiting the slow Ca(2+) activated K(+) current I(sAHP) via a G-protein coupled mechanism. This phenomenon is associated with Ca(2+) mobilization and protein-kinase activation and ultimately leads to modulation of ion channels responsible for intrinsic electrical properties such as firing adaptation. The significance for CNS function of these newly emerging metabotropic kainate receptors is poorly understood and as yet proteomic analysis of kainate receptors has yielded little information on signaling molecules associated with the kainate receptor ionophore. This chapter covers the key findings that have led to the proposal that high-affinity postsynaptic kainate receptors trigger a form of metabotropic signaling regulating I(sAH P) and neuronal firing in CA3 hippocampal neurons.
Kainate receptors (KARs) are one of the three subtypes of ionotropic glutamate receptors in the CNS. These receptors are widely expressed pre- and postsynaptically throughout the brain. Thus, kainate receptor activation mediates a large variety of pre- and postsynaptic effects on either glutamatergic or GABAergic synaptic transmission. Although ionotropic functions for KAR have been described in multiple brain regions, there is considerable evidence from various CNS regions that KARs activation modulates GABA release through either G-protein dependent metabotropic pathway or secondary activation of G-protein coupled receptors. In the present chapter, we provide further evidence supporting that these two pathways are also involved in the modulation of GABA release in specific basal ganglia nuclei. Because of their more subtle effects on neurotransmisison regulation than other ionotropic glutamate receptors, KARs represent interesting targets for the future development of pharmacotherapy for basal ganglia diseases.
Kainate receptors (KARs) are members of the family of ionotropic glutamate receptors (iGluRs) which also include NMDA and AMPA receptors. As ionotropic receptors, KARs have been characterized, pre and postsynaptically, in several brain regions. In this chapter we review evidence that suggests that KARs mediate some of their effects without invoking ion-fluxes. Beginning with seminal experiments described some ten years ago, when the notion of a metabotropic action of KAR was first posited in the modulation of GABA release from hippocampal interneurons, increasingly, there have been reports indicating that some KAR functions overtly depend on G-protein activation and involve the participation of intracellular signalling cascades. Thus, KAR activation instigates a cascade involving G(i/o), phospholipase C and protein kinase C to suppress the release of GABA and therefore underpins disinhibition of pyramidal cells in the CA1 region of the hippocampus. This type of metabotropic function of KARs in controlling GABA release represents an additional level of activity-dependent control of synaptic inhibition which is independent of any ionotropic activity of KARs.
Neuronal oscillations are thought to underlie interactions between distinct brain regions required for normal memory functioning. This study aimed at elucidating the neuronal basis of memory abnormalities in neurodegenerative disorders. Magnetoencephalography (MEG) was used to measure oscillatory brain signals in patients with Alzheimer s disease (AD), a neurodegenerative disease causing progressive cognitive decline, and mild cognitive impairment (MCI), a disorder characterized by mild but clinically significant complaints of memory loss without apparent impairment in other cognitive domains. Furthermore, to help interpret our AD/MCI results and to develop more powerful oscillatory MEG paradigms for clinical memory studies, oscillatory neuronal activity underlying declarative memory, the function which is afflicted first in both AD and MCI, was investigated in a group of healthy subjects. An increased temporal-lobe contribution coinciding with parieto-occipital deficits in oscillatory activity was observed in AD patients: sources in the 6 12.5 Hz range were significantly stronger in the parieto-occipital and significantly weaker in the right temporal region in AD patients, as compared to MCI patients and healthy elderly subjects. Further, the auditory steady-state response, thought to represent both evoked and induced activity, was enhanced in AD patients, as compared to controls, possibly reflecting decreased inhibition in auditory processing and deficits in adaptation to repetitive stimulation with low relevance. Finally, the methodological study revealed that successful declarative encoding and retrieval is associated with increases in occipital gamma and right hemisphere theta power in healthy unmedicated subjects. This result suggests that investigation of neuronal oscillations during cognitive performance could potentially be used to investigate declarative memory deficits in AD patients. Taken together, the present results provide an insight on the role of brain oscillatory activity in memory function and memory disorders. Muistin toiminta edellyttää eri aivoalueiden joustavaa yhteistoimintaa. Hermosolujen oskillatorinen toiminta saattaa olla keskeinen tekijä näiden vuorovaikutusten taustalla. Tämän tutkimuksen tarkoituksena oli selvittää Alzheimerin taudin ja sitä edeltävän lievän kognitiivisen heikentymisen (mild cognitive impairment, MCI) taustalla olevia aivotoiminnan häiriöitä. Alzheimerin tauti on yleisin hermostoa rappeuttava degeneratiivinen sairaus, joka aiheuttaa etenevää kognitiivisen suorituskyvyn heikentymistä. MCI on puolestaan nimitys Alzheimerin tautia usein edeltävälle oireyhtymälle, johon liittyy lievää mutta kliinisesti merkittävää muistitoimintojen heikentymistä. MCI-potilailla on todettu kohonnut riski sairastua Alzheimerin tautiin. Aivotoiminnan oskillatorista toimintaa mitattiin magnetoenkefalografialla (MEG), joka välittää tietoa aivokuoren hermosolujen toiminnan aiheuttamista heikoista sähkömagneettisista muutoksista. Tutkimuksessa verrattiin aivojen oskillatorisen toiminnan piirteitä Alzheimerin taudista ja MCI:stä kärsivien potilaiden sekä terveiden koehenkilöiden välillä. Lisäksi tutkimuksessa selvitettiin muistitoimintojen hermostollista taustaa mittaamalla MEG-oskillaatioita terveiltä nuorilta koehenkilöiltä. Tulosten perusteella aivojen oskillatorisen toiminnan synnyttävä aivokuoren hermosoluverkosto on muuttunut Alzheimerin taudissa. Terveillä koehenkilöillä ja MCI-potilailla niin sanottu alfa-aktivaatio (8 12 Hz) syntyy päälaenlohkon ja takaraivolohkon alueilla, mutta Alzheimer-potilailla nämä oskillaatiot näyttivät syntyvän ohimolohkon alueilla. Toisaalta Alzheimer-potilailla havaittiin toistuvaan ääniärsykkeeseen vaihelukitun värähtelytoiminnan kiihtymistä, mikä saattaa liittyä puutteelliseen kuuloaivokuoren hermosolujen adaptaatioon. Nuorilla terveillä koehenkilöillä suoritetun menetelmällisen kokeen perusteella muistisuoriutuminen lisää takaraivolohkon korkeataajuuksista (60 90Hz) gamma-aktivaatiota ja oikean aivopuoliskon hidasta (4 8 Hz) theta-aktivaatiota. Tässä menetelmällisessä tutkimuksessa kehitetty asetelma saattaa osoittautua tulevaisuudessa merkitykselliseksi muistihäiriöiden kliinisen tutkimukselle. Kokonaisuudessaan tämän tutkimuksen tulokset osoittavat aivokuoren hermosolujen oskillatorisen toiminnan keskeisen merkityksen muistin toiminnan ja sen kliinisten häiriöiden taustalla.
Using electrophysiological techniques in the in vitro rat auditory cortex, we have examined how spontaneous acetylcholine (ACh) release modifies synaptic potentials mediated by glutamate and γ-aminobutyric acid (GABA). Single stimulus pulses to lower layer VI elicited in layer III a four-component (A-D) extracellular field response involving synaptic potentials mediated by glutamate and GABA. The cholinesterase inhibitor eserine (10–20 μM) or the cholinergic agonist carbachol (25–50 μM) depressed by 10–50% the glutamatergic components A and C, and the GABAergic components B and D. Atropine reversed the depressive effects of eserine and carbachol. A novel finding was that the degree of depression of component A varied inversely with stimulus intensity. However, during partial pharmacological antagonism of GABAA receptors, depression of A varied directly, not inversely, with stimulus intensity. Normally, then, depression of A is offset by reduced GABAergic inhibition of A. We also tested for differential depression of responses mediated by N-methyl-d-aspartate (NMDA) versus non-NMDA glutamate receptors. Following physiological and pharmacological isolation of the responses, eserine depressed the non-NMDA, but not the NMDA, receptor-mediated potential. Since the isolated NMDA potential still could be depressed by carbachol, the data suggested that activation of NMDA receptors may reduce spontaneous ACh release. In support of this, preincubation of slices in NMDA (10–20 μM) largely prevented eserine's, but not carbachol's, depression of components A and B.
These results permit three conclusions of relevance to cortical information processing: (1) spontaneous ACh release tonically depresses synaptic potentials mediated by glutamate and GABA; (2) ACh depresses responses to weak inputs to a greater degree than responses to strong inputs; (3) activation of NMDA receptors may “feed-back” to reduce ACh release, a mechanism that could place regulation of local ACh release under glutamatergic afferent control.
Coronal and horizontal slices of the lumbar and sacral spinal cord, respectively, of ovariectomized adult rats, either treated with estrogen (OVX+E) or untreated (OVX), were used to test the neuronal actions of TRH and its metabolite, cyclo(His-Pro) (or cHP). Both coronal slices, which possess only short stumps of ventral roots (VRs), and horizontal slices, in which long sections of VRs were preserved, were used for extracellular recording of single motor and other types of neurons. Methodological comparisons between these two types of slices showed that the length of VRs preserved had no significant effect on the characteristics of motoneurons (MNs). In coronal slices, MNs in medial and lateral lamina IX (MNM and MNL, respectively) were identified by antidromic activation. Of these lumbar MNs, estrogen treatment lowered the antidromic activation threshold for MNM but not MNL. Because MNM innervate the back muscles crucial for the execution of the estrogen-dependent lordosis, the observed estrogen effect may contribute to the hormone's induction of the sexual behavior. The recorded MNs and other types of neurons were subjected to bath applications of TRH, cHP, and neurotransmitters. TRH was found to be capable of evoking an early, shorter-lasting neuronal excitation and/or a late, longer-lasting modulation of neuronal responses to transmitters. Each neuronal action could occur with or without the other, and the occurrence of the excitation did not affect the probability of whether a modulation would occur later. The modulatory, but not the excitatory, action appeared to be shared by cHP, because cHP could also modulate neuronal responses in similar, if not identical, ways as TRH did, but could neither stimulate neurons nor mimic TRH in desensitizing TRH-evoked excitation. The modulatory actions of the two peptides were not affected by estrogen. Although the excitatory action was desensitized by repeated TRH applications, the modulatory action did not appear to be attenuated but instead was often enhanced by repeated administrations of TRH and/or cHP. These results, together with the essentially identical findings from our previous study on hypothalamic neurons, indicate that the excitatory and the modulatory actions of TRH are independent of each other and, hence, are mediated by different subcellular mechanisms.
