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The hypothalamic paraventricular and lateral parabrachial nuclei receive collaterals from raphe nucleus neurons: A combined double retrograde and immunocytochemical study
Abstract
Retrograde tracer injections of fluorescein- and rhodamine-labelled latex microspheres centered in the parvicellular zone of the hypothalamic paraventricular nucleus and pontine lateral parabrachial nucleus revealed that 36% of the labelled neurons in the dorsal raphe nucleus send collaterals to both structures. These cells were organized in a well-distinguishable cluster within the dorsal raphe nucleus. By combining retrograde tracing with immunocytochemistry, it was found that less than 8% of the double-labelled cells stained positively for serotonin. Of the remaining raphe nuclei that were examined, only the median raphe nucleus contributed a minor nonserotoninergic projection to the paraventricular or lateral parabrachial nuclei. Few of the retrogradely labelled cells in the median raphe nucleus contained both tracers. These results suggest that nonserotoninergic and serotoninergic neurons in the dorsal raphe nucleus, via collateral branching, may simultaneously influence the activity of two central nervous system nuclei involved in autonomic control.
... The dorsal raphe nucleus mediates the CO 2 arousal reflex via its projection to the lateral parabrachial nucleus (Petrov et al., 1992;Smith et al., 2018;Kaur et al., 2020). Other projections, sometimes involving collateral fibers, target predominantly forebrain regions, including the lateral hypothalamic nucleus, bed nucleus of the stria terminalis, central amygdala, and vestibular nuclei (Vertes, 1991;Petrov et al., 1992;Halberstadt and Balaban, 2006;Vasudeva et al., 2011;McDevitt et al., 2014). ...
... The dorsal raphe nucleus mediates the CO 2 arousal reflex via its projection to the lateral parabrachial nucleus (Petrov et al., 1992;Smith et al., 2018;Kaur et al., 2020). Other projections, sometimes involving collateral fibers, target predominantly forebrain regions, including the lateral hypothalamic nucleus, bed nucleus of the stria terminalis, central amygdala, and vestibular nuclei (Vertes, 1991;Petrov et al., 1992;Halberstadt and Balaban, 2006;Vasudeva et al., 2011;McDevitt et al., 2014). Descending projections target the hypoglossal, trigeminal and facial motor nuclei Guo et al., 2020), retrotrapezoid nucleus (Rosin et al., 2006), locus coeruleus (Luppi et al., 1995;Schwarz et al., 2015), periaqueductal gray (Vertes, 1991), and PPTg (Vertes, 1991;Steininger et al., 1992; Figure 6B). ...
... The main ascending inputs to the lateral parabrachial nucleus come from the spinal cord and NTS (Palmiter, 2018). Other inputs come from the pre-Bötzinger and Bötzinger complexes (Tan et al., 2010;Yang and Feldman, 2018), Kölliker-Fuse nucleus (Song et al., 2012a;Geerling et al., 2017), rVRG (Holstege, 1989), NTS (Beckstead et al., 1980;Herbert et al., 1990;McGovern et al., 2015b;Kawai, 2018;Yu et al., 2022), retrotrapezoid nucleus (Rosin et al., 2006;Bochorishvili et al., 2012;Silva et al., 2016a), locus coeruleus Yang et al., 2021), dorsal raphe (Petrov et al., 1992;Quattrochi et al., 1998;Kaur et al., 2020), bed nucleus of stria terminalis , central amygdala Yang et al., 2021), paraventricular nucleus (Zheng et al., 1995;Geerling et al., 2010;Singh et al., 2022), spinal trigeminal nucleus (Panneton et al., 2006;Zhang et al., 2018), paratrigeminal nucleus (Saxon and Hopkins, 1998;Caous et al., 2001;McGovern et al., 2015b;Driessen et al., 2018), PPTg (Quattrochi et al., 1998;Lima et al., 2019b), and fastigial nucleus (Teune et al., 2000). ...
Respiration is a brain function on which our lives essentially depend. Control of respiration ensures that the frequency and depth of breathing adapt continuously to metabolic needs. In addition, the respiratory control network of the brain has to organize muscular synergies that integrate ventilation with posture and body movement. Finally, respiration is coupled to cardiovascular function and emotion. Here, we argue that the brain can handle this all by integrating a brainstem central pattern generator circuit in a larger network that also comprises the cerebellum. Although currently not generally recognized as a respiratory control center, the cerebellum is well known for its coordinating and modulating role in motor behavior, as well as for its role in the autonomic nervous system. In this review, we discuss the role of brain regions involved in the control of respiration, and their anatomical and functional interactions. We discuss how sensory feedback can result in adaptation of respiration, and how these mechanisms can be compromised by various neurological and psychological disorders. Finally, we demonstrate how the respiratory pattern generators are part of a larger and integrated network of respiratory brain regions.
... with reported values ranging from 15% to >97% (O'Hearn and Molliver, 1984;Beitz et al., 1986;Ma et al., 1991;Petrov et al., 1992;Van Bockstaele et al., 1993;Datiche et al., 1995;Kirifides et al., 2001;Kim et al., 2004). It is known that the supramammillary nucleus (Kiss et al., 2002) and the inferior colliculus (Schwarz and Schwarz, 1992) are targeted by projections arising from glutamatergic DRN cells. ...
... Anterograde tracing studies have demonstrated that the DRN projects to PBN (Sim and Joseph, 1993;Vertes and Kocsis, 1994), PVN (Vertes, 1991;Sim and Joseph, 1993;Larsen et al., 1996;Morin and Meyer-Bernstein, 1999), CeA (Vertes, 1991;Sim and Joseph, 1993;Morin and Meyer-Bernstein, 1999;Commons et al., 2003), and the insular and infralimbic cortices (Vertes, 1991;Morin and Meyer-Bernstein, 1999). These findings have been confirmed using retrograde tracing (Imai et al., 1986;Seroogy and Fallon, 1989;Ma et al., 1991;Uryu et al., 1992;Petrov et al., 1992Petrov et al., , 1994Li et al., 1993;Champagne et al., 1998;Hasue and Shammah-Lagnado, 2002), and the projections were found to originate from both serotonergic and nonserotonergic neurons. Numerous retrograde tracing studies have reported that individual DRN neurons send axon collaterals to two or more functionally related target structures (van der Kooy and Kuypers, 1979;van der Kooy and Hattori, 1980;de Olmos & Heimer, 1980;Imai et al., 1986;Van Boeckstaele et al., 1993;Kirifides et al., 2001). ...
... Interestingly, components of the vestibulo-parabrachial network are already known to be targets of collateralized DRN projections. For example, DRN cells send branching collaterals to the lateral PBN and the PVN (Petrov et al., 1992), and Petrov and colleagues (Petrov et al., 1994) have reported that the DRN is a source of collateralized projections to the CeA and the PVN. Importantly, preliminary retrograde tracing studies in our laboratory indicate that individual DRN cells send collateralized projections to CeA and the vestibular nuclei (Halberstadt et al., 2001). ...
Preclinical and clinical evidence indicates that the serotonergic system regulates processing in the vestibular nuclei and in pathways linking balance function with emotional responses and affect. Previous studies conducted in this laboratory demonstrated that the serotonergic innervation of the vestibular nuclei is derived largely from the dorsal raphe nucleus (DRN), and revealed that the DRN also sends a nonserotonergic projection to the vestibular nuclei. The purpose of these experiments was to characterize the organization of the serotonergic and nonserotonergic components of the DRN projection to the vestibular nuclei. In Chapter 3, we describe retrograde tracing experiments that examined whether DRN cells send collateralized projections to the vestibular nuclei and central amygdaloid nucleus (CeA), regions involved in the clinical linkage between disorders of balance control and anxiety, and concluded that a subset of the serotonergic and nonserotonergic projections to the vestibular nuclei also project to CeA.Chapter 4 describes experiments with the anterograde tracer biotinylated dextran amine (BDA) that identified the terminal distribution of DRN projections within the vestibular nuclei. This study revealed that DRN projections descending in the ventricular plexus and the medial longitudinal fasciculus terminate within distinct vestibular terminal fields. In Chapter 5, BDA was used in combination with the serotonergic neurotoxin 5,7-dihydroxytryptamine (5,7-DHT) to selectively anterogradely trace nonserotonergic DRN projections to the vestibular nuclei. These experiments demonstrated that nonserotonergic DRN projections descend exclusively within the ventricular plexus and terminate primarily within the periventricular aspect of the vestibular nuclei.The purpose of the experiments in Chapter 6 was to map the distribution of serotonergic DRN terminals within the vestibular nuclei; 5,7-DHT was injected directly into DRN and silver staining was used to visualize the resulting pattern of terminal degeneration. It appears that projections from serotonergic DRN neurons terminate within both medial and lateral regions of the vestibular nuclei.Based on these findings, we conclude that major differences exist in the course of descent and termination patterns of serotonergic and nonserotonergic DRN projections to the vestibular nuclei, indicating that serotonergic and nonserotonergic cells give rise to distinct DRN projection systems that may selectively modulate processing within specific functional domains of the vestibular nuclei.
... SERT is the target of serotonin selective reuptake inhibitors (SSRIs), and acute administration increases serotonin release preferentially in the raphe nuclei [71][72][73][74]. Genetic ablation of SERT results in elevated anxiety-like behavior in adult mice, an effect which is modulated by differences in genetic background [75] and is accompanied by fivefold increases in extracellular 5-HT concentrations [76], desensitization and downregulation of 5-HT 1A Rs in the raphe, hypothalamus, septum, and amygdala [77], 5-HT 1A R-dependent reduction in spontaneous firing rates in 5-HTergic neurons in the DRN [78], downregulation of 5-HT 1B Rs in serotonergic axon terminals [79], and disorganization of vibrissae-stimulated barrel fields in the somatosensory cortex [80]. ...
... Exposure to an open field under low-light conditions increases c-Fos-like immunoreactivity in the BLA of rats, while high-light conditions selectively increase c-Fos-like immunoreactivity in the posterior and ventromedial portions of the BLA [71], as well as in ventral hippocampal [72] and dorsomedial and caudodorsal raphe cells [73] which project to the basolateral amygdala. The administration of anxiogenic drugs with multiple mechanisms (the 5-HT 2 R non-selective agonist m-chlorophenyl piperazine [mCPP], the adenosine non-selective antagonist caffeine, the a 2 -adrenoceptor antagonist yohimbine, and the partial benzodiazepine receptor inverse agonist N-methyl-beta-carboline-3-carboxamide [FG-7142]) increase c-Foslike immunoreactivity in parvalbumin-positive GABAergic neurons of the BLA [74]. Activation of the BLA has also been observed in mice receiving aversive ultrasound in the home cage, and this Fos expression is not changed when footshocks are administered before ultrasound exposure [35]. ...
... Exposure of rats to an open field elicits c-Fos-like expression in diverse neurons of the BLA [71,72], including neurons which receive projections from the ventral hippocampus [72] and from the DRN [73], and inescapable stress and microinjection of the CRF 2 R agonist urocortin 2 in the DRN increase serotonin release in the BLA [86,87]. Anxiogenic drugs with different molecular targets (mCPP, caffeine, yohimbine, FG-7142) activate GABAergic interneurons in the BLA which expresses parvalbumin [74]. These interneurons express high levels of the 5-HT 2A receptor [88,89]; the increased serotonergic efflux in the BLA, thus, leads to increased GABA release by these interneurons [89,90]. ...
While anxiogenic stimuli activate neurons in the raphe and lead to serotonin release in limbic forebrain targets, panicogenic stimuli do not necessarily do so. For example, escape performance in an elevated T-maze does not increase c-Fos-like immunoreactivity in the raphe. Nonetheless, the observation of a panicolytic role of serotonin in the PAG and an anxiogenic role in the amygdala and hippocampus suggests that the raphe is not a homogeneous structure. In fact, the dorsal raphe can be divided at least into six subregions based on cytoarchitecture and distribution of serotonergic neurons. These comprise the rostral (DRr), dorsal (DRD), ventral (DRV), lateral wing (lwDR), caudal (DRC), and interfascicular (DRI) portions (Fig. 5.1). Among those, the rostral, ventral, and interfascicular subregions play little role in the control of defense responses, and discussion of their functions can be found elsewhere. Here, we will discuss evidence for a role of the DRD, lwDR and DRC in anxiety and fear.
... However, the serotonin neurons and their neuronal targets mediating the effect of the serotonin system on CO 2 arousal have not been identified. In this regard, DR neurons are known to project to the lateral PB [14][15][16] , are CO 2 responsive, and injecting the DR with low pH (acidic) solution causes wakefulness in wildtype mice, but not in mice lacking serotonin neurons 17 . Based on the proposed role of the DR serotonergic neurons in CO 2 chemosensitivity, we hypothesized that the DR may supply a serotonergic input to the PBel CGRP neurons via 5HT 2a receptors to sensitize them to CO 2 -responsive inputs. ...
... However, deletion of medullary serotonin neurons in Egr2-Pet1 mice reduces the ventilatory response to CO 2 , suggesting that the medullary serotonin neurons innervate and facilitate the medullary cell groups that promote ventilation, but not the ascending arousal system. Because both medullary and DR serotonin neurons are known to be CO 2 responsive 8,9 , and DR neurons were previously found to target lateral PB [14][15][16] , it therefore seemed likely that the DR neurons may contribute to CO 2 arousal by means of their projections to the PB. A recent study supported this idea by showing that stimulation of the DR by microinjection of acidic cerebrospinal fluid woke up wild-type mice, but not Lmx1b l/fl/p ::ePet1-Cre mice that lack serotonin neurons 17 . ...
During obstructive sleep apnea, elevation of CO 2 during apneas contributes to awakening and restoring airway patency. We previously found that glutamatergic neurons in the external lateral parabrachial nucleus (PBel) containing calcitonin gene related peptide (PBel CGRP neurons) are critical for causing arousal during hypercapnia. However, others found that genetic deletion of serotonin (5HT) neurons in the brainstem also prevented arousal from hypercapnia. To examine interactions between the two systems, we showed that dorsal raphe (DR) 5HT neurons selectively targeted the PBel. Either genetically directed deletion or acute optogenetic silencing of DR Sert neurons dramatically increased the latency of mice to arouse during hypercapnia, as did silencing DR Sert terminals in the PBel. This effect was mediated by 5HT 2a receptors which are expressed by PBel CGRP neurons. Our results indicate that the serotonergic input from the DR to the PBel via 5HT 2a receptors is critical for modulating the sensitivity of the PBel CGRP neurons that cause arousal to rising levels of blood CO 2 .