The effects of acetylcholine (ACh) in processing acoustical information in the inferior colliculus (IC) of awake horseshoe bats (Rhinolophus rouxi) were examined with single cell recordings and microiontophoresis. Cholinergic agonists, acetylcholine and carbachol raised the stimulus evoked discharge in 37% and suppressed responses in 16% of the sample. They did not alter the shapes of tuning curves and rate-intensity functions but the latter showed parallel shifting. The nicotinic antagonist, hexamethonium raised neuronal activity in 52% of neurons without affecting discharge patterns. The nonspecific muscarinic antagonist atropine was mostly inhibitory (62% of units) and caused changes in temporal discharge patterns by affecting the tonic response component. The selective muscarinic ml antagonist pirenzepine, also had an inhibitory effect (37% of units) and predominantly influenced the tonic response component. The selective m2 antagonist, gallamine however produced mainly excitatory effects (64% of units) and changed temporal discharge patterns by adding tonic response components. These findings may indicate a differential pre- and postsynaptic synaptic distribution of m1/m2 receptors in the inferior colliculus as reported for other brain structures. The results indicate that ACh plays a neuromodulatory transmitter role in the auditory midbrain by setting the level of neuronal activity. Its exact function in particular behavioral contexts remains to be determined, since the origin of cholinergic innervation of the mammalian IC is still unclear.
We have recently shown that in the gustatory cortex of the rat, taste learning enhances protein tyrosine phosphorylation and taste memory is blocked by muscarinic antagonists. A major protein whose tyrosine phosphorylation is stimulated by taste learning in cortex is a 180 kDa synaptic glycoprotein identified as the NMDA receptor subunit 2B (NR2B). Here we report that microinjection of carbachol into the taste cortex modulates protein tyrosine phosphorylation similarly to the effect of unfamiliar taste, and that a 180 kDa protein whose tyrosine phosphorylation is enhanced in vivo by carbachol is NR2B. These data, combined with our previous findings, are in line with the hypothesis that muscarinic input plays a role in encoding new items in memory, and that tyrosine phosphorylation of NR2B is involved in this process.
The muscarinic antagonist scopolamine blocks conditioned taste aversion (CTA) when microinjected bilaterally into the rat insular cortex shortly before the exposure of the rat to a novel taste (the conditioned stimulus, CS) in CTA training. Scopolamine has no effect when microinjected shortly after the exposure to the novel taste or shortly before the application of the malaise-inducing agent (unconditioned stimulus, UCS). Scopolamine does not affect sensory, motor and retrieval mechanisms required for performing the CTA task, and does not block CTA when injected into another cortical area. The effect of scopolamine is independent of the taste used as CS. Furthermore, microinjection of scopolamine into the insular cortex shortly before the pre-exposure to a new taste in a latent inhibition paradigm, impairs the attenuation of CTA by that pre-exposure. Other muscarinic antagonists, pirenzepine and AF DX-116, have an effect similar to that of scopolamine. Comparison of the dose-dependency curves of the muscarinic antagonists suggests a predominant role in CTA for M2 subtype receptors. Carbachol, a muscarinic agonist, also impairs the encoding of taste in the insular cortex, but the results are confounded by the ability of that ligand to induce seizures. Our findings suggest that cholinergic neuromodulation participates in processing the CS in the gustatory cortex in CTA, either by encoding novelty at the cellular level, or by instructing the neural circuits to store the novel taste representation.
We investigated the effect of nicotine, a nicotinic agonist, and D-cycloserine (DCS, a partial glycine-B agonist of the N-methyl-D-aspartate (NMDA) receptor complex) on aging-induced defects of water maze (WM) spatial navigation in rats. Nicotine (0.3 mg kg-1, s.c.) or DCS (10 mg kg-1, i.p.) enhanced acquisition of the WM task. A combination of subthreshold doses of nicotine (0.1 mg kg-1) and DCS (3 mg kg-1) improved WM acquisition. A subthreshold dose of a competitive NMDA antagonist, CPP (1 mg kg-1), blocked the effect of nicotine (0.3 mg kg-1) on WM acquisition. A nicotine antagonist, mecamylamine (10 mg kg-1), impaired WM acquisition and had no effect on retention, but did not block the effect of DCS 10 mg kg-1. The results suggest that nicotine and DCS synergistically enhance spatial navigation in aged rats.
Neurons in layer II of the entorhinal cortex (EC) are key elements in the temporal lobe memory system because they integrate and transfer into the hippocampal formation convergent sensory input from the entire cortical mantle. EC layer II also receives a profuse cholinergic innervation from the basal forebrain that promotes oscillatory dynamics in the EC network and may also implement memory function. To understand the cellular basis of cholinergic actions in EC, we investigated by intracellular recording in an in vitro rat brain slice preparation the muscarinic modulation of the electroresponsive properties of the two distinct classes of medial EC layer II projection neurons, the stellate cells (SCs) and non-SCs. In both SCs and non-SCs, muscarinic receptor activation with carbachol (CCh, 10-50 microM) caused atropine-sensitive (300 nM) membrane depolarization. In SCs, the CCh-induced membrane depolarization was associated with subthreshold membrane potential oscillations and "spike cluster" discharge, which are typically expressed by these cells on depolarization. CCh, however, caused a decrease of the dominant frequency of the membrane potential oscillations from 9.2 +/- 1.1 (SD) Hz to 6.3 +/- 1.1 Hz, as well as a decrease of the intracluster firing frequency from 18.1 +/- 1.7 Hz to 13.6 +/- 1.3 Hz. In addition, spike cluster discharge was less robust, and the cells tended to shift into tonic firing during CCh. In contrast to SCs, in non-SCs, CCh drastically affected firing behavior by promoting the development of voltage-dependent, long-duration (1-5 s) slow bursts of action potentials that could repeat rhythmically at slow frequencies (0.2-0.5 Hz). Concomitantly, the slow afterhyperpolarization (sAHP) was replaced by long-lasting plateau postdepolarizations. In both SCs and non-SCs, CCh also produced conspicuous changes on the action potential waveform and its afterpotentials. Notably, CCh significantly decreased spike amplitude and rate of rise, which suggests muscarinic modulation of a voltage-dependent Na+ conductance. Finally, we also observed that whereas CCh abolished the sAHP in both SCs and non-SCs, the membrane-permeant analogues of adenosine 3',5'-cyclic monophosphate, 8-(4-chlorophenylthio)-adenosine-cyclic monophosphate and 8-bromo-adenosine-cyclic-monophosphate, abolished the sAHP in SCs but not in non-SCs. The data demonstrate that cholinergic modulation further differentiates the intrinsic electroresponsiveness of SCs and non-SCs, and add support to the presence of two parallel processing systems in medial EC layer II that could thereby differentially influence their hippocampal targets. The results also indicate an important role for the cholinergic system in tuning the oscillatory dynamics of entorhinal neurons.
The present study is concerned with the ability of muscarinic actions of acetylcholine (ACh) to modulate glutamate and gamma-aminobutyric acid (GABA)-mediated synaptic transmission in the in vitro rat auditory cortex. Whole-cell patch clamp recordings were obtained from layer II-III pyramidal neurons, and the fast-EPSP (AMPA/kainate), fast-IPSP (GABA(A)), and slow-IPSP (GABA(B)), were elicited following a stimulus to deep gray/white matter. Acetyl-beta-methylcholine (MCh), a muscarinic receptor agonist, applied by either superfusion or iontophoresis, produced an atropine-sensitive increase or decrease in the amplitude of the fast-EPSP. The effect of MCh could be predicted by the response of the fast-EPSP to paired-pulse stimulation (i.e. a conditioning pulse followed 300 ms later by a test pulse). The fast-EPSP was decreased in amplitude by MCh in cases where the test-EPSP was suppressed in the pre-MCh condition, and increased in amplitude when the test-EPSP was facilitated. The fast- and slow-IPSPs were always reduced by MCh. In several experiments, the strength of synaptic inhibition was systematically modified by varying stimulus intensity. When the fast-EPSP was elicited in the absence of IPSPs, it was decreased in amplitude by MCh. However, when the fast-EPSP was elicited in conjunction with large IPSPs it was increased in amplitude during MCh. Because the magnitude of the fast-EPSP is influenced by the degree of temporal overlap with IPSPs, it was hypothesized that enhancement of the fast-EPSP was the result of disinhibition produced as a consequence of muscarinic reduction of GABAergic IPSPs. This view was supported by the finding that MCh could reduce the amplitude of pharmacologically isolated GABAergic IPSPs (i.e. elicited in the absence of glutamatergic transmission). Our results suggest that ACh at muscarinic receptors can modify fast glutamatergic neurotransmission differently as a function of strength of inhibition, to suppress that produced by 'weak' inputs and enhance that produced by 'strong' inputs.