... However, the serotonin neurons and their neuronal targets mediating the effect of the serotonin system on CO 2 arousal have not been identified. In this regard, DR neurons are known to project to the lateral PB [14][15][16] , are CO 2 responsive, and injecting the DR with low pH (acidic) solution causes wakefulness in wildtype mice, but not in mice lacking serotonin neurons 17 . Based on the proposed role of the DR serotonergic neurons in CO 2 chemosensitivity, we hypothesized that the DR may supply a serotonergic input to the PBel CGRP neurons via 5HT 2a receptors to sensitize them to CO 2 -responsive inputs. ...
... However, deletion of medullary serotonin neurons in Egr2-Pet1 mice reduces the ventilatory response to CO 2 , suggesting that the medullary serotonin neurons innervate and facilitate the medullary cell groups that promote ventilation, but not the ascending arousal system. Because both medullary and DR serotonin neurons are known to be CO 2 responsive 8,9 , and DR neurons were previously found to target lateral PB [14][15][16] , it therefore seemed likely that the DR neurons may contribute to CO 2 arousal by means of their projections to the PB. A recent study supported this idea by showing that stimulation of the DR by microinjection of acidic cerebrospinal fluid woke up wild-type mice, but not Lmx1b l/fl/p ::ePet1-Cre mice that lack serotonin neurons 17 . ...
Introduction
Serotoninergic dorsal raphe neurons (DRSert) are CO2 responsive, and mice lacking serotonin have impaired arousal to CO2. We showed that the neurons in external lateral parabrachial nucleus containing calcitonin gene related peptide (PBelCGRP), are required for CO2-arousal. PBelCGRP neurons also receive serotoninergic innervation from the DRSert. 5HT2A agonist restores CO2 responsiveness in mice lacking serotonin, suggesting that DRSert may modulate CO2 arousal by acting on 5HT2A receptors possibly on the PBel neurons.
Methods
We used serotonin transporter (Sert)-Cre mice to optogenetically inhibit DRSert neurons and their terminals in the PBel. We injected AAV-FLEX-ArchT into the DR and implanted an optical fiber just above it in one set of Sert-Cre mice and bilaterally in the PBel in another set. All mice were instrumented for sleep and optogenetics and were tested for EEG arousals to 10% CO2. Latencies of arousal were compared with optogenetic inhibition of either the DR neurons or their terminals in the PBel with a 593nm laser light. We further tested whether a 5HT2A agonist (TCB-2) can reverse blockade of CO2 arousal in mice where DRSert terminals in PBel were inhibited. Finally, TCB-2 was injected in mice with PBelCGRP deletions and arousal latency to CO2 was compared.
Results
Compared to the control (Laser-OFF) condition, arousal latency to CO2 was significantly increased by photoinhibition of either the DRSert neurons (n=6; latency- 40.9 ± 6.4 vs. 13.81± 0.69 sec; F3, 17= 11.5; P< 0.001) or their terminals in PBel (n=8; latency-34.9 ± 2.3 sec vs. 16.62 ± 0.97sec, F1, 14= 56.9; P< 0.001). This was reversed by the 5HT2A agonist TCB-2 (5mg/kg), as it reduced the latency to CO2 arousal in mice with photoinhibition of terminals in PBel from 35.48 ± 7.31 sec to 16.24 ± 1.06 sec (F3, 9= 8.05; P= 0.006), but had no effect in mice with PBelCGRP neurons deletions.
Conclusion
The serotonin system modulate CO2-arousals by the DRSert input to the PBel. TCB-2 reversed the effect of inhibition of DRSert terminals in the PBel, but not in mice with PBelCGRP deletions, suggests that DRSert modulate PBelCGRP neurons through 5HT2a receptors.
Support
NIH- 2P01 HL095491 and NS112175
... The PBN receives taste and visceral inputs relayed through the NTS. Interestingly, it also has been shown to receive serotonergic innervation from the dorsal raphe (Petrov et al., 1992 ). This is unusual, since the vast majority of projections from the dorsal raphe are to the forebrain. ...
... This is unusual, since the vast majority of projections from the dorsal raphe are to the forebrain. Moreover, a substantial number of dorsal raphe neurons were reported to send collateral projections to both the PBN and the PVH in the hypothalamus (Petrov et al., 1992). Neurons of the PBN express both 5HT2CR and 5HT1BR (Bruinvels et al., 1993; Wright et al., 1995). ...
Maintenance of energy balance requires regulation of the amount and timing of food intake. Decades of experiments utilizing pharmacological and later genetic manipulations have demonstrated the importance of serotonin signaling in this regulation. Much progress has been made in recent years in understanding how central nervous system (CNS) serotonin systems acting through a diverse array of serotonin receptors impact feeding behavior and metabolism. Particular attention has been paid to mechanisms through which serotonin impacts energy balance pathways within the hypothalamus. How upstream factors relevant to energy balance regulate the release of hypothalamic serotonin is less clear, but work addressing this issue is underway. Generally, investigation into the central serotonergic regulation of energy balance has had a predominantly “hypothalamocentric” focus, yet non-hypothalamic structures that have been implicated in energy balance regulation also receive serotonergic innervation and express multiple subtypes of serotonin receptors. Moreover, there is a growing appreciation of the diverse mechanisms through which peripheral serotonin impacts energy balance regulation. Clearly, the serotonergic regulation of energy balance is a field characterized by both rapid advances and by an extensive and diverse set of central and peripheral mechanisms yet to be delineated.
... The RN is known to project to the PVN. [76][77][78][79] We carried out a series of experiments to study the functional importance of serotonergic projection from the RN in the PVN. (A) Unlike OXT +/+ (n = 15) rat pups, saline-treated OXT À/À (n = 33) rat pups showed no preference for maternal bedding over other dams' bedding. ...
Maternal affiliation by infants is the first social behavior of mammalian animals. We report here that elimination of the Tph2 gene essential for serotonin synthesis in the brain reduced affiliation in mice, rats, and monkeys. Calcium imaging and c-fos immunostaining showed maternal odors activation of serotonergic neurons in the raphe nuclei (RNs) and oxytocinergic neurons in the paraventricular nucleus (PVN). Genetic elimination of oxytocin (OXT) or its receptor reduced maternal preference. OXT rescued maternal preference in mouse and monkey infants lacking serotonin. Tph2 elimination from RN serotonergic neurons innervating PVN reduced maternal preference. Reduced maternal preference after inhibiting serotonergic neurons was rescued by oxytocinergic neuronal activation. Our genetic studies reveal a role for serotonin in affiliation conserved from mice and rats to monkeys, while electrophysiological, pharmacological, chemogenetic, and optogenetic studies uncover OXT downstream of serotonin. We suggest serotonin as the master regulator upstream of neuropeptides in mammalian social behaviors.
... Previous studies have reported that DR 5-HT neurons project to both lateral geniculate body and superior colliculus with collateral axons for processing visual information [46]. The collateral 5-HT branching was also found to innervate both paraventricular nucleus of the hypothalamus (PVN) and lateral parabrachial nucleus (PBN) for autonomic control [47]. Using retrograde AAV for neural tracing, we found both DR and MnR 5-HT neurons sent projections to ZIR. ...
Objective
Rostral zona incerta (ZIR) evokes feeding by sending GABA transmission to paraventricular thalamus (PVT). Although central serotonin (5-HT) signaling is known to play critical roles in the regulation of food intake and eating disorders, it remains unknown whether raphe 5-HT neurons functionally innervate ZIR-PVT neural pathway for feeding control. Here, we sought to reveal how raphe 5-HT signaling regulates both ZIR and PVT for feeding control.
Methods
We used retrograde neural tracers to map 5-HT projections in Sert-Cre mice and slice electrophysiology to examine the mechanism by which 5-HT modulates ZIR GABA neurons. We also used optogenetics to test the effects of raphe-ZIR and raphe-PVT 5-HT projections on feeding motivation and food intake in mice regularly fed, 24 h fasted, and with intermittent high-fat high-sugar (HFHS) diet. In addition, we applied RNAscope in situ hybridization to identify 5-HT receptor subtype mRNA in ZIR.
Results
We show raphe 5-HT neurons sent projections to both ZIR and PVT with partial collateral axons. Photostimulation of 5-HT projections inhibited ZIR but excited PVT neurons to decrease motivated food consumption. However, both acute food deprivation and intermittent HFHS diet downregulated 5-HT inhibition on ZIR GABA neurons, abolishing the inhibitory regulation of raphe-ZIR 5-HT projections on feeding motivation and food intake. Furthermore, we found high-level 5-HT1a and 5-HT2c as well as low-level 5-HT7 mRNA expression in ZIR. Intermittent HFHS diet increased 5-HT7 but not 5-HT1a or 5-HT2c mRNA levels in the ZIR.
Conclusions
Our results reveal that raphe-ZIR 5-HT projections dynamically regulate ZIR GABA neurons for feeding control, supporting that a dynamic fluctuation of ZIR 5-HT inhibition authorizes daily food intake but a sustained change of ZIR 5-HT signaling leads to overeating induced by HFHS diet.
... One outlier in the control group and 2 outliers in the DRN-TPH2-KO group were detected in (I), but were kept in the dataset as presented. Abbreviations: DRN, dorsal Raphe nuclei; DRN-TPH2-KO, TPH2 knockout in the DRN; SEM, standard error of the mean; TPH2, tryptophan hydroxylase 2.Tryptophan hydroxylase 2 deletion in the DRN decreases c-fos expression in hypothalamic POMC neurons 5-hydroxytryptamine neurons in the DRN send dense projections to the forebrain, including various nuclei in the hypothalamus(45,(59)(60)(61). Of note, POMC neurons in the arcuate nucleus of the hypothalamus (ARH) play a well-established role in mediating the anorexigenic actions of 5-HT(58, 62, 63). ...
Central 5-hydroxytryptamine (5-HT), which is primarily synthesized by tryptophan hydroxylase 2 (TPH2) in the dorsal Raphe nuclei (DRN), plays a pivotal role in the regulation of food intake and body weight. However, the physiological functions of TPH2 on energy balance have not been consistently demonstrated. Here we systematically investigated the effects of TPH2 on energy homeostasis in adult male and female mice. We found that the DRN harbors similar amount of TPH2 + cells in control male and female mice. Adult-onset TPH2 deletion in the DRN promotes hyperphagia and body weight gain only in male mice, but not in female mice. Ablation of TPH2 reduces hypothalamic pro-opiomelanocortin (POMC) neuronal activity robustly in males but only to a modest degree in females. Deprivation of estrogen by ovariectomy (OVX) causes comparable food intake and weight gain in female control and DRN-TPH2-KO mice. Nevertheless, disruption of TPH2 blunts the anorexigenic effects of exogenous E2 and abolishes E2-induced activation of POMC neurons in OVX female mice, indicating that TPH2 is indispensable for E2 to activate POMC neurons and to suppress appetite. Together, our study revealed that TPH2 in the DRN contributes to energy balance regulation in a sexually dimorphic manner.
... The decrease in sodium appetite during hypovolemia, by conditionally knocking out Htr2c from the LPBN (LPBN Htr2cKO mice), suggests that ECF volume may regulate serotonergic signaling in the LPBN, which in turn controls sodium intake by modulation of LPBN Htr2c neuronal activity. We found serotonergic projections to the LPBN from the DR and MnR, consistent with other reports 37,38 . The MnR is of particular interest because electrical stimulation of MnR has been shown to increase blood pressure, which was attenuated in the presence of an inhibitor of tryptophan hydroxylase 39 . ...
Sodium appetite is a powerful form of motivation that can drive ingestion of high, yet aversive concentrations of sodium in animals that are depleted of sodium. However, in normal conditions, sodium appetite is suppressed to prevent homeostatic deviations. Although molecular and neural mechanisms underlying the stimulation of sodium appetite have received much attention recently, mechanisms that inhibit sodium appetite remain largely obscure. Here we report that serotonin 2c receptor (Htr2c)-expressing neurons in the lateral parabrachial nucleus (LPBNHtr2c neurons) inhibit sodium appetite. Activity of these neurons is regulated by bodily sodium content, and their activation can rapidly suppress sodium intake. Conversely, inhibition of these neurons specifically drives sodium appetite, even during euvolemic conditions. Notably, the physiological role of Htr2c expressed by LPBN neurons is to disinhibit sodium appetite. Our results suggest that LPBNHtr2c neurons act as a brake against sodium appetite and that their alleviation is required for the full manifestation of sodium appetite. High concentrations of sodium are normally unpalatable. This study shows a neural population in the brainstem that suppresses appetite for sodium. Reducing the activity of these neurons can drive ingestion of high concentrations of sodium.
... Serotonin activity in the PVN has been shown to be a positive modulator of HPA axis responses (Feldman et al., 2000;Pan and Gilbert, 1992). Similarly to cholinergic innervation, the PVN is relatively devoid of serotonergic terminals, with only a small contribution of ascending afferents from the raphe nuclei (Petrov et al., 1992;Swanson and Sawchenko, 1983). There is a much higher density of serotonergic fibers innervating the neutrophil immediately outside of the PVN . ...
The stress response—originally described by Hans Selye as “the nonspecific response of the body to any demand made upon it”—is chiefly mediated by the hypothalamic-pituitary-adrenal (HPA) axis and is activated by diverse sensory stimuli that inform threats to homeostasis. The diversity of signals regulating the HPA axis is partly achieved by the complexity of afferent inputs that converge at the apex of the HPA axis: this apex is formed by a group of neurosecretory neurons that synthesize corticotropin-releasing hormone (CRH) in the paraventricular nucleus of the hypothalamus (PVN). The afferent synaptic inputs onto these PVN-CRH neurons originate from a number of brain areas, and PVN-CRH neurons respond to a long list of neurotransmitters/neuropeptides. Considering this complexity, an important question is how these diverse afferent signals independently and/or in concert influence the excitability of PVN-CRH neurons. While many of these inputs directly act on the postsynaptic PVN-CRH neurons for the summation of signals, accumulating data indicates that they also modulate each other's transmission in the PVN. This mode of transmission, termed heterosynaptic modulation, points to mechanisms through which the activity of a specific modulatory input (conveying a specific sensory signal) can up- or down-regulate the efficacy of other afferent synapses (mediating other stress modalities) depending on receptor expression for and spatial proximity to the heterosynaptic signals. Here, we review examples of heterosynaptic modulation in the PVN and discuss its potential role in the regulation of PVN-CRH neurons' excitability and resulting HPA axis activity.