Protein kinase C is an important intracellular signaling molecule. Many of its ten isoforms are highly expressed in brain, and protein kinase C has been implicated in the regulation of the activity of receptors of several major neurotransmitters, including glutamate, acetylcholine, glycine, and gamma-aminobutyric acid. These neurotransmitters and their receptors are present in central auditory pathways, suggesting their role in auditory signal processing. Although they may be important modulators of the function of these neurotransmitter receptors, the distribution of protein kinase C isoforms in central auditory systems has not been well characterized. By using immunocytochemistry with specific antibodies, we studied the distribution of immunoreactivity of four isoforms of protein kinase C, betaI, betaII, gamma, and gamma, in central auditory systems of rat brain. Each of these protein kinase C isoforms was found to have a unique distribution in the auditory brainstem and cortex, supporting a role for these isoforms of protein kinase C in different aspects of auditory sensory processing.
EEG and single-unit techniques have been used to study the EEG correlates of cellular firing in the neocortex, n. reticularis (RT) and "specific" thalamic nuclei, and the cholinergic forebrain area (nucleus basalis, NB). Neuronal firing was related to the ongoing behavior of the rat. In addition, using a 16-channel neocortical recording/mapping system, we studied the effects of ibotenic acid lesion of NB, RT, and other thalamic nuclei on the patterns and spatial distribution of neocortical electrical activity. The majority of neurons in neocortex, NB, and RT increased their firing rates during walking, as compared to during immobility, with concurrent decrease of delta power in the neocortical EEG. During immobility, high-voltage spindles (HVS; greater than 1 mV) were occasionally recorded from the neocortex. Depth profiles of HVS and slow delta waves were different in the neocortex. Neocortical cells decreased their discharge frequency during the positive portion of delta waves recorded in layers V and VI. All cells in the neocortex and specific thalamic nuclei fired rhythmically and phase-locked to the spike component of HVS. RT neurons showed an opposite phase relationship and fired mainly during the wave component of HVS. Half of the NB neurons also showed phasic modulation with HVS. Circumscribed lesion of RT and extensive damage of other thalamic regions, including the intralaminar nuclei, suppressed HVS but had no effect on the neocortical EEG correlates of behavior. In sharp contrast, damage to the NB resulted in a dramatic increase of slow delta waves on the side of the lesion, mimicking the effect of scopolamine administration. We suggest that the NB plays a key role in neocortical arousal by directly activating the neocortex and by suppressing the rhythm generation in the RT-thalamocortical circuitry. We further suggest that the NB system may serve as a structural basis for the concept of the generalized ascending activation of Moruzzi and Magoun (1949).
Tumor-promoting phorbol esters such as 12-O-tetradecanoylphorbol-13-acetate (TPA) directly activate in vitro Ca2+-activated, phospholipid-dependent protein kinase (protein kinase C), which normally requires unsaturated diacylglycerol. Kinetic analysis indicates that TPA can substitute for diacylglycerol and greatly increases the affinity of the enzyme for Ca2+ as well as for phospholipid. Under physiological conditions, the activation of this enzyme appears to be linked to the receptor-mediated phosphatidylinositol breakdown which may be provoked by a wide variety of extracellular messengers, eventually leading to the activation of specific cellular functions or proliferation. Using human platelets as a model system, TPA is shown to enhance the protein kinase C-specific phosphorylation associated with the release reaction in the total absence of phosphatidylinositol breakdown. Various phorbol derivatives which have been shown to be active in tumor promotion are also capable of activating this protein kinase in in vitro systems.
In the mammalian neocortex, the EEG reflects the state of behavioral arousal. The EEG undergoes a transformation, known as activation, during the transition from sleep to waking. Abundant evidence indicates the involvement of the neurotransmitter acetylcholine (ACh) in EEG activation; however, the cellular basis of this involvement remains unclear. We have used electrophysiological techniques with in vivo and in vitro preparations to demonstrate actions of endogenous ACh on neurons in auditory neocortex. In vivo stimulation of the nucleus basalis (NB), a primary source of neocortical ACh, (1) elicited EEG activation via cortical muscarinic receptors, (2) depolarized cortical neurons, and (3) produced a change in subthreshold membrane potential fluctuations from large-amplitude, slow (1-5 Hz) oscillations to low-amplitude, fast (20-40 Hz) oscillations. The NB-mediated change in pattern of membrane potential fluctuations resulted in a shift of spike discharge pattern from phasic to tonic. Stimulation of afferents in the in vitro neocortex elicited cholinergic actions on putative layer 5 pyramidal neurons. Acting via muscarinic receptors, endogenous ACh (1) reduced slow, rhythmic burst discharge and facilitated higher-frequency, single-spike discharge in burst-generating neurons, and (2) facilitated the appearance and magnitude of intrinsic membrane potential oscillations. These in vivo and in vitro observations suggest that neocortical activation results from muscarinic modulation of intrinsic neural oscillations and firing modes. Rhythmic-bursting pyramidal neurons in layer 5 may act as cortical pacemakers; if so, then modifying their discharge characteristics could alter local cortical networks. Larger, intercortical networks could also be modified, due to the widespread projections of NB neurons. Thus, NB cholinergic neurons may play a critical role in producing different states of neocortical function.
Neuronal nicotinic acetylcholine receptor of the alpha 4/non-alpha (alpha 4/n alpha) type was reconstituted in Xenopus oocytes after nuclear injection of cDNA expression vectors. Functional neuronal receptor was only formed when the two subunits alpha 4 and n alpha were coinjected, neither alpha 4 nor n alpha alone being effective. Responses to bath application of acetylcholine (AcCho) have been measured in voltage clamp. AcCho doses as low as 10 nM induce currents of up to 50 nA. Dose-response studies indicate a Kd of about 0.77 x 10(-6) M and a Hill coefficient of 1.5, thus predicting more than one AcCho binding site per receptor molecule. The current-voltage relationship of AcCho-induced currents presents a strong inward rectification. Responses to AcCho were compared to those of three other agonists: L-nicotine, carbachol, and 1,1-dimethyl-4-phenylpiperazinium iodide (DMPP). Sensitivities to AcCho, nicotine, and DMPP are quite similar. Sensitivity to carbachol is much lower, but the currents are otherwise indistinguishable from those induced by AcCho. Five AcCho antagonists--neuronal bungarotoxin (kappa-bungarotoxin), tubocurarine (TC), hexamethonium bromide (Hex), decamethonium bromide (Dec), and mecamylamine (Mec)--have been tested. Neuronal bungarotoxin has no effect on the alpha 4/n alpha channel, whereas 2.5 microM TC reduces by half the current peak evoked by 1 microM AcCho. The block by TC is independent of membrane voltage. By contrast, the block of AcCho-induced currents by Hex or Dec is strongly voltage dependent, suggesting that these substances enter the channel. The block by Mec is detectable at concentrations as low as 100 nM when applied together with 1 microM AcCho and is voltage independent. Hex, Dec, and Mec are effective only when AcCho is present. While the effects of all other agents are fully reversible, the Mec block is persistent.
Intracellular recordings were obtained from neurons in layer II/III of rat frontal cortex. Single-electrode current- and voltage-clamp techniques were employed to compare the sensitivity of excitatory postsynaptic potentials (EPSPs) and iontophoretically evoked responses to N-methyl-D-aspartate (NMDA) to the selective NMDA antagonist D-2-amino-5-phosphonovaleric acid (D-2-APV). The voltage dependence of the amplitudes of the EPSPs before and after pharmacologic changes in the neuron's current-voltage relationship was also examined.
To investigate excitatory postsynaptic potentials (EPSPs), intracellular recordings were performed in layer II/III neurons of the rat medial frontal cortex. The average resting membrane potential of the neurons was more than -75 mV and their average input resistance was >20 MΩ. The amplitudes of the action potentials evoked by injection of depolarizing current pulses were >100 mV. The electrophysiological properties of the neurons recorded were similar to those of regular-spiking pyramidal cells.
The mechanisms of action of acetylcholine (ACh) in the guinea-pig neocortex were investigated using intracellular recordings from layer V pyramidal cells of the anterior cingulate cortical slice. At resting membrane potential (Vm = -80 to -70 mV), ACh application resulted in a barrage of excitatory and inhibitory post-synaptic potentials (p.s.p.s) associated with a decrease in apparent input resistance (Ri). ACh, applied to pyramidal neurones depolarized to just below firing threshold (Vm = -65 to -55 mV), produced a short-latency hyperpolarization concomitant with p.s.p.s and a decrease in Ri, followed by a long-lasting (10 to greater than 60 s) depolarization and action potential generation. Both of these responses were also found in presumed pyramidal neurones of other cortical regions (sensorimotor and visual) and were blocked by muscarinic, but not nicotinic, antagonists. The ACh-induced hyperpolarization possessed an average reversal potential of -75.8 mV, similar to that for the hyperpolarizing response to gamma-aminobutyric acid (GABA; -72.4 mV) and for the i.p.s.p. generated by orthodromic stimulation (-69.6 mV). This cholinergic inhibitory response could be elicited by ACh applications at significantly greater distance from the cell than the slow depolarizing response. Blockade of GABAergic synaptic transmission with solution containing Mn2+ and low Ca2+, or by local application of tetrodotoxin (TTX), bicuculline or picrotoxin, abolished the ACh-induced inhibitory response but not the slow excitatory response. In TTX (or Mn2+, low Ca2+) the slow excitatory response possessed a minimum onset latency of 250 ms and was associated with a voltage-dependent increase in Ri. Application of ACh caused short-latency excitation associated with a decrease in Ri in eight neurones. The time course of this excitation was similar to that of the inhibition seen in pyramidal neurones. Seven of these neurones had action potentials with unusually brief durations, indicating that they were probably non-pyramidal cells. ACh blocked the slow after-hyperpolarization (a.h.p.) following a train of action potentials, occasionally reduced orthodromically evoked p.s.p.s, and had no effect on the width or maximum rate of rise or fall of the action potential. It is concluded that cholinergic inhibition of pyramidal neurones is mediated through a rapid muscarinic excitation of non-pyramidal cells, resulting in the release of GABA. In pyramidal cells ACh causes a relatively slow blockade of both a voltage-dependent hyperpolarizing conductance (M-current) which is most active at depolarized membrane potentials, and the Ca2+-activated K+ conductance underlying the a.h.p.(ABSTRACT TRUNCATED AT 400 WORDS)
This book stemmed from an IBRO symposium that took place in Leipzig, from 12 to 14 August 1987. Local organizers were D. Biesold and V. Bigl from the Department of Neurochemistry at the Paul Flechsig Institute for Brain Research. Some of the contributors to this book were members of the International Program Committee: L. L. Butcher, M.-M. Mesulam, G. Pepeu, and M. Steriade. Leipzig was chosen as the site of a meeting devoted to brain cholinergic systems because some historical steps in this domain are related to the German city. Indeed, after the initial description of the substantia innominata (die ungennante Marksubstanz) by J. C. Reil (1809), T. Meynert, one of the founders of scientifically oriented psychiatry, designated this structure as Ganglion der Hirnschenkelschlinge in a volume published in Leipzig (1872). The name of Meynert was linked to the nucleus basalis by A. Koelliker in a Handbook, also published in Leipzig (1896).
Since edited volumes rarely allow the expression of coherence between the numerous authors, we decided to ask for chapters from only a limited number of participants and we invited other colleagues, who could not attend the symposium, to submit their contribution. Our goal was to present current data and concepts about the morphology, physiology, and pathology of brainstem and basal forebrain cholinergic systems controlling the excitability of the thalamus and cerebral cortex. It is a pleasure to extend our thanks to all colleagues, whose names and affiliations are listed, for taking time from busy lives to survey their fields of interest. We also express our appreciation to Oxford University Press for an agreeable collaboration and support in the preparation of this book.
Slices of sensorimotor and anterior cingulate cortex from guinea pigs were maintained in vitro and bathed in a normal physiological medium. Electrophysiological properties of neurons were assessed with intracellular recording techniques. Some neurons were identified morphologically by intracellular injection of the fluorescent dye Lucifer yellow CH. Three distinct neuronal classes of electrophysiological behavior were observed; these were termed regular spiking, bursting, and fast spiking. The physiological properties of neurons from sensorimotor and anterior cingulate areas did not differ significantly. Regular-spiking cells were characterized by action potentials with a mean duration of 0.80 ms at one-half amplitude, a ratio of maximum rate of spike rise to maximum rate of fall of 4.12, and a prominent afterhyperpolarization following a train of spikes. The primary slope of initial spike frequency versus injected current intensity was 241 Hz/nA. During prolonged suprathreshold current pulses the frequency of firing adapted strongly. When local synaptic pathways were activated, all cells were transiently excited and then strongly inhibited. Bursting cells were distinguished by their ability to generate endogenous, all-or-none bursts of three to five action potentials. Their properties were otherwise very similar to regular-spiking cells. The ability to generate a burst was eliminated when the membrane was depolarized to near the firing threshold with tonic current. By contrast, hyperpolarization of regular-spiking (i.e., nonbursting) cells did not uncover latent bursting tendencies. The action potentials of fast-spiking cells were much briefer (mean of 0.32 ms) than those of the other cell types.(ABSTRACT TRUNCATED AT 250 WORDS)
1. Intracellular recordings were obtained from neurons of the guinea pig sensorimotor cortical slice maintained in vitro. Under control recording conditions input resistances, time constants, and spiking characteristics of slice neurons were well within the ranges reported by other investigators for neocortical neurons in situ. However, resting potentials (mean of -75 mV) and spike amplitudes (mean of 93.5 mV) were 10-25 mV greater than has been observed in intact preparations. 2. Current-voltage relationships obtained under current clamp revealed a spectrum of membrane-rectifying properties at potentials that were subthreshold for spike generation. Ionic and pharmacologic analyses suggest that subthreshold membrane behavior is dominated by voltage-sensitive, very slowly inactivating conductances to K+ and Na+. 3. Action potentials were predominantly Na+ dependent under normal conditions but when outward K+ currents were reduced pharmacologically, it was possible, in most cells, to evoke a non-Na+-dependent, tetrodotoxin-(TTX) insensitive spike, which was followed by a prominent depolarizing after-potential. Both of these events were blocked by the Ca2+ current antagonists, Co2+ and Mn2+. 4. A small population of neurons generated intrinsic, all-or-none burst potentials when depolarized with current pulses or by synaptic activation. These cells were located at a narrow range of depths comprising layer IV and the more superficial parts of layer V. 5. Spontaneous excitatory synaptic potentials appeared in all neurons. Spontaneous inhibitory events were visible in only about 10% of the cells, and in those cases apparently reversed polarity at a level slightly positive to resting potential. Stimulation of the surface of the slice at low intensities evoked robust and usually concurrent excitatory and inhibitory synaptic potentials. Unitary inhibitory postsynaptic potentials (IPSPs) reversed at levels positive to rest. Stronger stimulation produced a labile, long-duration, hyperpolarizing IPSP with a reversal potential 15-20 mV negative to the resting level. 6. Neocortical neurons in vitro retain the basic membrane and synaptic properties ascribed to them in situ. However, the array of passive and active membrane behavior observed in the slice suggests that cortical neurons may be differentiated by specific functional properties as well as by their extensive morphological diversity.
Layer V pyramidal cells of the motor cortex of cats are necessary for short-latency blink conditioning (Woody et al., 1974). Increases in their excitability support production of this learned motor response (Woody et al., 1970; Woody and Engel, 1972; Woody and Black-Cleworth, 1973; Brons and Woody, 1980). Early evidence suggested (Woody and Black-Cleworth, 1973) that a decreased postsynaptic conductance might support the excitability increase. Further studies indicate that the input resistance (R
m) of these cells is increased by acetylcholine (ACh), cGMP, and cGMP-dependent protein kinase (cGPK) but not by cAMP or Ca2+ (Woody et al., 1978, 1986; Swartz and Woody, 1979, 1984; Wallis et al., 1982; Bartfai et al., 1985; Woody and Gruen, 1986b). The increase in R
m is persistent when depolarization-induced cell discharge accompanies application of ACh, cGMP, or cGPK. Recent studies using single-electrode voltage-clamp techniques in vivo now provide direct evidence that ACh and cGPK decrease a net outward current in these cells (Woody and Gruen, 1986a). We suggest that a persistent decrease in this current may mediate the excitability increase that supports short-latency conditioned blinking.
In recent years substantial advancement has occurred toward understanding integrative functions of the nervous system. Acetylcholine (ACh), dopamine (DA), norepinephrine (NE), and serotonin (5-HT), are substances that have gained general recognition as neurotransmitters in the sense of having a direct influence on the magnitude of membrane potential. These substances are now also characterized as neuromodulators, a designation which encompasses observations of complex electrophysiologic and metabolic effects in target neurons that occur in addition to, or are independent of, simple excitatory and inhibitory changes in the membrane potential (for reviews, see Dismukes, 1979; Kupfermann, 1979; Daly et al., 1980; Kaczmarek and Levitan, 1987).
Cloning and sequence analysis of DNA complementary to porcine cerebral
messenger RNA encoding the muscarinic acetylcholine receptor predict the
complete amino-acid sequence of this protein. Expression of the
complementary DNA produces functional muscarinic receptor in Xenopus
oocytes. The muscarinic receptor is homologous with the
β-adrenergic receptor and rhodopsin in both amino-acid sequence and
suggested transmembrane topography.
Exogenously applied muscarinic agonists—for example, acetylcholine (ACh) and acetyl-beta-methacholine (MCh)—modify frequency receptive fields in auditory cortex of unanesthetized animals in a frequency-specific rather than global manner. The present study sought to relate these findings to endogenous actions of ACh by using the anticholinesterase agents eserine sulphate and soman (0-1,2,2-trimethylpropylmethylphosphonofluoridate) to facilitate the effects of endogenous ACh. Frequency receptive fields (FRF) were determined by presenting sequences of different isointensity tones before, during, and after application of ACh, MCh, eserine, or soman; also the cholinesterase blockers were applied between applications of ACh or MCh. The major effects produced by the inhibitors were similar to those of the agonists. Predominant effects were frequency-specific changes in FRF. Further, eserine and soman, similar to ACh and MCh, produced shifts in the best frequency (BF) of FRF due mainly to coordinated depression of responses to the BF and increased responses to adjacent, non-BF. The results indicate that exogenous and endogenous ACh, acting via muscarinic receptors, can significantly influence the physiological functioning of cortical neurons and consequently their processing of sensory information.
Previously we reported that acetylcholine (ACh) and acetyl-beta-methacholine (MCh) modify responses of neurons in auditory cortex to individual frequencies. The purpose of this study was to determine whether muscarinic agonists produce frequency-specific alterations or general changes in cellular responses. Frequency-specific modifications would be evident in alterations of frequency receptive fields (FRF) that differed across frequencies while general effects would be seen as changes that were more or less the same over frequencies. Responses of single neurons to designated sets of tones were recorded in the auditory cortex of chronically prepared awake cats before, during, and following ejection of ACh or MCh by iontophoresis or micropressure using multibarrel micropipettes. Frequency receptive fields were determined by presenting isointensity tones across a range of frequencies including the cell's best frequency (BF) to tone onset. FRF for “off” and “sustained (through)” responses were also determined quantitatively. The effects of ACh and MCh were predominantly frequency-specific (77%, 39/51 cells); general changes (19%, 10/51) and no effects (4%, 2/51) were less likely. Frequency-specific effects involved both facilitation and reduction of the same response component to different frequencies within the same neuron. For responses to tone onset (but not “through” and “off” responses), agonists were more likely to produce a decrease at the BF while simultaneously increasing responses to other frequencies. Agonists could increase or decrease frequency selectivity. Effects of agonists could be blocked by atropine, suggesting involvement of muscarinic receptors.