This article is part of the Special Issue entitled ‘Hypothalamic Control of Homeostasis’.
... Virtually all brain nuclei implicated in energy-balance regulation receive serotonergic afferents, including the PVH, dorsomedial hypothalamus, and lateral hypothalamus. [330][331][332][333] Food restriction decreases serotonin levels in the hypothalamus. 241 In turn, serotonin decreases food intake in humans and rodents, whether it is given systemically or centrally. ...
Anorexia nervosa (AN) is a psychiatric illness with minimal effective treatments and a very high rate of mortality. Understanding the neurobiological underpinnings of the disease is imperative for improving outcomes and can be aided by the study of animal models. The activity-based anorexia rodent model (ABA) is the current best parallel for the study of AN. This review describes the basic neurobiology of feeding and hyperactivity seen in both ABA and AN, and compiles the research on the role that stress-response and reward pathways play in modulating the homeostatic drive to eat and to expend energy, which become dysfunctional in ABA and AN.
... The 5-HT system is a known activator of the HPA axis, where selective SSRIs and 5-HT agonists increase corticosterone production during and independent of stress (Heisler et al., 2007). While direct innervation of the PVN has been reported (Petrov et al., 1992(Petrov et al., , 1994Williamson and Viau, 2007), 5-HTergic fibers from the DR also heavily innervate the GABAergic neurons of the PVN-surround (Sawchenko et al., 1983). Therefore, CRF-mediated changes in 5-HT output could modulate this axis through a disinhibition of medial parvocellular neurons. ...
Neuropsychiatric diseases represent a major public health burden worldwide; due to gaps in our understanding of the pathogenic mechanisms of disease, approximately 30% of patients are refractory to treatment. Activation of neuroinflammatory signaling cascades has shown promise as a contributing factor for disease development. This hypothesis is driven in part by an intriguing overlap of symptoms in patients with neuropsychiatric and immunological disorders. Patients with depression, bipolar disorder, schizophrenia, and autism commonly present with immune dysfunction, while patients with multiple sclerosis, lupus, and rheumatoid arthritis often experience severe mood disturbances. Diversity in presentation of symptoms, however, has posed a research challenge to our mechanistic understanding of this link.^ In contrast to the complexity of modeling specific diseases, altered sensitivity to stress is a well-documented vulnerability marker across neuropsychiatric disorders. Of relevance to clinical advancement, aspects of stress behavior and physiology can be modeled and measured in animals, where core components of the stress axis are conserved in humans and rodents.^ Thus, we performed an examination of the neuroinflammatory regulation of stress behavior and physiology. Using a genetic model of stress sensitivity, we report the discovery that anti-inflammatory treatment ameliorates hypothalamic-pituitary-adrenal axis dysregulation, identifying the dorsal raphe (DR) as a locus of heightened responsivity. We then demonstrated sex differences in this brain region in response to the stress neuropeptide, corticotropin-releasing factor, suggesting that differences in its responsivity may underlie sex differences in vulnerability to stress-related disorders. Finally, we used a transgenic approach to show that neuroinflammation localized specifically to the DR results in dysregulated stress behavior and physiology through interactions with the serotonergic neurotransmitter system.^ Overall, this work demonstrates that hyper- or hypo-function of the DR, based on genetic susceptibility, sex, or neuroinflammatory insult, can result in altered stress physiology and behavior. Though the DR has previously been identified as a potential locus of dysregulation, here we establish the specific, mechanistic link between risk factors for stress-related disorders. We present evidence of quantitative changes to this brain region and its functional output, and demonstrate that differences in responsivity of the DR may underlie vulnerability to stress-related disorders.
... Conversely, activation of CRH1 receptors in the LC increases the discharge activity of NE neurons (Page and Abercrombie, 1999;Jedema and Grace, 2004). Similarly, DRN 5-HT neurons project to the PVN (Petrov et al., 1992) and activate the HPA axis through 5-HT 1A and 5-HT 2 receptors (Zhang et al., 2004;Osei-Owusu et al., 2005;Heisler et al., 2007) while conversely, CORT tonically inhibits 5-HT 1A receptor expression (Laaris et al., 1995;Wissink et al., 2000). ...
... An increase in EDA results from slight increases in eccrine sweat gland activity which is directly controlled by the hypothalamus (Davison and Koss, 1975;Shibasaki and Crandall, 2010). Since most areas of the hypothalamus receive serotonergic inputs from the DRN (Sawchenko et al., 1983;Willoughby and Blessing, 1987;Petrov et al., 1992), it is therefore also possible that increases in EDA during flow were conditioned by rather direct modulations of hypothalamic influences on sympathetic sudomotor nerve activation. ...
Previously, experimentally induced flow experiences have been demonstrated with perfusion imaging during activation blocks of three minutes length to accommodate with the putatively slowly evolving "mood" characteristics of flow (Ulrich et al., 2014). Here, we used functional magnetic resonance imaging (fMRI) in a sample of 23 healthy, male participants to investigate flow in the context of a typical fMRI block design with block lengths as short as 30 s. To induce flow, demands of arithmetic tasks were automatically and continuously adjusted to the individual skill level. Compared against conditions of boredom and overload, experience of flow was evident from individuals' reported subjective experiences and changes in electrodermal activity. Neural activation was relatively increased during flow, particularly in the anterior insula, inferior frontal gyri, basal ganglia, and midbrain. Relative activation decreases during flow were observed in medial prefrontal and posterior cingulate cortex, and in the medial temporal lobe including the amygdala. Present findings suggest that even in the context of comparably short activation blocks flow can be reliably experienced and is associated with changes in neural activation of brain regions previously described. Possible mechanisms of interacting brain regions are outlined awaiting further investigation which should now be possible given the greater temporal resolution compared to previous perfusion imaging.
... In the brain, serotonin (5-hydroxytryptamine (5-HT)) is produced in the dorsal and medial raphe nucleus (DRN and MRN, respectively) with widespread projections throughout the brain including several nuclei within the hypothalamus (Sawchenko et al., 1983;Willoughby and Blessing, 1987;Petrov et al., 1992). Serotonin is synthesized from the essential amino acid tryptophan which is hydroxylated by tryptophan hydroxylase (TPH) to 5-hydroxytryptophan (5-HTP), which in its turn is decarboxylated to form serotonin (5-HT). ...
It is evident that eating in misalignment with the biological clock (such as in shift work, eating late at night and skipping breakfast) is associated with increased risk for obesity and diabetes. The biological clock located in the suprachiasmatic nucleus dictates energy balance including feeding behavior and glucose metabolism. Besides eating and sleeping patterns, glucose metabolism also exhibits clear diurnal variations with higher blood glucose concentrations, glucose tolerance and insulin sensitivity prior to waking up. The daily variation in plasma glucose concentrations in rats, is independent of the rhythm in feeding behavior. On the other hand, feeding itself has profound effects on glucose metabolism, but differential effects occur depending on the time of day. We here review data showing that a disturbed diurnal eating pattern results in alterations in glucose metabolism induced by a disrupted circadian clock. We first describe the role of central serotonin on feeding behavior and glucose metabolism and subsequently describe the effects of central serotonin on the circadian system. We next explore the interaction between the serotonergic system and the circadian clock in conditions of disrupted diurnal rhythms in feeding and how this might be involved in the metabolic dysregulation that occurs with chronodisruption.
Copyright © 2015. Published by Elsevier Ltd.
... ). Connectional studies also demonstrate that the 87 LPBN is monosynaptically connected with the AP and 88 dorsal raphe nucleus (DRN) (Lanca and van der Kooy 89 1985;Petrov et al., 1992). Serotonin from these nuclei90 can be released into the LPBN and, in turn, can facilitate 91 this inhibitory pathway, related to the modulation of 92 sodium intake and excretion (Lanca and van der Kooy, 93 1985; Godino et al., 2010). ...
Our aim was to analyze the participation of inhibitory and stimulatory signals in the temporal dissociation between sodium depletion (SD) induced by peritoneal dialysis (PD) and the appearance of sodium appetite (SA), particularly 2h after PD, when the rats are hypovolemic/natremic but SA is not evident. We investigated the effects of bilateral injections of the serotonin (5-HT) receptor antagonist, methysergide, into the lateral parabrachial nucleus (LPBN) on hypertonic NaCl and water intake 2h vs 24h after PD. We also studied plasma renin activity (PRA) and aldosterone concentration 2h vs 24h after PD. Additionally, we combined the analysis of brain Fos-ir with the detection of double immunoreactivity in 5HT and oxytocinergic (OT) cells 2 h after PD. Bilateral LPBN injections of methysergide (4 μg/200 nl at each site) increased NaCl intake when tested 2h after PD compared to controls. We found a significant increase in PRA and aldosterone concentration after PD but no differences between 2 h and 24h after PD. We also found for the first time a significant increase 2h after PD in the number of Fos-ir neurons in the brainstem nuclei that have been shown to be involved in the inhibition of SA. In summary, the results show that 5HT-mechanisms in the LPBN modulate sodium intake during the delay of SA when the renin angiotensin aldosterone system (RAAS) is increased. In addition, the activation of brainstem areas previously associated with the satiety phase of SA is in part responsible for the temporal dissociation between SD and behavioral arousal.
Copyright © 2015. Published by Elsevier Ltd.
... However, lesions of LC do reduce ACTH and corticosterone responses to acute stress (Ziegler et al. 1999), and there are suggestions that LC may impact PVN indirectly via limbic structures [reviewed in (Herman and Cullinan 1997)]. DRN has collateral serotonergic projections to CNA and PVN (Petrov et al. 1992(Petrov et al. , 1994, and 5-HT agonists enhance PVN activity as indicated by increased corticosterone levels and Fos expression (Mikkelsen et al. 2004). Indirect pathways may also play a role in serotonergic regulation of PVN (Herman and Cullinan 1997). ...
Stress is considered to be an important cause of disrupted sleep and insomnia. However, controlled and experimental studies in rodents indicate that effects of stress on sleep-wake regulation are complex and may strongly depend on the nature of the stressor. While most stressors are associated with at least a brief period of arousal and wakefulness, the subsequent amount and architecture of recovery sleep can vary dramatically across conditions even though classical markers of acute stress such as corticosterone are virtually the same. Sleep after stress appears to be highly influenced by situational variables including whether the stressor was controllable and/or predictable, whether the individual had the possibility to learn and adapt, and by the relative resilience and vulnerability of the individual experiencing stress. There are multiple brain regions and neurochemical systems linking stress and sleep, and the specific balance and interactions between these systems may ultimately determine the alterations in sleep-wake architecture. Factors that appear to play an important role in stress-induced wakefulness and sleep changes include various monominergic neurotransmitters, hypocretins, corticotropin releasing factor, and prolactin. In addition to the brain regions directly involved in stress responses such as the hypothalamus, the locus coeruleus, and the amygdala, differential effects of stressor controllability on behavior and sleep may be mediated by the medial prefrontal cortex. These various brain regions interact and influence each other and in turn affect the activity of sleep-wake controlling centers in the brain. Also, these regions likely play significant roles in memory processes and participate in the way stressful memories may affect arousal and sleep. Finally, stress-induced changes in sleep-architecture may affect sleep-related neuronal plasticity processes and thereby contribute to cognitive dysfunction and psychiatric disorders.
... Neuroanatomical evidence indicates that the DRN sends 5HT projections to the LPBN [14,31], forming a key pathway to regulate homeostatic responses under hydroelectrolyte balance alterations. Previous investigations also demonstrated that serotonergic mechanisms in the LPBN play an inhibitory role in controlling sodium appetite following a variety of dipsogenic and/ or natriorexigenic stimuli [17,[32][33][34][35]. ...
In order to determine whether serotonergic (5HT) dorsal raphe nucleus (DRN) cells are involved in body sodium status regulation, the effect of a s.c. infusion of either 2 M or 0.15 M NaCl on 5HT DRN neuron firing was studied using single unit extracellular recordings. In separate groups of 2 M and 0.15 M NaCl-infused rats, water intake, oxytocin (OT) plasma concentration, urine and plasma sodium and protein concentrations were also measured. Also, to determine the involvement of particular brain nuclei and neurochemical systems in body sodium overload (SO), animals from both groups were perfused for brain immunohistochemical detection of Fos, Fos-OT and Fos-5HT expression. SO produced a significant increase in serotonergic DRN neuron firing rate compared to baseline and 0.15 M NaCl-infused rats. As expected, 2 M NaCl s.c. infusion also induced a significant increase of water intake, diuresis and natriuresis, plasma sodium concentration and osmolality, even though plasma volume did not increase as indicated by changes in plasma protein concentration. The distribution of neurons along the forebrain and brainstem expressing Fos after SO showed the participation of the lamina terminalis, extended amygdala, supraoptic and paraventricular hypothalamic nuclei in the neural network that controls osmoregulatory responses. Both Fos-OT immunoreactive and plasma OT concentration increased after s.c. hypertonic sodium infusion. Finally, matching the "in vivo" electrophysiological study, SO doubled the number of Fos-5HT immunolabeled cells within the DRN. In summary, the results characterize the behavioral, renal and endocrine responses after body sodium overload without volume expansion and specify the cerebral nuclei that participate at different CNS levels in the control of these responses. The electrophysiological approach also allows us to determine in an "in vivo" model that DRN 5HT neurons increase their firing frequency during an increase in systemic sodium concentration and osmolality, possibly to modulate sodium and water intake/excretion and avoid extracellular volume expansion.
... Our current findings replicate and extend these studies by demonstrating that the EB-induced increase in Pet-1, 5-HTT, and TPH-2 mRNA levels, is localized within the DRV subnucleus of the DRN. The significance of this relates to the fact that the DRV has reciprocal connections with the hypothalamus and parabrachial nucleus (Petrov et al., 1992;Peyron et al., 1998). As a result, the DRV may be in a position to directly regulate a portion of the neural network controlling food intake. ...