Structural analogs of oxotremorine have been employed to examine the relationship between the binding of agonists to muscarinic receptors in guinea pig cerebral cortex and the enhancement of inositol lipid turnover. Large differences were observed in the ability of the analogs to stimulate inositol phospholipid turnover, as measured both by the increase in labeling of phosphatidate and phosphatidylinositol from 32Pi in a nerve-ending fraction, and by the stimulated release of labeled inositol phosphates from slices of cerebral cortex, a direct measure of inositol lipid breakdown. The quaternary N+ analogs, oxotremorine-M and its N-methylacetamide derivative, were five to thirteen times as effective as oxotremorine. In contrast, methyl substitution of the pyrrolidone ring of oxotremorine resulted in a complete loss of agonist activity. Receptor occupancy data obtained from the displacement of labeled quinuclidinyl benzilate bound to receptors in a nerve-ending fraction indicated that the more efficacious agonists interacted with at least two affinity forms of the muscarinic receptor, whereas the less effective agonists bound to a single affinity form. Dose-response curves obtained in the presence of oxotremorine-M for inositol lipid turnover in both the nerve-ending fraction and slice preparation correlated with the occupancy of a single low-affinity form of the muscarinic receptor. The results suggest that the differential abilities of analogs of oxotremorine to enhance inositol lipid turnover in brain are closely related to the extent of agonist-induced conformational change in the muscarinic receptor.
Neurons of in vitro guinea pig and rat auditory cortex receive a complex synaptic pattern of afferent information. As many as four synaptic responses to a single-stimulus pulse to the gray or white matter can occur, an early-EPSP followed, sequentially, by an early-IPSP, late-EPSP. and late-IPSP. Paired pulse stimulation and pharmacological studies show that the early-IPSP can modify information transmission that occurs by way of the early-EPSP. Each of these four synaptic responses differed in estimated reversal potential and each was differentially sensitive to antagonism by pharmacological agents. DNQX (6,7-dinitroquinoxaline-2,3-dione), a quisqualate/kainate receptor antagonist, blocked the early-EPSP and the late-EPSP was blocked by the NMDA receptor antagonist APV (D-2-amino-5-phosphonovalerate). The early-IPSP was blocked by the GABA-a receptor antagonist bicuculline. and the late-IPSP by the GABA-b receptor antagonists 2-OH saclofen or phaclofen. Presentation of stimulus trains, even at relatively low intensities, could produce a long-lasting APV-sensitive membrane depolarization. Also discussed is the possible role of these synaptic potentials in auditory cortical function and plasticity.
The cerebral cortex receives a prominent cholinergic innervation which is thought to play an important role in regulating its normal function. Electrophysiological studies have shown that activation of cholinergic receptors results in a marked enhancement of excitatory stimuli onto cortical neurons and it has been suggested that this effect is secondary to the blockade of several voltage- and calcium-dependent potassium conductances in these cells. It is reported here that, in addition to these effects, activation of muscarinic receptors in the prefrontal cortex elicits the appearance of a slow calcium-dependent inward current in response to the generation of action potentials. This inward aftercurrent produces a slowly decaying depolarizing afterpotential which, when activated by stimulation of the cell, can summate with the carbachol-induced depolarization greatly increasing its magnitude. As a result the ability of muscarinic receptor to elicit a depolarization and excite cells in this region can be dramatically potentiated by evoked cell activation. This effect expands the range of mechanisms by which muscarinic receptors can facilitate excitatory inputs and provides a mechanism by which the association of brief excitatory stimuli to cholinergic stimulation can selectively enhance muscarinic responses among discrete cell populations in the cerebral cortex.
To determine if classical conditioning produces general or specific modification of responses to acoustic conditioned stimuli (CS), frequency receptive fields (RF) of neurons in guinea pig auditory cortex were determined before and up to 24 h after conditioning. Highly specific RF plasticity characterized by maximal increased responses to the CS frequency and decreased responses to the pretraining best frequency (BF) and other frequencies was observed in 70% of conditioning cases. These opposing changes were often sufficient to produce a shift in tuning such that the frequency of the CS became the new BF. CS frequency specific plasticity was maintained as long as 24 h. Sensitization training produced general increased responses across the RF without CS specificity. The findings indicate that associative processes produce systematic modification of the auditory system's processing of frequency information and exemplify the advantages of combining receptive field analysis with behavioral training in the study of the neural bases of learning and memory.
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The dendritic spines of many central neurons are generally thought to modulate the ability of individual synaptic conductances to depolarize the dendritic shaft. A compartmental analysis using typical spine dimensions shows that spine neck resistances are probably far too low to support such a function, because low conductance synapses act as time-varying current sources. However, the collective presence of all spines on a dendrite significantly modifies the electrical properties of the branch in ways which have previously been overlooked. In particular, they lower its input impedance and length constant, reducing the amplitude of the unitary excitatory postsynaptic potential as well as the strength of spatial summation. This enables a dendrite to integrate large numbers of synaptic inputs while occupying minimal volume. In this way, dendritic spines are analogous to axonal myelin, which also alters transcellular impedance in order to maximize neurite function and minimize volume. Unlike membrane resistance changes, spines have little effect on the membrane time-constant so they maintain a long window for temporal summation. Though spine shape and neck resistance do not significantly affect dendritic potentials, spine area does. Therefore, while changes in spine morphology probably do not directly potentiate the strength of individual synapses, changes in spine density can regulate the synaptic excitability of an entire dendrite.
1. The cellular mechanism by which acetylcholine (ACh) potentiates neuronal responses to N-methyl-D-aspartate (NMDA) was investigated in CA1 neurones of hippocampal slices using current- and voltage-clamp techniques. 2. Loading cells with 5'-guanylylimidodiphosphate (GppNHp) caused a gradual increase in response to NMDA. Pulses of ACh accelerated this increase. Guanosine 5'-O-(2-thiodiphosphate) (GDP beta S) blocked the potentiating effect of ACh on responses to NMDA. 3. Acute LiCl caused a gradual decrease in the potentiating effect of ACh, while the potentiation was completely prevented by 3 day chronic 6 mequiv/kg (I.P.) LiCl treatment and restored by acute treatment with 10 mM-inositol. 4. Loading cells with a general protein kinase inhibitor, H-7, enhanced the potentiating effect of ACh on responses to NMDA and blocked the effect of ACh on the after-hyperpolarization (AHP). 5. Ultraviolet irradiation of cells loaded with a photolabile inositol 1,4,5-trisphosphate (InsP3) caused a transient increase in responses to NMDA, while penetrating cells with active InsP3-containing pipettes caused a gradual BAPTA-sensitive increase in responses to NMDA. 6. Reducing the rate of InsP3 metabolism, with 2,3-diphosphoglyceric acid (DPG), caused an increase and prolongation of the potentiating effect of ACh, while blocking the InsP3 receptor with heparin prevented the cholinergic potentiation. 7. NMDA, by itself, potentiated subsequent responses to NMDA, an effect that was blocked when [Ca2+]i was chelated with BAPTA. NMDA and ACh were also found to compete in potentiating responses to NMDA. Finally, the cholinergic potentiation was blocked when cells were loaded with BAPTA. 8. We propose that activation of the InsP3 branch of the phosphoinositide pathway potentiated responses to NMDA and that InsP3 exerted this effect by elevating [Ca2+]i.
With the use of in vitro preparations and sophisticated electrophysiological recording techniques, we are beginning to understand in great detail how a wide variety of putative neurotransmitters alter neuronal excitability. Many of these recent results have changed our previous conceptualization of excitatory and inhibitory synaptic transmission. For example, as we reviewed, it is now clear that in many regions of brain, EPSPs, although mediated by a single neurotransmitter, glutamate, are composed of two components that subserve distinct functions. Similarly, the elucidation of the different properties of GABA(A) and GABA(B) receptors has demonstrated that inhibition in the brain is also not a single process but most likely has at least two components. It is also now well established that many neurotransmitter receptors are coupled to second messengers, the activation of which in turn alters ion channel activity. Many of these second messenger systems modulated voltage-dependent channels, resulting not in a simple membrane depolarization or hyperpolarization but rather an effect that depends on the membrane potential of the cell, a property that is constantly changing in both the spatial and temporal domains. This type of modulation can result in drastic changes in the cells' input-output properties. By being coupled to the same second messenger system or G protein, distinct neurotransmitter receptors may induce the same electrophysiological response. Figure 1 shows this convergence of action of various neurotransmitter receptors onto two types of ion channels, an inwardly rectifying K+ channel (I(K)) and a Ca2+-activated K+ channel (I(AHP). In many cases this convergence can be seen in a single type of neuron. Thus GABA(B), 5-HT(1a), A1, and probably SS receptors are coupled to the same K+ channel in CA1 hippocampal pyramidal cells. In locus coeruleus neurons, μ-opioid, α2-noradrenergic, SS, and probably GABA(B) receptors all activate the same K+ conductance. In lateral parabrachial neurons, μ-opioid, M2, and GABA(B) receptors increase the same K+ conductance. Finally, in substantia nigra neurons, GABA(B) and D2 receptors, activate the same K+ conductance. For I(AHP), all of the neurotransmitters listed act on CA1 hippocampal pyramidal cells and converge onto the same K+ conductance. The receptor subtype of the 5-HT effect and the coupling mechanism for the 5-HT and ACh effects have not been clearly established. In addition to convergence, the use of G proteins and second messenger systems to mediate the actions of neurotransmitters may offer additional advantages. For example, parts of a cell spatially segregated from a given receptor could be affected by activation of that receptor via its coupling to diffusible second messengers. Actions mediated by second messengers might be long lasting and more subject to subtle, long-term modulations. It is also likely that neurotransmitters that activate receptors coupled to G proteins and second messengers have functionally important biochemical effects in addition to directly modulating ion channels. Although different neurotransmitters often appear to have similar electrophysiological actions, they can also be shown to have quite distinct actions, primarily because they activate a variety of subtypes of receptors that are coupled to distinct G proteins and second messengers. Thus the same neurotransmitter can have distinct actions on different cells or different parts of the same cell because of the differential distribution of receptor subtypes. A corollary of this is that if a given neurotransmitter is known to have different actions in different brain regions, it most likely is because the receptors it is activating are distinct. Figures 2-4 show this striking divergence of action for five different neurotransmitters. Glutamate probably activates five types of receptor. Three of these, the AMPA, the kainate, and the NMDA receptors, cause a rapid increase in cation conductance without an intervening coupling molecule. A subtype of quisqualate receptor, which differs pharmacologically from the quisqualate receptor linked to ion channels (now generally referred to as the AMPA receptor), is coupled to PI turnover. The physiological consequence of activating this receptor has yet to be observed in CNS neurons. Activation of the AP4 receptor causes a presynaptic inhibition in a number of pathways within the CNS, perhaps by decreasing Ca2+ entry into the nerve terminal. There is only limited data that glutamate can act on this presynaptic receptor. γ-Aminobutyric acid activates two types of receptors. The GABA(A) recepors rapidly open Cl- channels without an intervening coupling molecule.