The most potent, naturally occurring estrogen, estradiol, is involved in the physiological control of food intake. Estradiol appears to exert its anorexigenic effect by activating nuclear estrogen receptors (ERs), which are expressed widely in peripheral tissues and in the brain. However, the relative contributions of peripheral and central ERs to estradiol’s anorexigenic effect are largely unknown. Experiment 1 utilized an antiestrogenic compound that fails to cross the blood brain barrier, ICI 182,780 (ICI), to determine whether blockade of either peripheral or central ERs could attenuate the anorexigenic effect of estradiol in ovariectomized (OVX) rats. Peripheral ICI treatment failed to attenuate estradiol’s anorexigenic effect at a dose that was sufficient to block a classic ER action in the periphery. In contrast, central ICI infusion blocked estradiol’s anorexigenic effect, suggesting that activation of central, but not peripheral, ERs is necessary for estradiol’s anorexigenic effect.
Estradiol appears to make rats more sensitive to the anorexigenic effects of serotonin (5-HT), a neurotransmitter that is released during meals and functions to inhibit meal size. For example, previous research from our lab revealed that estradiol increases the anorexigenic effects of fenfluramine (FEN), a serotonergic drug that produces a rapid increase in 5-HT neurotransmission. To begin to examine the mechanism by which estradiol increases the anorexigenic effects of 5-HT, Experiment 2 was designed to investigate whether estradiol increases the expression of serotonergic genes involved in the regulation of 5-HT neurotransmission, including pheochromocytoma 12 ETS domain transcription factor (Pet-1), 5-hydroxytryptophan transporter (5-HTT), and tryptophan-hydroxylase isoform 2 (TPH-2), in the midbrain raphe nuclei (dorsal and medial raphe nuclei) of OVX rats. Estradiol increased Pet-1, 5-HTT, and TPH-2 mRNA levels in midbrain raphe nuclei at a time that coincided with estradiol’s inhibitory effect on food intake. These findings demonstrate that estradiol increases the expression of serotonergic genes in midbrain raphe nuclei and this effect may increase serotonergic tone in a feeding-related, neural circuit. The 5-HT agonist, FEN, increases 5-HT neurotransmission by increasing the release of 5-HT, by decreasing the reuptake of 5-HT, and by activating postsynaptic 5-HT2C receptors. Because of this lack of specificity, it is hard to determine whether estradiol may be acting presynaptically, to increase release of 5-HT, or postsynaptically, to increase activation of excitatory 5-HT2C receptors. Experiment 3 was designed to test the latter hypothesis. The anorexigenic effect of mCPP, a selective 5-HT2C receptor agonist, was examined in estradiol- and vehicle-treated OVX rats. In support of our hypothesis, the magnitude of mCPP-induced anorexia was significantly greater in estradiol-treated rats, relative to oil-treated rats. This finding suggests that estradiol interacts with the postsynaptic 5-HT2C receptor to decrease food intake. Together, these findings provide the first demonstrations that activation of central ERs is necessary for estradiol’s anorexigenic effect, that a physiological regimen of estradiol treatment increases the expression of multiple serotonergic genes in the midbrain raphe nuclei, and that estradiol increases the anorexia produced by activation of postsynaptic 5-HT2C receptors. The results of the present studies suggest that estradiol interacts with the 5-HT system to control food intake in the female rat.
... The LC and NRD are the origins of the noradrenergic and serotonergic neurotransmitter systems, respectively (Mann 1983;Rossor 1988;Saper 1988). Both brain stem areas have strong connections with the nucleus basalis of Meynert (NBM), the prefrontal cortex, the hippocampus, the amygdala, the hypothalamus, and the SCN (Bobillier et al. 1976;Legoratti-Sanchez et al. 1989;Petrov et al. 1992;Vertes 1991). These areas, which play a role in (working) memory, (affective) behavior, and the rest-activity rhythm (Carpenter & Grossberg 1993;Collette et al. 1997;Salzmann 1992;Swaab 1997), are affected in AD (Blin et al. 1997;Braak & Braak 1991a;Goudsmit et al. 1990;Haroutunian et al. 1998;Swaab 1997). ...
... Chronic stress impacts the function of a number of brain regions. These regions include the dorsal raphe (DR) nucleus, which is a major source of serotonergic input to the mammalian forebrain and which has been implicated in the physiological and behavioral response to stress (Azmitia and Segal, 1978;Mamounas et al., 1991;Jacobs and Azmitia, 1992;Petrov et al., 1992;Maier et al., 1993;Lowry, 2002;Lowry et al., 2005). In addition, evidence suggests that chronic stress leads to affective disorders and impairs learning and memory both in humans and in laboratory animals, thus implicating the hippocampus in response to stress (Sapolsky, 2000;McEwen et al., 2002;Sheline et al., 2003;Duman, 2004;Dagyte et al., 2011). ...
Chronic exposure to stress is associated with a number of psychiatric disorders, but little is known about the epigenetic mechanisms that underlie the stress response or resilience to chronic stress. We investigated histone acetylation in 7 different brain regions of rats exposed to chronic social defeat stress: the dorsal hippocampus, ventral hippocampus, medial prefrontal cortex, basolateral amygdala, locus coeruleus, paraventricular thalamus, and dorsal raphe nucleus. This stress paradigm was unique in that it allowed rats to display resilience in the form of an active coping mechanism. We found that there was an increase in acetylation of H3K9 and bulk acetylation of H4K5,8,12,16 in the dorsal raphe nucleus of rats that were less resilient. Less resilient rats also displayed increased levels of H3K18 acetylation in the medial prefrontal cortex when compared to non-stressed controls. In the ventral hippocampus, there was an increase in acetylation of H3K18 and H4K12 in rats that were less resilient when compared to non-stressed control rats. In addition, there was a decrease in levels of H4K8 acetylation in both resilient and non-resilient rats as compared to controls. We assessed expression of histone modifying enzymes in the ventral hippocampus and the medial prefrontal cortex using quantitative real time PCR and found changes in expression of a number of targets. These included changes in Sirt1 and Sirt2 in the ventral hippocampus and changes in Kat5 in the medial prefrontal cortex. Overall, these results suggest that changes in histone acetylation and expression of histone modifying enzymes in these regions correlate with the behavioral response to stress in socially defeated rats.
... Baro-and cardiopulmonary receptor activation stimulates brainstem structures, such as the nucleus of the solitary tract (NTS), area postrema (AP) and the locus coeruleus (LC), in response to changes in extracellular volume or changes in arterial pressure. The NTS and the AP activate the LC neurones, which, in turn, send projections to integrative nuclei, such as the NPBL (lateral parabrachial nucleus), NDR (dorsal raphe nucleus) and CeA, which project to the Pa and the SON (37,38). The dorsal raphe nucleus, once activated, could act through direct projections or connections with the CeA to activate the release of oxytocin from the magnocellular neurones in the Pa and in the SON (39)(40)(41). ...
This study investigated the involvement of the oxytocinergic neurons that project into the central amygdala (CeA) in the control of electrolyte excretion and hormone secretion in unanaesthetised rats subjected to acute hypertonic blood volume expansion (BVE, 0.3 M NaCl, 2 ml/100 g of body weight over 1 min). Oxytocin and vasopressin mRNA expression in the paraventricular (Pa) and supra-optic nucleus (SON) of the hypothalamus were also determined using Real Time-PCR and in situ hybridization. Male Wistar rats with unilaterally implanted stainless steel cannulas in the CeA were used. Oxytocin (1 μg/0.2 μl), vasotocin, an oxytocin antagonist (1 μg/0.2 μl) or vehicle was injected into the CeA 20 min before the BVE. In rats treated with vehicle in the CeA, hypertonic BVE increased urinary volume, sodium excretion, plasma OT, vasopressin (AVP) and atrial natriuretic peptide (ANP) levels and also increased the expression of OT and AVP mRNA in the Pa and SON. In rats pre-treated with OT in the CeA, previously to the hypertonic BVE there were further significant increases in plasma AVP, OT and ANP levels, urinary sodium and urine output, as well as in gene expression (AVP and OT mRNA) in the Pa and SON compared to BVE alone. Vasotocin reduced sodium, urine output and ANP levels, but no changes were observed in plasma AVP and OT levels or in the expression of the AVP and OT genes in both hypothalamic nuclei. The present results suggest that oxytocin in the CeA exerts a facilitatory role in the maintenance of hydroelectrolyte balance in response to changes in extracellular volume and osmolality. © 2013 British Society for Neuroendocrinology.
... Although 5-HT2C receptormediated actions are best characterized in the ARC, 5HT2C receptor transcripts are widely expressed in hypothalamic regions which are especially implicated in appetite and energy balance, including the ARC, PVN, VMN, DMN, and LH (Hoffman and Mezey, 1989;Pasqualetti et al., 1999;Wright et al., 1995). These structures all receive 5-HTergic innervations from the mesencephalic dorsal and/or median raphe nuclei (Petrov et al., 1992;Steinbusch and Nieuwenhuys, 1981). ...
Short sleep duration has been suggested to be a risk factor for weight gain and adiposity. Serotonin (5-HT) substantially contributes to the regulation of sleep and feeding behavior. Although 5-HT predominately promotes waking and satiety, the effects of 5-HT depend on 5-HT receptor function. The 5-HT1A, 5-HT1B, 5-HT2A, 5-HT2C, 5-HT6, and 5-HT7 receptors reportedly contribute to sleep-waking regulation, whereas the 5-HT1B and 5-HT2C receptors contribute to the regulation of satiety. The 5-HT1B and 2C receptors may therefore be involved in the regulation of sleep-feeding. In genetic studies, 5-HT1B receptor mutant mice display greater amounts of rapid eye movement sleep (REMS) than wild-type mice, while displaying no effects on waking or slow wave sleep (SWS). On the other hand, 5-HT2C receptor mutant mice exhibit increased wakefulness and decreased SWS, without any effect on REMS. Moreover, the 5-HT2C receptor mutants display leptin-independent hyperphagia, leading to a middle-aged onset of obesity, whereas 5-HT1B receptor mutants do not display any effect on food intake. Thus, the genetic deletion of 5-HT2C receptors results in sleep loss-associated hyperphagia, leading to the late onset of obesity. This is a quite different pattern of sleep-feeding behavior than is observed in disturbed leptin signaling, which displays an increase in sleep-associated hyperphagia. In pharmacologic studies, 5-HT1B and 5-HT2C receptors upregulate wakefulness and downregulate SWS, REMS, and food intake. These findings suggest that 5-HT1B/2C receptor stimulation induces sleep loss-associated anorexia. Thus, the central 5-HT regulation of sleep-feeding can be dissociated. Functional hypothalamic proopiomelanocortin and orexin activities may contribute to the dissociated 5-HT regulation.
... These results suggested that activation of GABA B receptors in the DRN can modulate the activity of 5-HT and increase inter-male aggression in several test conditions. However, the DRN also contains large number of other types of neurons that are not serotonergic, and many of those neurons also project to distant brain nuclei (Beitz et al. 1986;Ma et al. 1991;Petrov et al. 1992;Van Bockstaele et al. 1993;Kanno et al. 2008, Kirifides et al. 2001Kim et al. 2004;Halberstadt and Balaban, 2006). Those neurons can also be affected by local baclofen treatment. ...
Pharmacological activation of GABA(B) receptors in the dorsal raphe nucleus (DRN) can escalate territorial aggression in male mice.
We characterized this escalated aggression in terms of its behavioral and environmental determinants.
Aggressive behavior of resident male (CFW or ICR mouse) was assessed in confrontations with a group-housed intruder. Either baclofen (0.06 nmol/0.2 μl) or vehicle (saline) was microinjected into the DRN 10 min before the confrontation. We examined baclofen-heightened aggression in five situations: aggression in a neutral arena and after social instigation (experiment 1), aggression during the light phase of the cycle (experiment 2), aggression without prior fighting experience (experiment 3), aggression toward a female (experiment 4), and aggression after defeat experiences (experiment 5). In addition, we examined the body targets towards which bites are directed and the duration of aggressive bursts after baclofen treatment.
Regardless of the past social experience, baclofen escalated aggressive behaviors. Even in the neutral arena and after defeat experiences, where aggressive behaviors were inhibited, baclofen significantly increased aggression. Baclofen increased attack bites directed at vulnerable body areas of male intruders but not toward a female and only in the dark. Also, baclofen prolonged the duration of aggressive bursts.
For baclofen to escalate aggression, specific stimulation (male intruder) and tonic level of serotonin (dark cycle) are required. Once aggressive behavior is triggered, intra-DRN baclofen escalates the level of aggression to abnormal levels and renders it difficult to terminate. Also, baclofen counteracts the effects of novelty or past experiences of defeat.
The brainstem is a neglected brain area in neurodegenerative diseases, including Alzheimer's and Parkinson's disease, frontotemporal lobar degeneration and autonomic dysfunction. In Depression, several observations have been made in relation to changes in one particular the Dorsal Raphe Nucleus (DRN) which also points toward as key area in various age-related and neurodevelopmental diseases. The DRN is further thought to be related to stress regulated processes and cognitive events. It is involved in neurodegeneration, e.g., amyloid plaques, neurofibrillary tangles, and impaired synaptic transmission in Alzheimer's disease as shown in our autopsy findings. The DRN is a phylogenetically old brain area, with projections that reach out to a large number of regions and nuclei of the central nervous system, particularly in the forebrain. These ascending projections contain multiple neurotransmitters. One of the main reasons for the past and current interest in the DRN is its involvement in depression, and its main transmitter serotonin. The DRN also points toward the increased importance and focus of the brainstem as key area in various age-related and neurodevelopmental diseases.
This review describes the morphology, ascending projections and the complex neurotransmitter nature of the DRN, stressing its role as a key research target into the neural bases of depression.