The present study shows that [3H]4-DAMP binds specifically, saturably, and with high affinity to muscarinic receptor sites in the rat brain. In homogenates of hippocampus, cerebral cortex, striatum, and thalamus, [3H]4-DAMP appears to bind two sub-populations of muscarinic sites: one class of high-affinity, low capacity sites (Kd less than 1 nM; Bmax = 45-152 fmol/mg protein) and a second class of lower-affinity, high capacity sites (Kd greater than 50 nM; Bmax = 263-929 fmol/mg protein). In cerebellar homogenates, the Bmax of [3H]4-DAMP binding sites was 20 +/- 2 and 141 +/- 21 fmol/mg protein for the high- and the lower-affinity site, respectively. The ligand selectivity profile for [3H]4-DAMP binding to its sites was similar for both the high- and lower-affinity sites; atropine = (-)QNB = 4-DAMP much greater than pirenzepine greater than AF-DX 116, although pirenzepine was more potent (16-fold) at the lower- than at the high-affinity sites. The autoradiographic distribution of [3H]4-DAMP sites revealed a discrete pattern of labeling in the rat brain, with the highest densities of [3H]4-DAMP sites present in the CA1 sub-field of Ammon's horn of the hippocampus, the dentate gyrus, the olfactory tubercle, the external plexiform layer of the olfactory bulb and layers I-II of the frontoparietal cortex. Although the distribution of [3H]pirenzepine sites was similar to that of [3H]4-DAMP sites in many brain regions, significant distinctions were apparent. Thus, both the ligand selectivity pattern of [3H]4-DAMP binding and the autoradiographic distribution of sites suggest that although the high-affinity [3H]4-DAMP sites may consist primarily of muscarinic-M3 receptors, the lower-affinity [3H]4-DAMP sites may be composed of a large proportion of muscarinic-M1 receptors.
We have hypothesized that auditory cortex plasticity involves modification of thalamocortical transmission by basal forebrain (BF) cholinergic neurons, and that this action may involve muscarinic receptors. In a first test of this hypothesis, we report that BF stimulation can suppress or facilitate, depending on the intensity of stimulation, auditory cortical responses elicited by thalamic stimulation. BF-mediated facilitation is antagonized by atropine, implicating muscarinic receptors. These data suggest that BF cholinergic neurons functionally modify auditory cortex by regulating thalamocortical transmission.
Slow excitatory postsynaptic potentials (EPSPs) were identified in rat neocortical slices. Such potentials, resistant to blockade of glutamate and gamma-aminobutyric acid-A (GABAA) receptors, were partially antagonized by muscarinic or beta-adrenergic antagonists separately, and completely blocked when these agents were added in combination. Slow EPSPs were enhanced by a cholinesterase inhibitor or catecholamine reuptake blockers. Spontaneous epileptic discharges induced by picrotoxin also triggered slow EPSPs. Such potentials were pharmacologically identical to those induced by electrical stimulation under normal conditions. A non-conventional mechanism for synaptic transmission is postulated to account for triggering of slow EPSPs by epileptic discharges.
In 1954, L-glutamate (Glu) and L-aspartate (Asp) were first suggested as being excitatory synaptic transmitters in the cerebral cortex. Since then, evidence has mounted steadily in favor of the view that Glu and Asp are major excitatory transmitters in the neocortex. Many of the experimental studies which reported how Glu/Asp came to satisfy the criteria for transmitters in the neocortex are reviewed here, according to the methods employed. Since the question of which particular synaptic sites in cortical neural circuits Glu/Asp operate as excitatory transmitters has not previously been reviewed, particular attention is given to efferent, afferent and intrinsic neural circuits of the visual and somatosensory cortices, where circuitry is relatively clearly delineated. Recent studies using chemical assays of released amino acids, high-affinity uptake mechanisms of Glu/Asp from nerve terminals, the direct micro-iontophoretic administration of Glu/Asp antagonists, and immunocytochemical techniques have demonstrated that almost all corticofugal efferent projections employ Glu/Asp as excitatory synaptic transmitters. Evidence indicating that thalamocortical afferent projections, including geniculocortical projections and some intrinsic connections are glutamatergic, is also reviewed. Thus, the results highlighted here indicate that the main framework of neocortical circuitry is operated by Glu/Asp. Pharmacological studies indicate that synaptic receptors for Glu/Asp can be classified into a few subtypes, including N-methyl-D-aspartate (NMDA) and quisqualate/kainate (non-NMDA) types. Some evidence indicating the sites of operation of NMDA and non-NMDA receptors in neocortical circuitry is reviewed, and the distinct, functional significance of these two types of Glu/Asp receptors in information processing in the neocortex is proposed.
Action of antagonists for excitatory amino-acid (EAA) receptors on extracellularly and intracellularly recorded responses of layer II/III cels to electrical stimulation of the underlying white matter were studied in a slice preparation of rat's visual cortex. Antagonists used were 2-amino-5-phosphonovalerate (APV) and 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX), which are selective antagonists for EAA receptors of N-methyl-D-aspartate (NMDA) and quisqualate/kainate (non-NMDA) type, respectively. In extracellular recordings, it was found that responses of almost all of the cells were suppressed by CNQX. In contrast, sensitivity to APV was different between cells with short-and long-latency responses; 81% of the former responses were not suppressed by APV, while about a half of the latter were suppressed. Excitatory postsynaptic potentials (EPSPs) evoked by white-matter stimulation were recorded intracellularly from 42 neurons. Most of polysynaptically elicited EPSPs were sensitive to AVP, whereas the majority of monosynaptic EPSPs, were not. CNQX almost completely suppressed EPSPs irrespective of monosynaptically or polysynaptically evoked, but in some cases slow EPSPs with low amplitude were spared. These CNQX-resistant EPSPs were elicited polysynaptically and had an anomalous voltage dependence, a characteristic of NMDA receptors. It is suggested that non-NMDA receptors contribute dominantly to first-order synaptic transmission while NMDA receptors participate substantially in second-order transmission so as to serve as a booster of outputs from visual cortex.
Muscarinic synaptic activation is known to be involved in cortical arousal as well as learning. Although simple increases in the electrical responsiveness of neurons might be the basis of arousal, the linkage of muscarinic transmission to the synaptic plasticity that might underlie learning is lacking. Most models of synaptic plasticity involve postsynaptic Ca2+ changes as a trigger for subsequent processes. We imaged muscarinic effects on free Ca2+ accumulation during intracellular recordings from CA3 pyramidal neurons in the guinea pig hippocampal slice. Muscarinic activation, either by repetitive stimulation of cholinergic fibers or by bath-applied carbachol, strongly increased intradendritic Ca2+ accumulation during directly evoked repetitive firing, in part by blocking a Ca(2+)-dependent K+ conductance. The effects of repetitive stimulation of cholinergic fibers were enhanced by the acetylcholine-esterase blocker eserine and blocked by the muscarinic antagonist atropine. These findings demonstrate a novel muscarinic reinforcement of Ca2+ changes during excitation, which are probably significant for synapse modification.
We have examined the activation of phosphoinositide metabolism by muscarinic agonists in rat cerebral cortex, in an attempt to delineate the mechanisms by means of which some selective antagonists inhibit this response in a manner that deviates from simple mass action law. The accumulation of [3H]inositol phosphates induced by the full agonist carbamylcholine in cell aggregates preparations was inhibited by muscarinic antagonists with the following order of potency: telenzepine greater than atropine greater than 4-diphenylacetoxy-N-methyl-piperidine methbromide greater than pirenzepine greater than hexahydro-sila-difenidol greater than AF-DX 116. The same order of potency was found for the competition of these antagonists with [3H]telenzepine binding to M1 muscarinic receptors. The inhibition of the formation of [3H]inositol phosphates activated by acetylcholine, carbamylcholine, and oxotremorine-M by pirenzepine and telenzepine showed biphasic curves, with 62-73% of the response being inhibited with high affinity. Atropine, AF-DX 116, and pirenzepine shifted the concentration-response curves of oxotremorine-M to the right in a parallel manner. However, pirenzepine at micromolar concentrations showed deviation from linearity of the Schild regression. The blockade by high concentrations of pirenzepine and telenzepine showed less than additive dose ratios when assayed in the presence of atropine, suggesting deviation of their antagonism from simple competition. However, after alkylation with propylbenzilylcholine mustard in the presence of low concentrations of pirenzepine, the response to carbamylcholine and oxotremorine-M showed monophasic inhibition curves by pirenzepine and linear Schild regression for this antagonist. These results support the interpretation that the formation of [3H]inositol phosphates is activated by multiple muscarinic receptor subtypes in rat cerebral cortex. The profile of affinities of muscarinic antagonists indicates that a major component of the response is activated by an M1 receptor subtype and a minor component is probably mediated by M3 muscarinic receptors when acetylcholine, carbamylcholine, or oxotremorine-M are used to stimulate the response. Conversely, pirenzepine inhibited the response induced by methacholine and bethanechol in a monophasic manner with high affinity (Ki = 13 nM), suggesting that these agonists can selectively stimulate phosphoinositide metabolism through activation of M1 muscarinic receptors in rat cerebral cortex.
Excitatory amino acid receptors are the mediators of synaptic transmission at many synapses that can undergo use-dependent modifications of synaptic efficiency. They also play an essential role in the induction of these plastic changes. Graham Collingridge and Wolf Singer describe how NMDA receptors can endow synapses with hebbian-like properties and discuss how these may be used by vertebrates for associative learning and experience-dependent modifications of synaptic connections during development. The role of AMPA receptors in the maintenance of long-term potentiation is also discussed.
This chapter describes the cholinergic system in Alzheimer disease (AD). Choline acetyltransferase (ChAT), the synthetic enzyme for acetylcholine (ACh), is consistently reduced by 50%–95% in cortex and hippocampus of AD patients compared to age-matched controls. Reductions are also observed in high-affinity choline uptake (HACU), in in vitro synthesis of ACh and release during depolarization, in presynaptic muscarinic and nicotinic receptor binding, in ACh and acetylcholinesterase (AChE) levels in cortex and in cerebrospinal fluid (CSF). The reduction of these presynaptic cholinergic markers is associated with a marked loss of cells in the nucleus basalis of Meynert, which project to cortex. The postsynaptic muscarinic receptor mechanisms appear to be relatively spared in AD patients. The reductions of cortical and CSF cholinergic markers are closely correlated with the extent of neuropathology and with the severity of cognitive impairment. The chapter also discusses the changes in ACh and Ch metabolism in aging animals and in the CSF of AD patients, which may be related to neuronal membrane breakdown and reduced uptake of Ch by cholinergic neurons.
This chapter discusses the pharmacological muscarinic receptor subtypes. The introduction of several selective muscarinic receptor antagonists has permitted the classification of muscarinic receptors into three pharmacological receptor subtypes, M1, M2, and M3, with indications for a fourth type, and has played a role in prompting the discovery of the molecular forms of the receptor. The distribution of muscarinic receptor subtypes in body tissues are described by hybridization techniques and has provided insight in elucidating the pharmacologically defined receptors present in smooth muscle and striatum, using pirenzepine (PZ), AF-DX 116, hexahydrosiladifenidol (HHSiD), and methoctramine (METH). Affinity constants for equilibrium binding of PZ to muscarinic receptors in membranes from the brain and peripheral tissues are presented in the chapter. The tissue selectivity profile correlated well with the pharmacological activity of the drug determined in vivo, and in vitro on isolated organ preparations. The property of PZ to discriminate highly between the M1 and M2 receptors lead to the detection of heterogeneity of receptors in the rat striatum and to the characterization of the receptor subtypes with other selective compounds.
Intracellular recording techniques were used to compare the effects of the M2 muscarinic antagonists, AF-DX 116 and gallamine, on membrane potential (Vm), input resistance (Ri), responses induced by methacholine, muscarinic slow postsynaptic potentials and action potentials in the superior cervical ganglion of the rabbit. Gallamine or AF-DX 116 antagonized methacholine-induced or synaptically-evoked muscarinic hyperpolarization, without having significant effect on depolarization induced by methacholine or synaptically. The drug AF-DX 116 reduced evoked muscarinic hyperpolarizing potentials, without significant change in Vm or Ri, recorded in the absence of muscarinic stimulation. In contrast to AF-DX 116, gallamine elicited a concentration-dependent depolarization of the membrane, with a corresponding increase in Ri, when tested in the absence of muscarinic stimulation. These effects of gallamine were accompanied by an increase in duration and decrease in the slope of the descending phase of the action potential. Blockade by gallamine of evoked hyperpolarization was independent of membrane depolarization and readily occurred when gallamine-induced depolarization was prevented by clamping Vm at its pre-gallamine level. The effects of gallamine were maintained during its presence and reversed upon washing with gallamine-free physiological solution. These results indicate that AF-DX 116 and gallamine have a specificity for antagonism of muscarinic responses, mediated by receptors of the M2 type in the superior cervical ganglion. However, gallamine, while an effective antagonist of M2 responses, also has the ability to modify the electrical characteristics of ganglion cells and thus may modify ganglionic transmission by mechanisms other than antagonism of receptors.
Intracellular recordings were obtained from pyramidal neurons in layer 5 of rat somatosensory and visual cortical slices maintained in vitro. When directly depolarized, one subclass of pyramidal neurons had the capacity to generate intrinsic burst discharges and another generated regular trains of single spikes. Burst responses were triggered in an all-or-none manner from depolarizing afterpotentials in most bursting neurons. Regular spiking cells responded to electrical stimulation of ascending afferents with a typical EPSP-IPSP sequence, whereas IPSPs were hard to detect in bursting cells. Orthodromic activation of the latter evoked a prominent voltage-dependent depolarization that could trigger a burst response. Intracellularly labelled bursting and regular spiking cells were located in layer 5b, but had distinctly different morphologies. Bursting neurons had a large pyramidal soma, a gradually emerging apical dendrite, and an extensile apical and basal dendritic tree. Their axonal collateral arborization was predominantly limited to layers 5/6. In contrast, regular spiking cells had a more rounded soma with abruptly emerging apical dendrite, a smaller dendritic arborization, and 2 to 8 ascending axonal collaterals that arborized widely in the supragranular layers. Both bursting and regular spiking cells had main axons that entered the subcortical white matter.
Acetylcholinesterase (AChE)-rich projections from basal forebrain to neocortex cerebri were characterized in the present study. The purpose was to investigate 3 aspects of these projections in rats and mice that have been incompletely described in previous work: intracortical organization of the fibers, subcortical pathways and axonal branching patterns of individual basal forebrain neurons. AChE histochemistry, lesions and Golgi impregnations were the principal strategies employed in this light microscopic study. The moderately dense, AChE-stained innervation of neocortex can be altered by intracortical lesions. The results depended on the region involved and the orientation of the lesion. Sagittal knife cuts had barely detectable effects, regardless of sites. Coronal knife cut lesions in medial cortex resulted in substantial loss of staining in cingulate and medial occipital fields. In contrast, coronal lesions of lateral or anterior cortex produce only small zonal reductions in staining. The interpretation of the latter findings that we favor is that AChE-rich basal forebrain fibers enter lateral/anterior cortex and branch densely there, but in tangentially limited and overlapping terminal domains. Observations on the topography and targets of AChE-rich basal forebrain cortical afferents revealed that the fibers could be grouped based on certain characteristics. Three sets of fibers were distinguishable: anterior pathway innervating cortex of the frontal pole. These fibers were traceable to the region of the substantia innominata/nucleus basalis. They crossed the neostriatum and external capsule in the sagittal plane, forming in 3 dimensions an orderly sheet-like array of fibers bridging the anteroventral surface of the neostriatum with nearby polar cortex medial pathway innervating cingulate and medial occipital cortex. Emerging predominantly from the region of the diagonal band, the fibers run caudally as a triangular bundle in deep layer VI of cingulate cortex. lateral pathway innervating most of remaining lateral neocortex. The fibers radiate out from substantia innominata/nucleus basalis with a complex 3-dimensional organization. In all pathways, fibers enter and initially run within layer VI before ascending pialward, although the intracortical course in layer VI differs between pathways. These fibers primarily terminate in layer V with a secondary concentration in layer I. However, the latter appears to receive substantial AChE-stained inputs from other sources, possibly intracortical, as well. The pathways overlap at their respective boundary zones. This system is comparably organized in rats and mice.(ABSTRACT TRUNCATED AT 400 WORDS)
The nucleus basalis of Meynert (NBM) has recently been identified as the primary source of acetylcholine (ACh) in the cerebral cortex. Damage to the NBM in Alzheimer's disease appears to account for the consistent loss of cholinergic markers in cortex associated with this disease. These findings have drawn attention to the possible functions of the NBM and the other components of the basal forebrain cholinergic system. Cholinergic neurotransmission has long been thought to be involved in memory and arousal, and current research has provided new insights into the specific role of the NBM in these processes. Recent findings have also led to new concepts of how the NBM may interact with other neural structures. This review discusses some of the recent hypotheses of the functions of the NBM, particularly its possible role in learning.
The cloning of cDNAs and genes for five different muscarinic acetylcholine receptors provides a new basis for characterizing muscarinic receptor function. Studies of the cloned receptors when introduced into cells not expressing endogenous receptors have allowed the initial identification of two classes of functional response. The m1, m3 and m5 receptors belong to a class characterized by agonist-induced stimulation of phosphatidylinositol metabolism and are structurally more related to each other than they are to the m2 and m4 receptors, which belong to a class associated with agonist-induced inhibition of adenylate cyclase. While functional differences within these classes may yet be found, it appears likely that much of the difference between functionally similar receptors will be found to lie in their regulation.