Experiments were performed on anaesthetized rats and cats to examine the effects of cardiopulmonary afferent activation on neuronal activity in the nucleus ambiguus (NA) and the nucleus tractus solitarius (NTS). Furthermore, the role of 5-HT1B/1D and 5-HT3 receptors in the NTS was studied in rats including examinations into their role on identified inputs, such as cardiopulmonary afferent activation. Anaesthetized animals were instrumented to allow recording of cardiovascular and respiratory variables, neurones were recorded using glass microelectrodes and the central administration of compounds was carried out using ionophoresis. Cardiopulmonary afferents were activated by right atrial injections of the 5-HT3 receptor agonist phenylbiguanide (PBG), and NA and NTS neurones were identified by a combination of their response to vagus nerve stimulation and histological localisation. The majority of B-fibre cardiac vagal preganglionic neurones (CVPNs) of the NA were excited by right atrial PBG administration and this excitation was maintained in the absence of respiratory drive. The majority of NTS neurones recorded responded to right atrial PBG, with the highest proportions of these neurones (70%) excited by this stimulus. The 5-HT1B/1D receptor agonist sumatriptan decreased the activity of the majority of NTS neurones recorded and also attenuated both the vagal- evoked and cardiopulmonary afferent-evoked activation of NTS neurones. In contrast, the 5-HT1B receptor agonist CP-93,129 increased the activity of the majority of NTS neurones, and excited both vagal-evoked and cardiopulmonary afferent-evoked activation. The inhibitory action of sumatriptan on neuronal activity was attenuated in the majority of neurones in the presence of the 5-HT1D receptor antagonist ketanserin, whilst it was potentiated in the majority of neurones in the presence of the 5-HT1B receptor antagonist GR55562B. Ionophoretic PBG increased the activity of the majority of NTS neurones and potentiated the vagal-evoked and cardiopulmonary afferent-evoked activation of neurones. The 5-HT3 receptor antagonists granisetron and ondansetron and the NMDA receptor antagonist AP-5 attenuated this PBG- evoked increase in activity in the majority of neurones. Granisetron and AP-5 also attenuated the cardiopulmonary afferent-evoked activation of the majority of NTS neurones. These data suggest that the cardiopulmonary reflex-evoked bradycardia is mediated in part by B-fibre CVPNs in the NA. In addition, 5-HT1B and 5-HT3 receptors play an excitatory role, whilst 5-HT1D receptors play an inhibitory role in the functioning of the NTS. Furthermore, the excitatory action of 5-HT3 receptors is mediated in part via the release of glutamate, acting at NMDA receptors, and these 5-HT3 receptors are tonically active during cardiopulmonary afferent transmission in the NTS.
Le neurotransmetteur sérotonine contrôle une grande variété de mécanismes physiologiques et comportementaux. Chez les humains, les mutations qui affectent la monoamine oxydase (MAO), l’enzyme qui dégrade la sérotonine, sont hautement délétères. Pourtant, le morphotype cavernicole aveugle de l’espèce Astyanax mexicanus (téléostéens) porte une mutation (P106L) induisant une perte de fonction partielle de cette enzyme, et semble prospérer dans son habitat souterrain naturel. Cette thèse décrit les effets de cette mutation, depuis l’échelle moléculaire à l’échelle des populations, afin de mieux cerner sa contribution à l’évolution des traits neuro-comportementaux du poisson cavernicole lors de son adaptation au milieu souterrain.Dans une première publication, nous avons établi 4 lignées de poissons, correspondant aux deux morphotypes de l’espèce Astyanax mexicanus (poisson cavernicole et poisson vivant en surface), chacun porteur ou non de la mutation P106L. La mutation P106L affecte le stress en augmentant l’amplitude de la réponse aigue au stress, tout en diminuant l’anxiété. Nous avons aussi étudié la distribution des allèles mutés dans les populations naturelles au Mexique et trouvé que l’allèle mutant est présent dans plusieurs grottes. L’évolution de l’allèle muté sous sélection ou dérive génétique est discutée.Dans une seconde publication, nous avons étudié les conséquences structurelles et biochimiques de la mutation P106L. La réduction de l’activité enzymatique de la MAO mutée est probablement due à une réduction de la flexibilité de l’une des boucles formant l’entrée au site actif, limitant l’accès des substrats. Des mesures d’HPLC ont montré des perturbations majeures de taux de sérotonine, dopamine et noradrénaline (et métabolites) dans le cerveau des poissons mutants. La mutation P106L mao est donc entièrement responsable du déséquilibre monoaminergique observé dans le cerveau des poissons cavernicoles. Enfin, l’effet de la mutation est partiellement compensé par une réduction de l’activité de synthèse de la sérotonine par la TPH. Nos résultats en révèlent plus sur les spécificités des systèmes monoaminergiques des poissons.Enfin, pour aggraver l’inhibition de la MAO, nous avons généré le premier poisson mutant knockout pour la MAO. Les mutants KO homozygotes sont chétifs et meurent durant les premières semaines de développement. Contrairement au système dopaminergique, le système sérotoninergique des poissons KO homozygotes est très fortement altéré : aucun neurone positif à la sérotonine n’est détecté dans l’hypothalamus.
Angiotensin II (ANG II) acts on AT1 receptors located in the organum vasculosum and subfornical organ (SFO) of the lamina terminalis as a main facilitatory mechanism of sodium appetite. The brain serotonin (5‐HT) system with soma located in the dorsal raphe nucleus (DRN) provides a main inhibitory mechanism. In the present work, we first investigated the existence of ANG II AT1 receptors in serotonergic DRN neurons. Then, we examined whether whole body sodium depletion affects the gene expression of the AT1a receptor subtype and the presumed functional significance of AT1 receptors. Using confocal microscopy, we found that tryptophan hydroxylase‐2 and serotonin neurons express AT1 receptors in the DRN. Immunofluorescence quantification showed a significant reduction in 5‐HT content, but no change in AT1 receptor expression or AT1/5‐HT colocalisation in the DRN after sodium depletion. Whole body sodium depletion also significantly increased Agtr1a mRNA expression in the SFO and DRN. Oral treatment with the AT1 receptor antagonist losartan reversed the changes in Agtr1a expression in the SFO, but not the DRN. Losartan injection into either the DRN or the mesencephalic aqueduct had no influence on sodium depletion‐induced 0.3 M NaCl intake. The results indicate the expression of Agtr1a mRNA in the DRN and SFO as a marker of sodium depletion. They also suggest that serotonergic DRN neurons are targets for ANG II. However, the function of their AT1 receptors remains elusive.
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Background:
Brainstem-focused mechanisms supporting transcutaneous auricular VNS (taVNS) effects are not well understood, particularly in humans. We employed ultrahigh field (7T) fMRI and evaluated the influence of respiratory phase for optimal targeting, applying our respiratory-gated auricular vagal afferent nerve stimulation (RAVANS) technique.
Hypothesis:
We proposed that targeting of nucleus tractus solitarii (NTS) and cardiovagal modulation in response to taVNS stimuli would be enhanced when stimulation is delivered during a more receptive state, i.e. exhalation.
Methods:
Brainstem fMRI response to auricular taVNS (cymba conchae) was assessed for stimulation delivered during exhalation (eRAVANS) or inhalation (iRAVANS), while exhalation-gated stimulation over the greater auricular nerve (GANctrl, i.e. earlobe) was included as control. Furthermore, we evaluated cardiovagal response to stimulation by calculating instantaneous HF-HRV from cardiac data recorded during fMRI.
Results:
Our findings demonstrated that eRAVANS evoked fMRI signal increase in ipsilateral pontomedullary junction in a cluster including purported NTS. Brainstem response to GANctrl localized a partially-overlapping cluster, more ventrolateral, consistent with spinal trigeminal nucleus. A region-of-interest analysis also found eRAVANS activation in monoaminergic source nuclei including locus coeruleus (LC, noradrenergic) and both dorsal and median raphe (serotonergic) nuclei. Response to eRAVANS was significantly greater than iRAVANS for all nuclei, and greater than GANctrl in LC and raphe nuclei. Furthermore, eRAVANS, but not iRAVANS, enhanced cardiovagal modulation, confirming enhanced eRAVANS response on both central and peripheral neurophysiological levels.
Conclusion:
7T fMRI localized brainstem response to taVNS, linked such response with autonomic outflow, and demonstrated that taVNS applied during exhalation enhanced NTS targeting.
Inter-male aggression is an essential component of social behavior in organisms from insects to humans. However, when expressed inappropriately, aggression poses significant threats to the mental and physical health of both the aggressor and the target. Inappropriate aggression is a common feature of numerous neuropsychiatric disorders in humans and has been hypothesized to result from the atypical activation of reward circuitry in response to social targets. The lateral habenula (LHb) has recently been identified as a major node of the classical reward circuitry and inhibits the release of dopamine from the midbrain to signal negative valence. Here, we discuss the evidence linking LHb function to aggression and its valence, arguing that strong LHb outputs to the ventral tegmental area (VTA) and the dorsal raphe nucleus (DRN) are likely to play roles in aggression and its rewarding components. Future studies should aim to elucidate how various inputs and outputs of the LHb shape motivation and reward in the context of aggression.
Serotoninergic neurons are integral parts of central and/or peripheral nervous networks in diverse forms of invertebrates and vertebrates (Parent 1981a,b), suggesting that these neurons provide animals - across phylogeny - with capacities essential for adapting to changing internal and/or external demands. Clues as to their functional role(s) may already be gained from their morphology: giant metacerebral serotoninergic neurons in molluscs have abundantly collateralized axons (Cottrell 1977) as do some of the large multipolar, extensively ramifying serotoninergic neurons of the mammalian raphe nuclei/ extraraphe reticular 5-HT cell groups (Waterhouse et al. 1986; Vertes 1991; Van Boeckstaele et al. 1993; Vertes and Kocsis 1994; Holmes et al. 1994), implying that they innervate multiple target networks along the neuraxis. This enables them to coordinate and harmonize activities (or response properties) in diverse networks with state-dependent determinants such as the prevailing level of central motor tone, (somato)-sensory processing and autonomic regulation across the sleep/wake cycle (Jacobs and Fornal 1993). By way of abundantly collateralizing, a limited number of neurons are capable of modulating electrical activity and afferent input responsivity in multiple targets in a coordinate fashion. Therefore, certain serotoninergic neurons of the brainstem represent archetypical reticular-type multipolar neurons which resemble the Golgi type 1 neurons of the brainstem reticular core described by Scheibel and Scheibel(1958).
Background:
Magnocellular neurosecretory neurons of the hypothalamic supraoptic (SON) and paraventricular (PVN) nuclei synthesize vasopressin and oxytocin in response to signals generated by osmoreceptors and baroreceptors and, respectively, by receptors of the nipples and cervix.
Methods:
We analyzed the literature identifying relevant articles dealing with synaptic inputs of neural afferent pathways to Vasopressin-and Oxytocin-secreting neurons of SON and PVN.
Results:
This article focuses on the multisynaptic pathways involved in the regulation of Vasopressin and Oxytocin secretion.
Conclusion:
An updated topographic description of the afferent pathways involved in the regulation of VPergic and OTergic neurons and their synaptic inputs inducing the stimulus-secretion-coupling has been depicted.
The goal of the present study was to characterize the anatomical and neurochemical relationships that the raphe nuclear complex maintains with respect to lateralized and centralized components of the ventricular system. From this investigation, we 1) determined the ipsilateral vs, contralateral distribution of raphe efferents to the ependymal wall of the lateral ventricle, 2) assessed the degree of collateralization of individual ependymal projection neurons to other sites along the ventricular path, 3) compared the topography of raphe neurons that project to the ventricular lining as well as the lumen of the fourth and lateral ventricles, and 4) evaluated the neurochemical identity of raphe neurons that innervate the ventricular system. After tracer injections into the lateral ventricle, labeled cells were distributed evenly on both sides of the midline in the dorsomedial subregion of the intermediate dorsal raphe nucleus. Further rostrally, labeled cells were clustered bilaterally above the medial longitudinal fasciculi and extended into the median raphe nucleus. Injections that involved the ependymal wall of the lateral ventricle resulted in prominent ipsilateral labeling within the dorsal raphe nucleus, just ventral to the cerebral aqueduct. Most of the labeled cells in this latter group had collateral projections to other sites along the ventricular path. Most of the ventricle projection cells contained serotonin but not nicotinamide adenine dinucleotide phosphate diaphorase. These findings indicate that the raphe nuclear complex is topographically organized with respect to the ventricular system. Selected subsets of serotoninergic dorsal raphe neurons may influence discrete segments of the ventricular system independently as well as coordinate functions throughout the system through axon collaterals to other sites along the ventricular neuraxis. J. Comp. Neurol. 399:61-72, 1998. (C) 1998 Wiley-Liss, Inc.
Serotonin (5-HT) is implicated in the regulation of both behavioral arousal and a brainstem alerting system that operates in wakefulness and in rapid eye movement sleep (REM). Activation of the brainstem alerting system is marked by the presence of ponto-geniculooccipital (PGO) waves that occur in association with orienting in wakefulness and spontaneously in REM. Local application of serotonergic agents into REM and PGO wave regulatory regions can alter REM, but there is conflicting evidence as to whether 5-HT in the brainstem can independently influence PGO wave generation. A potential site of action of 5-HT outside the brainstem is the amygdala, which can influence arousal as well as neurobiological responses to novel and significant stimuli. The amygdala also modulates the occurrence and amplitude of PGO waves. We discuss the linkages between arousal and alerting systems and the role 5-HT may play in their regulation at brainstem and amygdalar sites.
The central autonomic system is defined by the cell groups that receive direct or disynaptically relayed input from the nucleus of the solitary tract, or that contribute projections to autonomic preganglionic neurons or their presynaptic inputs. The cell groups identified by these properties also project extensively among themselves, forming a true network. In this chapter, we identify the cell groups from the medulla and spinal cord through the brainstem, hypothalamus, thalamus, amygdala, and cortex that contribute to the central autonomic system. We then review the connections within the central autonomic network; the pathways that provide visceral sensory input to, and visceral motor output from the network; and the connections of the central autonomic system with cognitive, endocrine, and behavioral systems that must be integrated with autonomic responses. We also consider the neurotransmitters used by this network, and how they contribute to its functions.
It is well established that the brainstem contains discrete groups of serotonin-containing neurons with extensive axonal processes that distribute throughout the neuroaxis. Serotonergic neurons have been implicated in a range of functions prominently including the modulation of various events and states of sleep. We describe the efferent and afferent projections of the dorsal raphe (DR) and the median raphe (MR) nuclei. DR fibers distribute widely throughout the forebrain to dopamine-containing nuclei of the ventral midbrain, the lateral hypothalamus, the midline thalamus, amygdala, the dorsal and ventral striatum and adjoining regions of the basal forebrain, and most of the cortex. In contrast to the DR, the MR is a midline/paramidline system of projections. Specifically, MR fibers mainly distribute to forebrain structures lying on or close to the midline including the medial mammillary and supramammillary nuclei, posterior and perifornical nuclei of hypothalamus, midline and intralaminar nuclei of thalamus, lateral habenula, medial zona incerta, diagonal band nuclei, septum and hippocampus. Overall, MR projections to the cortex are light. With few exceptions, DR and MR project to separate, non-overlapping regions of the forebrain - or, in effect, DR and MR share the serotonergic innervation of the forebrain. Although their outputs are distinct, DR and MR receive common sets of afferent projections from limbic cortices, the medial and lateral preoptic areas, lateral habenula, the perifornical, lateral and dorsomedial nuclei of hypothalamus, and several brainstem nuclei prominently including the midbrain and pontine central gray, locus coeruleus, laterodorsal tegmental nucleus and caudal raphe groups. In addition to common afferents, DR receives significant projections from bed nucleus of stria terminalis, lateral septum, diagonal band nuclei, substantia nigra and the tuberomammillary nucleus, while MR receives distinct projections from the medial septum, mammillary nuclei and the interpeduncular nucleus. There are few projections from the amygdala to either DR or MR. In effect, the DR and MR are positioned to integrate of vast array of information from the brainstem and limbic forebrain and through their extensive axonal network influence virtually all parts of the neuro.
The dorsal raphe nucleus (DRN) is involved in the control of several physiological functions, including nociceptive modulation. This nucleus is one of the main sources of serotonin to the CNS and neuromodulators such as opioids and GABA may be are important for its release. This study evaluated the influence of serotonergic, GABAergic and opioidergic stimulation, as well as their interactions in the DRN, on vocalization nociceptive response during a peripheral noxious stimulus application in guinea pigs. Morphine (1.1 nmol), bicuculline (0.50 nmol) and alpha-methyl-5-HT (1.6 nmol) microinjection on the DRN produces antinociception. The antinociception produced by morphine (1.1 nmol) and alpha-methyl-5-HT (1.6 nmol) into the DRN was blocked by prior microinjection of naloxone (0.7 nmol). The alpha-methyl-5-HT effect blocked by naloxone may indicate the existence of 5-HT2A receptors on enkephalinergic interneurons within the dorsal raphe. Pretreatment with muscimol (0.26 nmol) also prevented the antinociceptive effect caused by morphine (1.1 nmol) when administered alone at the same site, indicating an interaction between GABAergic and opioidergic interneurons. The antinociception produced by bicuculline (0.5 nmol) in the DRN was blocked by prior administration of 8-OH-DPAT (0.5 nmol), a 5-HT1A agonist. This may indicate that the 5-HT autoreceptor activation by 8-OH-DPAT at DRN effector neurons can oppose the bicuculline disinhibition effect applied to the same effectors. Thus, we suggest that 5-HT2 receptor activation in the DRN promotes endorphin/enkephalin release that may disinhibit efferent serotonergic neurons of this present structure by inhibiting GABAergic interneurons, resulting in antinociception.
In previous studies beneficial effects of peripheral electrical or tactile nerve stimulation were observed on memory and affective behaviour in patients with probable Alzheimer's disease. In the present study, it was investigated whether electrical and tactile stimulation applied simultaneously to Alzheimer patients would exceed the effects which were observed following treatment by each type of stimulation separately. Our data reveal that the simultaneous application of the two types of stimulation had a beneficial effect on non-verbal and verbal long-term recognition memory. In addition, patients who were treated participated more in activities of daily living, and were more interested in social contacts. In spite of these positive results, comparisons with those of previous studies suggest that a combination of electrical and tactile stimulation does not yield more effects than application of each type of stimulation separately.
Women are twice as likely as men to suffer from stress-related affective disorders. Corticotropin-releasing factor (CRF) is an important link between stress and mood, in part through its signaling in the serotonergic dorsal raphe (DR). Development of CRF receptor-1 (CRFr1) antagonists has been a focus of numerous clinical trials but has not yet been proven efficacious. We hypothesized that sex differences in CRFr1 modulation of DR circuits might be key determinants in predicting therapeutic responses and affective disorder vulnerability.
Male and female mice received DR infusions of the CRFr1 antagonist, NBI 35965, or CRF and were evaluated for stress responsivity. Sex differences in indices of neural activation (cFos) and colocalization of CRFr1 throughout the DR were examined. Whole-cell patch-clamp electrophysiology assessed sex differences in serotonin neuron membrane characteristics and responsivity to CRF.
Males showed robust behavioral and hypothalamic-pituitary-adrenal axis responses to DR infusion of NBI 35965 and CRF, whereas females were minimally responsive. Sex differences were also found for both CRF-induced DR cFos and CRFr1 co-localization throughout the DR. Electrophysiologically, female serotonergic neurons showed blunted membrane excitability and divergent inhibitory postsynaptic current responses to CRF application.
These studies demonstrate convincing sex differences in CRFr1 activity in the DR, where blunted female responses to NBI 35965 and CRF suggest unique stress modulation of the DR. These sex differences might underlie affective disorder vulnerability and differential sensitivity to pharmacologic treatments developed to target the CRF system, thereby contributing to a current lack of CRFr1 antagonist efficacy in clinical trials.
Systemic administration of the serotonin (5-HT) releaser/reuptake inhibitor, d-fenfluramine decreases consumption of food in mammals. This hypophagic action involves loci at several levels of the neuraxis. Indirect evidence implicates the parabrachial nucleus (PBN) of the pons as one of these regions. Consistent with this hypothesis, unilateral infusion of d-fenfluramine (200, 280, and 400 nmol/0.5 μl) directly into the lateral PBN (LPBN) of male rats reduced food intake by 33%, 56%, and 66% from baseline (7.3±0.7 g) during a 30-min test with chow. Infusions lateral, medial, and dorsal to the PBN were ineffective. Stimulating 5-HT1B receptors in the PBN also reduces feeding. Administration of the selective 5-HT1B agonist CP-93,129 (3-(1,2,5,6-tetrahydropyrid-4-yl)pyrrolo[3,2-b]pyrid-5-one) (0, 0.625, 2.5, and 10 nmol/0.5 μl) into the PBN reduced food intake by 25–79%. The selective 5-HT1B antagonist SB-216641 (N-[3-[3-(dimethylamino(ethoxy]-4-methoxyphenyl]-2′-methyl-4′-(5-methyl-1,2,4-oxadiazol-3-yl)-[1,1′-biphenyl]-4-carboxamide) (2.5 nmol) completely blocked the hypophagic action of the approximate ED50 doses of CP-93,129 (2.5 nmol) and d-fenfluramine (280 nmol). These data strongly suggest that directly or indirectly activating 5-HT1B receptors in the LPBN inhibits feeding and implicates this pontine region in the serotonergic regulation of eating and satiation.
In the present study, the hypothesis was tested that peripheral tactile nerve stimulation by massage would improve various aspects of affective behavior in patients with probable Alzheimer's disease. It was assumed that peripheral tactile stimulation might activate higher-level brain structures (e.g. the hypothalamus). The present study revealed that patients who were stimulated with tactile stimulation felt less depressed, less anxious, more well tempered, and were more alert. Apart from that, their personal orientation and their environmental orientation in place improved, they were more interested in social contacts and they participated more in activities of daily living. However, the observed effects could not be maintained over a period of six weeks following treatment.
Serotonin was first isolated from serum by Rapport et al. 1,2 and was subsequently found to be present in the brain and to function as a neurotransmitter. 3–5 Over the course of the last 50 years there has been an explosion of knowledge of the serotonergic system. The relatively
recent development of selective serotonin reuptake inhibitors (SSRIs) and their effectiveness in treating a vast array of
conditions (e.g. depression, anxiety, obesity, bulimia, aggression, obsessive compulsive disorder and post-traumatic stress
disorder) has only served to heighten interest into the multitude of functions that serotonin influences via its actions within
the central nervous system. 614 Two of these functions of serotonin—the activation of various neuroendocrine systems and modulation
of the sleep/wake cycle—wiU be dealt with in this review. Particular emphasis will be placed on the possible role that neuroendocrine
systems may play in mediating some of the effects of serotonin on the three major components of the sleep/wake cycle: wakefulness,
slow wave sleep (SWS) and rapid eye-movement (REM) sleep
Recent observations have demonstrated that the ovarian steroid estrogen, acting through one of its receptors, specifically
estrogen receptor-α (ERα), in distinct regions in the central nervous system (CNS), is responsible for body weight regulation.
The ventral medial hypothalamus (VMH) contains two key nuclei, the arcuate (ARC), which contains populations of neurons that
regulate food intake and body weight, and the ventral medial nucleus (VMN). The VMN alone has not been found to have a prominent
role in the regulation of food intake and body weight, but does influence energy expenditure. ERα is expressed in the CNS,
and estrogenic activation of ERα in discrete brain nuclei is required for the coordinated control of food intake and energy
expenditure. Further, we discuss data suggesting that the effects of estrogens on food intake and energy expenditure are mediated
by estrogenic enhancement/augmentation of peripheral and neuronal signals in distinct hypothalamic neurons. In sum, estrogens
are critical regulators of body weight homeostasis.
KeywordsFood intake-Body weight-Energy homeostasis-Estrogens-Estrogen receptor alpha-Central nervous system
The opioid peptide enkephalin emerges as a major neuromodulator in the regulation and integration of the physiologic response in stressful conditions. The paraventricular nucleus of the hypothalamus is a coordinating center of neuroendocrine and autonomic functions. However, the detailed distribution of the enkephalin fibers and terminals in the paraventricular nucleus and the sources of enkephalinergic innervation are not well defined. In the present study, we used immunocytochemistry for the proenkephalin-derived octapeptide met-arg6-gly7-leu8 enkephalin to determine the distribution of enkephalin-immunoreactive fibers and somata within paraventricular nucleus. Without colchicine pretreatment, enkephalinergic fibers were prominent mainly in the ventromedial part of the parvicellular subdivision of the paraventricular nucleus, appearing in coronal sections as a dense collection of short segments of enkephalin-immunoreactive fibers. In the periventricular portion of the paraventricular nucleus, enkephalin-immunoreactive fibers produced a moderate plexus of short enkephalin-immunoreactive fibers dorsoventrally oriented. With colchicine treatment, a dense cluster of enkephalin-immunoreactive cell bodies was located in the dorsomedial and the dorsal parts of the parvicellular subdivisions. These enkephalin-immunoreactive neurons were small (< 10 μm) to medium sized (10–15 μm), with round and elongated shapes. Retrograde transport of wheat germ-conjugated gold particles. WGA-apoHRP-Au, from the paraventricular nucleus, combined with immunocytochemistry for enkephalin revealed that the major sources of extrahypothalamic enkephalin afferents to the paraventricular nucleus are provided by enkephalin neurons neurons in the lateral reticular nucleus and the paragigantocellularis reticular nucleus of the medulla (∼20% of retrogradely labeled neurons within this nucleus were double labeled) and in the nucleus solitary tract (∼10% of retrogradely labeled neurons within thisnucleus were double labeled). Retrogradely labeled enkephalin neurons were also observed in the medial preoptic area, median preoptic nucleus, dorsomedial hypothalamic nucleus, lateral septum and hypothalamic arcuate nucleus. These enkephalinergic pathways from the medulla and the forebrain could represent an anatomical substrate underlying the opioid effects on paraventricular neurons during physiological processes, such as a cardiovascular regulation, feeding or stress responses.
It is well established that the brainstem contains discrete groups of serotonin-containing neurons with extensive axonal processes that distribute throughout the neuroaxis. Serotonergic neurons have been implicated in a range of functions including sleep/wakefulness, feeding, affect/emotion, thermoregulation, and cognitive behaviors. We describe the projections and some functional properties of the two major serotonergic cell groups of the brain, the dorsal raphe (DR) and the median raphe (MR) nuclei. DR fibers distribute widely throughout the forebrain to dopamine-containing nuclei of the ventral midbrain, the lateral hypothalamus, the midline thalamus, amygdala, the dorsal and ventral striatum and adjoining regions of the basal forebrain, and most of the cortex. By contrast with DR, MR is a midline/paramidline system of projections. Specifically, MR fibers mainly distribute to forebrain structures lying on or close to the midline including the medial mammillary and supramammillary nuclei, posterior and perifornical nuclei of the hypothalamus, midline and intralaminar nuclei of the thalamus, lateral habenula, medial zona incerta, diagonal band nuclei, septum and hippocampus. Overall, MR projections to the cortex are light. With few exceptions, DR and MR project to separate, non-overlapping regions of the forebrain — or, in effect, DR and MR share the serotonergic innervation of the forebrain. We discuss roles for DR in sleep/wake control, feeding/appetite and mood/affect, and for MR in the control of electroencephalographic (EEG) activity of the hippocampus — or states of hippocampal EEG desynchronization.
Contextual fear is followed by significant reductions in rapid eye movement sleep (REM) that are regulated by the central nucleus of the amygdala (CNA). Corticotropin-releasing factor (CRF) plays a major role in regulating the stress response as well as arousal, and CRF in CNA is implicated in stress-related behavior. To test the hypothesis that CRF regulation of CNA is involved in fear-induced alterations in REM, we determined the effects of microinjections into CNA of the CRF1 antagonist, antalarmin (ANT) on fear-induced reductions in REM. We also evaluated c-Fos activation in the hypothalamic paraventricular nucleus (PVN), locus coeruleus (LC), and dorsal raphe nucleus (DRN) to determine whether activation of these regions was consistent with their roles in regulating stress and in the control of REM.
On separate days, rats were subjected to baseline and 2 shock training sessions (S1 and S2). Five days later, the rats received bilateral microinjections of ANT (4.8 mM) or vehicle (VEH) prior to exposure to the fearful context. Sleep was recorded for 20 h in each condition. Freezing was assessed during S1, S2, and context. Separate groups of rats received identical training and microinjections or handling control (HC) only, but were sacrificed 2 h after context exposure to assess c-Fos expression.
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Compared to baseline, S1 and S2 significantly reduced REM. Exposure to the fearful context reduced REM in VEH treated rats, whereas REM in ANT treated rats did not differ from baseline. ANT did not significantly alter freezing. Fear-induced c-Fos expression was decreased in PVN and LC after ANT compared to VEH.
The results demonstrate that CRF receptors in CNA are involved in fear-induced reductions in REM and neural activation (as indicated by c-Fos) in stress and REM regulatory regions, but not in fear-induced freezing.
Previous reports indicate that the midbrain periaqueductal gray and the central nucleus of the amygdala are interconnected but the organization of these projections has not been characterized. We have analyzed this reciprocal circuitry using anterograde and retrograde tracing methods and image analysis. Our findings reveal that innervation of periaqueductal gray from the central nucleus of the amygdala is extensive and discretely organized along the rostrocaudal axis of periaqueductal gray. In addition, the reciprocal projection from periaqueductal gray to the central nucleus of the amygdala is more extensive and more highly organized than previously suggested.