In dissociated cellular preparations of adult rat cortex, M1 and M2 muscarinic receptors were shown to mediate phosphoinositide metabolism and cAMP inhibition, respectively. Additionally, in dissociated striatum, an M2 receptor was shown to inhibit the level of cAMP. The components of "receptor reserve" in these three receptor-effector systems were evaluated by the method of partial receptor inactivation and the dissociation constants for the full agonist carbachol were determined. In dissociated cellular preparations of cortex, and in the presence of 10 mM lithium ion, carbachol (EC50 = 116 microM) activated an M1 receptor subtype (atropine Ki = 0.9 nM; pirenzepine Ki = 9.3 nM) to elicit up to a 7-fold release over the basal level of [3H]inositol 1-phosphate. Carbachol was 100-fold more potent (EC50 approximately 1 microM) in the inhibition of forskolin-elevated [3H]cAMP levels in both the cortex (maximally 28%) and striatum (maximally 49%). Pirenzepine blocked the [3H]cAMP inhibition responses to carbachol in cortex and striatum with Ki values of 334 nM and 313 nM, respectively, which indicated that cortical and striatal M2 receptors mediate [3H]cAMP inhibition. The equilibrium dissociation constants for the full agonist carbachol in mediating these two biochemical responses were determined after partial receptor inactivation with propylbenzilylcholine mustard. The results indicate that cortical M1 receptor-mediated phosphoinositide metabolism is elicited by carbachol through a low affinity agonist-receptor complex (carbachol Kd = 90 microM). However, the cortical and striatal M2 receptor-mediated inhibition of [3H]cAMP is mediated by a high affinity agonist-receptor complex (carbachol Kd = 8.5 microM and 2.9 microM, respectively). Thus, the agonist is bound in a low affinity active conformation of the M1 receptor but the agonist is bound in a high affinity active conformation of the M2 receptor. In contrast to the cortical M1-phosphoinositide system, the central M2 receptors exhibited a significant receptor reserve in their mediation of [3H]cAMP inhibition, as elicited by the full agonist carbachol. Whereas the ratio of Kd/EC50 for carbachol was 0.9 at the cortical M1 receptor, this ratio was 9.4 and 3.2 at the cortical M2 and striatal M2 receptors, respectively.
The ability of 16 well-known cholinergic agonists to compete with 1 nM [3H]pirenzepine for the high (KH) and low (KL) affinity states of M1 muscarine receptors was studied, using rabbit hippocampal membranes suspended in 20 mM Tris buffer containing 1 mM MnCl2, at pH 7.4 and 25 degrees C. The hippocampus, like the cerebral cortex, has primarily m1 and m3 subtypes of M1 receptors; these receptors have very similar affinities for pirenzepine and for agonists, and both receptors are coupled to the activation of phospholipase C. KH values varied more than 10,000-fold, and did not correlate with prior measurements of the ability of agonists to promote cerebral excitation or phosphoinositide turnover in cortical tissue. KL values varied more than 1,000-fold, and also did not correlate with agonist activities. In contrast, KL/KH ratios for individual agonists varied from 66 for cis-dioxolane to near 1 for oxotremorine, and correlated closely with data concerning the relative physiological and biochemical effectiveness of the agonists. Each agonist showed only a single affinity in 0.2 mM guanyl-5'-yl imidodiphosphate. Thus binding measurements of KL/KH appear to be a new way to screen cholinergic agonists for relative efficacy at m1 or m3 receptors coupled to their native G proteins. Whereas both quaternary and tertiary agonists are known to activate M2 receptors, only the quaternary agents tested were highly effective M1 agonists. The five agonists which have been tested for improving human memory all appear to have low efficacy at M1 receptors.
Argiotoxin 636, (AR636), a synaptic antagonist from orb weaver spider venom, is shown to produce reversible blockade of excitatory transmission in CA1 pyramidal neurons of the in vitro rat hippocampus. Microtopical application of AR636 (5-50 nM) resulted in a concentration-dependent suppression of the amplitude of the dendritic field EPSP recorded from stratum radiatum, and the amplitude of the population spike recorded from stratum pyramidale in response to stimulation of the Schaffer collaterals. The maximum effect of AR636 occurred at about 15-25 min. These effects were reversible after washing with toxin-free physiological solution with the rate of recovery having an inverse relationship to the concentration of AR636. In contrast to the effects observed with orthodromic stimulation, the amplitude of the antidromic spike was not affected by exposure to AR636. The temporal pattern of GABAergic paired-pulse inhibition was unaffected by exposure to AR636. Neuronal discharge elicited by pressure ejection of L-glutamate was abolished by AR636, whereas, responses to L-aspartate were not significantly affected. These data suggest that AR636 functions as a selective antagonist of glutamate-mediated synaptic transmission in rat hippocampus.
Subtypes of muscarinic cholinergic receptors have been proposed to exist, but the biochemical responses mediated by the putative subtypes are unknown. In the present study, muscarinic receptor-mediated phosphoinositide breakdown and inhibition of adenylate cyclase activity were characterized in rat brain as well as rat parotid and heart. To study whether these responses are mediated by separate subtypes of muscarinic receptors, the potencies of agonists and antagonists were determined in both assays. Antagonist potencies were calculated by Schild analysis. In the brain, the putatively selective muscarinic receptor antagonist, pirenzepine, exhibited Ki values of 21 nM in the assay of phosphoinositide breakdown and 310 nM in the assay of adenylate cyclase activity. Similarly, using radioligand binding techniques, it distinguished two binding sites with Kd values of 12 and 168 nM. The antagonist, atropine, on the other hand, was equipotent in the two biochemical assays and the radioligand binding assay with Ki values of approximately 1 to 2 nM. In peripheral tissues with robust muscarinic receptor-mediated phosphoinositide (parotid) and adenylate cyclase (heart) responses, pirenzepine exhibited a similar selectivity (19-fold) for the phosphoinositide assay that was seen in the brain, but it was 6- to 7-fold less potent in both peripheral tissues than in the central nervous system. In addition, the potencies of pirenzepine in binding and functional studies in each tissue were not as well correlated as in the brain. Atropine and other antagonists were 4- to 9-fold selective for inhibiting oxotremorine-stimulated phosphoinositide breakdown in the peripheral tissues.(ABSTRACT TRUNCATED AT 250 WORDS)
The aim of the present study was to identify in the rat the overall input‐output pattern of connections of the primary auditory field, with special attention to the topographical organization of the geniculocortical auditory projection. By using cytoarchitectural criteria, three temporal cortical fields were distinguished in the rat: Te1, Te2, and Te3. The primary auditory field Te1 is characterized by a relatively specific differentiation of its layers when compared with other temporal fields. The afferent and efferent connections of Te1 were identified by using the retrograde and anterograde transport of wheat germ agglutinin conjugated with horseradish peroxidase (WGA‐HRP).
The results indicate that Te1 is connected by a dense and reciprocal system of fibers with the auditory thalamus. Based on the nomenclature of Morest ('64) in the cat, five cytoarchitectural subdivisions of the medial geniculate complex (MG) were identified in the rat: ventral (MGv), dorsal (MGd), medial (MGm), suprageniculate (Sg), and peripeduncular (PPA). The major rostrocaudal extent of the MGv is connected to Te1. The surrounding cortical fields Te2 and Te3 do not receive a projection from the MGv, except from its most caudal pole. The MGv projection is topographically organized. When the deposit area of the tracer is shifted from dorsal to ventral upon Te1, the corresponding labeled zone within the MGv moves from rostral to caudal, whereas a cortical displacement of the deposit area of the tracer from rostrodorsal to caudoventral leads to a medial to lateral shift of the labeled zone in the MGv. In addition, more dorsal parts of the MGv project on more dorsal sectors of Te1.
Te1 receives a sparser, topographically organized projection from the deep dorsal subdivision of the MGd. The MGm and the lateral part of the posterior group of thalamic nuclei (Pol) also distribute fibers to the primary auditory field.
Te1 is reciprocally connected by a system of callosal fibers with the contralateral homotypic cortex. Finally, Te1 sends fibers to the dorsal and, to a lesser extent, external cortices of the inferior colliculus, caudomedial caudateputamen complex, and caudoventral thalamic reticular nucleus.
Advances in understanding the phosphoinositide cycle have helped unravel the chain of events initiated by muscarinic receptor stimulation. Hydrolysis of membrane phosphoinositides generates both diacylglycerol, an activator of protein kinase C, and inositol phosphates. In the nervous system, muscarinic receptors elicit a wide range of electrophysiological responses. Recent studies have made progress in identifying which of these neuronal muscarinic actions are mediated by activation of protein kinase C. Paradoxically, protein kinase C also exerts a strong inhibitory influence on muscarinic responses. This complex set of actions suggests that in addition to mediating certain muscarinic responses, protein kinase C also blocks signal transduction as part of a feedback mechanism.
Muscarinic acetylcholine receptors (mAChRs), like many other neurotransmitter and hormone receptors, transduce agonist signals by activating G proteins to regulate ion channel activity and the generation of second messengers via the phosphoinositide (PI) and adenylyl cyclase systems. Human mAChRs are a family of at least four gene products which have distinct primary structures, ligand-binding properties and patterns of tissue-specific expression. To examine the question of whether functional differences exist between multiple receptor subtypes, we have investigated the ability of each subtype to regulate PI hydrolysis and adenylyl cyclase when expressed individually in a cell lacking endogenous mAChRs. We show that the HM2 and HM3 mAChRs efficiently inhibit adenylyl cyclase activity but poorly activate PI hydrolysis. In contrast, the HM1 and HM4 mAChRs strongly activate PI hydrolysis, but do not inhibit adenylyl cyclase, and in fact can substantially elevate cAMP levels. Interestingly, the subtypes that we find to be functionally similar are also more similar in sequence. Our results indicate that the different receptor subtypes are functionally specialized.