Multiple or single discrete injections of wheatgerm agglutinin‐horseradish peroxidase into several rostrocaudal levels of periaqueductal gray retrogradely labeled a substantial population of neurons, predominantly located in the medial division of the central nucleus of the amygdala. Tracer injections into the central nucleus revealed a high degree of spatial organization in the projection from this nucleus to periaqueductal gray. Two discrete longitudinally directed columns in dorsomedial and lateral/ventrolateral periaqueductal gray are heavily targeted by central amygdalar inputs throughout the rostral one‐half to two‐thirds of periaqueductal gray. Beginning at the level of dorsal raphe and continuing caudally, inputs from the central nucleus terminate more uniformly throughout the ventral half of periaqueductal gray.
In addition, a substantial population of periaqueductal gray neurons were retrogradely labeled from the central nucleus of the amygdala; these were heterogeneously distributed along the rostrocaudal axis of periaqueductal gray, and included both raphe and non‐raphe neurons. Thus, the present study demonstrates that periaqueductal gray receives heavy, highly organized projections from the central nucleus of the amygdala and in turn, has reciprocal connections with the central nucleus.
Previous studies have demonstrated that longitudinally organized columns of output neurons located in dorsomedial and lateral/ventrolateral periaqueductal gray project to the ventral medulla. Thus, there may be considerable overlap between the two longitudinally organized terminal input columns from the central nucleus of the amygdala and the two longitudinal columns of descending projection neurons from periaqueductal gray to the ventral medulla. The central nucleus of the amygdala has been implicated in a variety of emotional/cognitive functions ranging from fear and orienting responses, defensive and aversive reactions, associative conditioning, cardiovascular regulation, and antinociception. Many of these same functions are strongly represented in the periaqueductal gray. It is noteworthy that the present results demonstrate that lateral periaqueductal gray, a preeminent central trigger site for behavioral and autonomic components of the defense/aversion response, is heavily targeted by inputs from the central nucleus of the amygdala at all levels of periaqueductal gray. Thus, the central nucleus of the amygdala to periaqueductal gray projection may be involved in the neural integration of behavioral, antinociceptive and autonomic responses with emotional state. In addition, the present demonstration of extensive reciprocal connections between the central nucleus of the amygdala and periaqueductal gray represents a route via which functional activity represented in periaqueductal gray may gain access to a forebrain structure long implicated m the integration of the cognitive and autonomic components of emotional behavior. Thus, the periaqueductal gray to central nucleus of the amygdala projection may provide a relatively direct linkage between critical species‐preserving behavioral reactions and a forebrain structure capable of influencing multiple nodal points in the descending autonomic system.
The hypothalamus is a major source of afferents to the parabrachial nucleus (PB), but the neurotransmitters in this pathway are largely unknown. In this study, we examine the neuropeptide immunoreactivities of neurons in the hypothalamus that project to the PB by using the combined retrograde fluorescence-immunofluorescence method. After injections of the fluorescent tracer fast blue into the PB, retrogradely labeled neurons were observed in the paraventricular, dorsomedial, ventromedial, median preoptic, and anteroventral periventricular hypothalamic nuclei; in the dorsal, retrochiasmatic, and lateral hypothalamic areas; and in the medial and lateral preoptic areas. Our results show that at least five distinct neuropeptide-immunoreactive cell populations in the hypothalamus project to the PB. In the perifornical lateral hypothalamus, many neurotensin (NT)-, corticotropin-releasing factor-, dynorphin (DYN)-, angiotensin II (AII)-, and galanin-like immunoreactive (-ir) neurons were retrogradely labeled. A cluster of retrogradely labeled neurons in the juxtacapsular lateral hypothalamus stained with an antiserum against alpha-melanocyte stimulating hormone (alpha MSH). Over 50% of the retrogradely labeled cells in the arcuate nucleus were adrenocorticotropin (ACTH)-or alpha MSH-ir. Many alpha MSH- and ACTH-ir, and a few DYN-, NT- and AII-ir neurons in the retrochiasmatic area were retrogradely labeled. Only small numbers of double-labeled neurons were found in the paraventricular nucleus, and, of these, enkephalin-ir and dynorphin-ir neurons were the most common. Somatostatin-ir cells in the hypothalamus were rarely double-labeled. The chemical coding of these hypothalamic projections to the PB may provide important clues to the functional organization of these descending pathways.
The present investigation examined the distributions of immunoreactive neurotensin (NT), cholecystokinin octapeptide (CCK), substance P (SP), methionine enkephalin (ENK), vasoactive intestinal polypeptide (VIP), somatostatin (SS), rat neurophysin II (RNP II), vasopressin (VP), oxytocin (OXY), tyrosine hydroxylase (TH), and serotonin in the parabrachial nuclear complex (PB) of the rat. All of these substances were localized to the PB and they appeared to be chemoarchitecturally organized within the complex. The lateral subdivision (PBL) was organized mediallateral and ventral-dorsal. Specifically NT, CCK, and SP immunoreactive fibers were found to be the most dense in the ventral aspect of the PBL. The distribution of NT-containing fibers was similar to the pattern of CCK-containing fibers and these were localized primarily to the central zone of the PBL. Immunoreactive SP fibers and cells were found in the external and internal zones ventrally and surrounding the dorsal and dorsolateral nuclei in the PBL. Somatostatin, ENK and VIP were found to be the most dense in the dorsal PBL. Serotonin- and TH-containing cells and fibers were found in both the PBL and PBM. These results, coupled with the observations of neuronal connections of the PB and the known functions of this region, underscore the potential involvement for these neuropeptides and monoamines in limbic-brainstem mechanisms of autonomic control.
The localization and distribution of serotonin (5-hydroxytryptamine, 5-HT) has been studied with the indirect immunofluorescence technique using a highly specific and well-characterized antibody to 5-HT. In neuron systems 5-HT was found to be primarily present with a distribution similar to that observed in basic mappings carried out with the formaldehyde-induced fluorescence method. In addition to the nine areas originally described, several other areas in the mesencephalon and rhombencephalon appeared to contain widely distributed 5-HT-positive perikarya. In the median eminence 5-HT fluorescent mast cells could be visualized. No 5-HT-positive nerve cell bodies could be observed either in the telencephalon or diencephalon.
The dorsal and median raphe nuclei in rats were electrically stimulated and blood pressure and heart rate were recorded. Stimulation of each raphe nucleus caused an increase in blood pressure without affecting heart rate. The size of the increase in blood pressure depended upon the stimulus-intensity.Significant increases were already obtained with 5 sec. trains of 0.3 msec., 200 μA stimuli given at a frequency of 50 Hz. The increases in blood pressure could be obtained with electrodes within the raphe nuclei.Pretreating rats with para-chlorophenylalanine (pCPA, 100 mg/kg.day for 3 days) significantly diminished the increases in blood pressure obtained during electrical stimulation of the median raphe nucleus. However, similar pretreatment did not affect blood pressure rises induced by dorsal raphe stimulation.These data are discussed in relation to the role of central serotoninergic mechanisms in cardiovascular control.
Immunocytochemical staining after controlled proteolytic treatment of the sections with pronase revealed widespread distribution of neuronal cell bodies with somatostatin-like immunoreactivity (SLI) in the rat forebrain. SLI-positive neurons were found in regions of the neocortex, the pyriform cortex, the cingulate cortex, the striatum, the olfactory tract and tubercle, the nucleus accumbens, the septum, and the hypothalamus. These results are consistent with previous radioimmunoassay findings and suggest the presence of large somatostatin-like (possibly precursor) molecules in the neurons stained for SLI after pronase treatment.
The rat hypothalamus was studied at the light microscopic level with the use of single and double immunocytochemical staining methods. It was shown that the rat supraoptic and paraventricular hypothalamic nuclei, and their accessory neurosecretory nuclei, do not contain magnocellular somatostatin neurons. The distribution of the hypothalamic parvocellular somatostatin cells is described. The parvocellular component of the rat hypothalamic paraventricular nucleus is, at least partly, composed of somatostatin cells: they form a fairly well circumscribed periventricular cell mass. The rat suprachiasmatic nuclei contain separate somatostatin neurons and vasopressin neurons. Scattered somatostatin cells are present in the entire arcuate nucleus. In addition to the periventricular somatostatin cells located in the preopticanterior hypothalamic area in the arcuate nucleus, the rat hypothalamus also contains numerous scattered somatostatin cells located distant from the third ventricle.
Lesions in the midbrain median but not in the dorsal raphe nucleus significantly decreased the serotonin (5-HT) content of the hippocampus (61%), medial preoptic area (49%), suprachiasmatic nucleus (70%) and anterior hypothalamic area (60%). Electrolytic lesions restricted to either the median or dorsal raphe nucleus produced significant reductions in the 5-HT concentration of the caudate-putamen, anterolateral hypothalamic area (45%) and arcuate nucleus (48--58%). The fall in caudate-putamen 5-HT level was significantly greater after the dorsal (66%) than after the median (24%) raphe lesion. Neither lesion significantly affected the 5-HT content of the posterolateral hypothalamic area, the ventromedial hypothalamic nucleus or the dorsomedial hypothalamic nucleus. Thus, like their differential projections to the caudate-putamen and hippocampus, the mesencephalic dorsal and median raphe nuclei appear to innervate different preoptico-hypothalamic nuclei and areas. Whereas the median raphe nucleus seems to be the primary source of 5-HT fibers to the suprachiasmatic nucleus, anterior hypothalamic area and medial preoptic area, the 5-HT inputs to the anterolateral hypothalamic area and arcuate nucleus appear to derive from both the dorsal and median raphe nuclei.
Projections from the medulla to the parabrachial complex of the rat were examined for their content of neuropeptide Y-, angiotensin II- or galanin-like immunoreactivity using combined retrograde tracing and immunohistochemical techniques. Rhodamine-labelled latex microspheres were stereotaxically injected into discrete nuclei of the parabrachial complex. After survival of two to five days, colchicine (100 μg in 10 μ1 saline) was injected into the cisterna magna. One day later, rats were perfused and the brainstems were prepared for visualization of the retrograde tracer and immunoreactivity of one of the three peptides.
The parabrachial nucleus (PB) is the main relay for ascending visceral afferent information from the nucleus of the solitary tract (NTS) to the forebrain. We examined the chemical organization of solitary-parabrachial afferents by using combined retrograde transport of fluorescent tracers and immunohistochemistry for galanin (GAL), cholecystokinin (CCK), and corticotropin-releasing factor (CRF).
Each peptide demonstrated a unique pattern of immunoreactive staining. GAL-like immunoreactive (-ir) fibers were most prominent in the “waist” area, the inner portion of external lateral PB, and the central and dorsal lateral PB subnuclei. Additional GAL-ir innervation was seen in the medial and external medial PB subnuclei. GAL-ir perikarya were observed mainly rostrally in the dorsal lateral, superior lateral, and extreme lateral PB.
CCK-ir fibers and terminals were most prominent in the outer portion of the external lateral PB; some weaker labeling was also present in the central lateral PB. CCK-ir cell bodies were almost exclusively confined to the superior lateral PB and the “waist” area, although a few cells were seen in the Kölliker-Fuse nucleus.
The distribution of CRF-ir terminal fibers in general resembled that of GAL, but showed considerably less terminal labeling in the lateral parts of the dorsal and central lateral PB, and the external medial and KöUlliker-Fuse subnuclei. The CRF-ir cells were most numerous in the dorsal lateral PB and the outer portion of the external lateral PB; rostrally, scattered CRF-ir neurons were seen mainly in the central lateral PB.
After injecting the fluorescent tracer Fast Blue into the PB, the distribution of double-labeled neurons in the NTS was mapped. GAL-ir cells were mainly located in the medial NTS subnucleus; 34% of GAL-ir cells were double-labeled ipsilaterally and 7% contralaterally. Conversely, 17% of the retrogradely labeled cells ipsilaterally and 16% contralaterally were GAL-ir. CCK-ir neurons were most numerous in the dorsomedial subnucleus of the NTS and the outer rim of the area postrema. Of the CCK-ir cells, 68% in the ipsilateral and 10% in the contralateral NTS were double-labeled, whereas 15% and 10%, respectively, of retrogradely labeled cells were CCK-ir. In the area postrema, 36% of the CCK-ir cells and 9% of the Fast Blue cells were double-labelled CRF-ir neurons were more widely distributed in the medial, dorsomedial, and ventrolateral NTS subnuclei, but double-labeled cells were mainly seen in the medial NTS. Of CRF-ir cells in the NTS, 26% ipsilaterally and 8%contralaterally were retrogradely labeled by the PB injections. Conversely, of retrogradely labeled cells in the NTS, 4% ipsilaterally and 6% contralaterally were CRF-ir.
Our results suggest that the functional specificity of NTS afferents may be maintained by their selective termination in particular PB subnuclei. In addition, the neuropeptides found in these pathways may provide chemical coding for the relay of specific types of visceral sensory information to the PB.
By a double-labeling method combining the retrograde tracing of horseradish peroxidase and the immunocytochemical technique, serotonin-, substance P- or leucine-enkephalin-like immunoreactive neurons in the midbrain periaqueductal gray (PAG) and the nucleus raphe dorsalis (DR) of the rat were found to send projection fibers to the central amygdaloid nucleus bilaterally with an ipsilateral dominance. These PAG neurons were chiefly distributed in the ventrolateral PAG subdivision and the ventral parts of medial PAG subdivision at the middle and caudal levels of PAG.
In this study, we utilized a double retrograde axonal tracing technique to investigate the possible existence of collateralized axonal projections from raphe nuclei 'pontis' and 'magnus' to both medial preoptic area (MPA) and cervical spinal cord (C1-C2). Following microinjections of fluorescent tracers Fast blue (FB) and Diamidino yellow (DY) within MPA and C1-C2, substantial numbers of FB and DY single-labeled neurons, as well as FB-DY double-labeled neurons have been found within raphe nuclei 'pontis' and 'magnus'.
Immunohistochemical and axonal transport techniques were used to characterize the origin and distribution of galanin-immunoreactive inputs to the paraventricular (PVH) and supraoptic (SO) nuclei of the hypothalamus in the rat. In the parvicellular division of the PVH, the most prominent inputs were confined to the anterior and periventricular parts of the nucleus rostrally and the dorsal and ventral medial subdivisions caudally; the galaninergic inputs to the magnocellular division of PVH and SO were very sparse and were preferentially distributed to regions containing predominantly oxytocinergic neurons. A combined retrograde transport-immunohistochemical method was employed to identify sources of these projections. Galanin immunoreactivity was found to coexist with dopamine-beta-hydroxylase (DBH) immunoreactivity in subsets of retrogradely labeled neurons of the A1 and A6 (locus coeruleus) catecholamine cell groups; no evidence was adduced for the presence of galanin in adrenergic (i.e., phenylethanolamine-N-methyltransferase-positive) neurons that project to the PVH. Apart from minor contributions from the mesencephalic raphe nuclei, no other brainstem cell groups contributed to the galaninergic innervation of the PVH. In the forebrain, the most prominent grouping of doubly labeled cells was centered in the rostral part of the dorsomedial nucleus of the hypothalamus (DMH), though significant numbers were also found in the lateral hypothalamic area, the arcuate nucleus, and the medial preoptic area. In experiments designed to define the subnuclear specificity of some galanin-containing inputs to the PVH, iontophoretic deposits of the anterogradely transported plant lectin, Phaseolus vulgaris-leucoagglutinin (PHA-L), were placed in the A1 and A6 cell groups and in the DMH. Sections through the PVH were prepared so as to allow colocalization of anterogradely transported PHA-L and galanin immunoreactivity in individual fibers and varicosities. Consistent with the retrograde transport data, the greatest degree of galanin-PHA-L correspondence was seen after lectin deposits in the DMH, and over 80% of the doubly labeled varicosities were confined to the anterior, periventricular, and medial parvicellular subdivisions of the nucleus. The galanin-containing projection from the locus coeruleus was most circumscribed, with the vast majority of doubly labeled varicosities confined to the periventricular and adjoining aspects of the anterior and medial parvicellular subdivisions.(ABSTRACT TRUNCATED AT 400 WORDS)
Brainstem afferents to the magnocellular basal forebrain were studied by using tract tracing, immunohistochemistry and extracellular recordings in the rat. WGA‐HRP injections into the horizontal limb of the diagonal band (HDB) and the magnocellular preoptic area (MgPA) retrogradely labelled many neurons in the pedunculopontine and laterodorsal tegmental nuclei, dorsal raphe nucleus, and ventral tegmental area. Areas with moderate numbers of retrogradely labelled neurons included the median raphe nucleus, and area lateral to the medial longitudinal fasciculus in the pons, the locus ceruleus, and the medial parabrachial nucleus. A few labelled neurons were seen in the substantia nigra pars compacta, mesencephalic and pontine reticular formation, a midline area in the pontine central gray, lateral parabrachial nucleus, raphe magnus, prepositus hypoglossal nucleus, nucleus of the solitary tract, and ventrolateral medulla. A similar but not identical distribution of labelled neurons was seen following WGA‐HRP injections into the nucleus basalis magnocellularis.
The possible neurotransmitter content of some of these afferents to the HDB/MgPA was examined by combining retrograde Fluoro‐Gold labelling and immunofluorescence. In the mesopontine tegmentum, many retrogradely labelled neurons were immunoreactive for choline acetyltransferase. In the dorsal raphe nucleus, some retrogradely labelled neurons were positive for serotonin and some for tyrosine hydroxylase (TH); however, the majority of retrogradely labelled neurons in this region were not immunoreactive for either marker. The ventral tegmental area, substantia nigra pars compacta, and locus ceruleus contained retrogradely labelled neurons which were also immunoreactive for TH. Of the retrogradely labelled neurons occasionally observed in the nucleus of the solitary tract, prepositus hypoglossal nucleus, and ventrolateral medulla, some were immunoreactive for either TH or phenylethanolamine‐N‐methyltransferase.
To characterize functionally some of these brainstem afferents, extracellular recordings were made from antidromically identified cortically projecting neurons, mostly located in the HDB and MgPA. In agreement with most previous studies, about half (48%) of these neurons were spontaneously active. Electrical stimulation in the vicinity of the pedunculopontine tegmerital and dorsal raphe nuclei elicited either excitatory or inhibitory responses in 21% (13/62) of the cortically projecting neurons.
These findings indicate that neurons in the magnocellular basal fore‐brain receive afferents from widely distributed brainstem neurons that contain acetylcholine, serotonin, dopamine, noradrenaline, adrenaline, and some unknown neurotransmitters. Through their actions on magnocellular basal forebrain neurons, these brainstem neurons may indirectly regulate the activity of neurons throughout the cerebral cortex.
In order to determine the responses of paraventricular nucleus neurones following activation of central serotonergic pathways, single unit activity was recorded and responses following electrical stimulation of the midbrain dorsal raphe nucleus were examined. Excitation was recorded from approximately 50% of the cells, independent of whether they were antidromically identified as projecting to the median eminence or unidentified. Approximately 20% of cells were inhibited by the stimulation, the majority of these being unidentified. Parachlorophenylalanine-induced inhibition of serotonin synthesis reduced hypothalamic serotonin levels by 77% and caused a significant reduction in the proportion of cells excited by the stimulation, whereas the inhibitory responses were not affected. Intracerebroventricular administration of the serotonergic neurotoxin, 5,7-dihydroxytryptamine, which caused similar reductions in hypothalamic serotonin content (77%), reduced still further the proportion of excitatory responses and also reduced the proportion of cells inhibited by the stimulation. The data obtained suggest that serotonin acts as an excitatory neurotransmitter in the paraventricular nucleus; this is discussed particularly with respect to the regulation of the hypothalamo-hypophysial-adrenocortical axis. The loss of inhibitory responses in 5,7-dihydroxytryptamine treated, as opposed to the parachlorophenylalanine treated, animals suggests that the serotonergic fibers innervating the recorded cells may contain a cosecreted substance that may have important physiological actions in the control of neuronal activity in the region recorded.
A combined retrograde tracing-immunofluorescent technique was used to identify the relationships between the cellular population projecting to the parabrachial nucleus and the serotonin-immunoreactive cell population of the area postrema in rats. The retrograde fluorescent tracer True Blue was injected in the parabrachial region and 3 days later the animals were perfused. Serial cryostat sections were processed for serotonin immunofluorescence. Three different groups of labeled cells were identified in the area postrema. First, True Blue-positive cells (up to 250/section) that project to the parabrachial nucleus were observed distributed throughout the area postrema. Second, in the pargyline (a monoamine oxidase inhibitor)-treated animals a large number of serotonergic cells (up to 125/section) was observed distributed throughout the area postrema. There was a tendency to a heavier distribution of serotonin-immunoreactive cells in the dorsal two-thirds of the area postrema. Third, double-labelled cells were also seen. Twenty percent of the True Blue-labelled cells projecting to the parabrachial nucleus were serotonin-immunoreactive. Thirty nine percent of the serotonin-immunoreactive population was retrogradely labelled with True Blue. Thus a new serotonergic pathway from the area postrema to the parabrachial nucleus is described; this pathway may be important in the ascending transmission and modulation of chemical and visceral sensory input.
The lateral parabrachial nucleus (LPBN) has been shown to be anatomically linked to a number of forebrain nuclei and medullary structures implicated in the control of body fluid balance and cardiovascular regulation. Although these connections suggest a role for the LPBN in body fluid homeostasis, there is currently little or no physiological or behavioral data to support this notion. The purpose of the present series of experiments was to determine the importance of the ventrolateral region of the LPBN (VLLPBN) in the behavioral response to various thirst challenges. Rats with electrolytic lesions of the VLLPBN and control rats were studied after administration of angiotensin II (ANG II) (1.5 and 3.0 mg/kg), isoproterenol (30 and 100 micrograms/kg), polyethylene glycol (20%) and hypertonic saline (4 and 12%). It was found that rats with lesions drank more in response to ANG II and isoproterenol administration than did control animals.
The use of avidin-biotin interaction in immunoenzymatic techniques provides a simple and sensitive method to localize antigens in formalin-fixed tissues. Among the several staining procedures available, the ABC method, which involves an application of biotin-labeled secondary antibody followed by the addition of avidin-biotin-peroxidase complex, gives a superior result when compared to the unlabeled antibody method. The availability of biotin-binding sites in the complex is created by the incubation of a relative excess of avidin with biotin-labeled peroxidase. During formation of the complex, avidin acts as a bridge between biotin-labeled peroxidase molecules; and biotin-labeled peroxidase molecules, which contains several biotin moieties, serve as a link between the avidin molecules. Consequently, a "lattice" complex containing several peroxidase molecules is likely formed. Binding of this complex to the biotin moieties associated with secondary antibody results in a high staining intensity.
To detect the cell bodies of neurones which project to the area of the hypothalamic para-ventricular nucleus, 10–40 nl of a solution containing horseradish peroxidase and poly-l-ornithine were pressure-injected into one paraventricular nucleus of the rat. After 24 or 48 h, the enzyme remaining at the site of injection was detected by the diaminobenzidine procedure. Retrogradely transported horse-radish peroxidase was visualized by using o-dianisidine as the chromogen substrate.The extent and the intensity of labelling correlated with the apparent volume of the injection site. Labelled cell bodies were observed, ipsilateral to the injection, in the mediobasal hypothalamus, in the limbic system (lateral septum, posteromedial amygdala, ventral subiculum) and in several cell clusters in the brain stem (dorsal raphe nucleus, locus coeruleus, parabrachial nucleus, nucleus of the solitary tract and lateral reticular nucleus). In some animals, light labelling in the organum vasculosum laminae terminalis and in the subfornical organ was observed. No labelled neurones could be detected in the spinal cord.
This review will focus on recent developments demonstrating how drugs can influence respiration via interactions with putative CNS neurotransmitter mechanisms. Thus, not all drugs that affect respiration are mentioned. The primary goal is to relate these drug studies to other background knowledge of anatomy and physiology of respiration in order to provide a framework upon which new drugs can be developed to aid in management of patients with respiratory failure.
Several lipid-mobilizing peptides occur in the pituitary, among them beta-lipotropin and "lipolytic peptide A and peptide B". The latter two peptides are distinct from beta-lipotropin and appear to be chemically related to the neurophysins. Immunohistochemistry has now revealed that the lipolytic peptide B of the pituitary is localized in the ACTH- and MSH-cells. In addition, immunoreactive peptide B was found in axons of the posterior lobe of the pituitary. Immunoreactive peptide B was found also in nerve fibers and nerve cell bodies in the hypothalamus, particularly in the hypothalamo-hypophyseal tract and in the magnocellular neuronal system. Immunoreactive nerve fibers were numerous also in the periventricular nucleus of the thalamus. The antiserum against peptide B cross-reacts with neurophysin I, and hence, it cannot be excluded that at least part of the immunostaining in the brain reflects the presence of the latter component. However, the regional distribution of immunoreactive peptide B and neurophysin was not identical. Therefore, it is possible that authentic peptide B occurs not only in the pituitary but also in the brain.
Immunofluorescence, induced in serotonergic neurons, was combined with retrograde axonal tracing by propidium iodide. Evidence is presented for the existence of serotonergic as well as non-serotonergic projections from the nucleus raphe dorsalis to the caudate-putamen complex.
The distribution of serotonin-immunoreactive varicosities in the paraventricular (PVH) and supraoptic (SO) nuclei was charted in normal immunohistochemical material and the probable cells of origin of these projections were identified using a combined retrograde transport-immunohistochemical method. The density of serotonergic fibers in the PVH and the SO is quite low relative to that in the immediately surrounding neuropil, in striking contrast to noradrenergic inputs to the nuclei. Immunoreactive fibers are concentrated in specific parts of the parvocellular division of the PVH, whereas in the magnocellular division of the nucleus, and in the SO, they are found mostly in regions where oxytocinergic cells predominate. These projections appear to arise from 3 distinct serotonergic cell groups (B7, B8 and B9) in the midbrain.
Alterations in the dynamics of brain serotonin biosynthesis can lead to changes in cardiovascular function. It appears that the activation of cerebral serotonin receptors produces a pressor effect in normotensive rats but produces a depressor effect in normotensive cats or dogs. On the other hand, reductions in the levels of serotonin can prevent the onset of hypertension in some experimental hypertensive models and lower the blood pressure of organisms with established hypertension. The ability of brain serotonin to modulate arterial blood pressure may be mediated by the influences of the serotonergic neuronal systems on efferent sympathetic activity. Finally, the reduction in sympathetic outflow produced by increasing brain serotonin levels in dogs protects the heart against ventricular fibrillation and may, therefore, constitute a reasonable adjunct in the management of high-risk, cardiac-arrest patients.
In summary, we have demonstrated the subnuclear organization of PB, and correlated this with the origins of its efferent connections. In general, PBm projects primarily to the insular, infralimbic and lateral frontal cortex, and to associated areas in the thalamus, hypothalamus and amygdala. PBl chiefly innervates the autonomic nuclei of the hypothalamus and related portions of the amygdala and the bed nucleus of the stria terminalis. KF is the main source of descending projections from PB to the region of the nucleus of the solitary tract, the ventrolateral medulla and the intermediolateral cell column in the thoracic spinal cord. Further subnuclear organization of the origins of these projections within the major PB subdivisions has been described in detail. While PB afferents tend to terminate in specific subnuclei, one cannot reliably predict from the functional properties of the major inputs to a subnucleus what information will be carried in its efferents. Further anatomical and physiological studies of the input-output relationships of single PB neurons will be necessary to help resolve this enigma. However, recent immunohistochemical observations suggest that the subnuclear organization of PB afferent and efferent connections may reflect, at least in part, their biochemical specificity.
Spontaneously firing units in the region of parabrachial nuclei (PB) and Kölliker-Fuse nuclei (KF) of 19 chloralose-anesthetized cats were monitored for changes in firing frequency during electrical stimulation of carotid sinus (CSN) and aortic depressor (ADN) nerves, of central nucleus of the amygdala (ACE), and of paraventricular nuclei of the hypothalamus (PVH). In the ipsilateral PB 64 of 189 and in the contralateral PB 9 of 103 units responded to CSN stimulation; 18 of 185 ipsilaterally and 7 of 97 contralaterally responded to ADN stimulation. Responses were primarily excitatory, and units were located primarily in the ventrolateral portion of the PB. Only 9 of 267 units responded to stimulation of both CSN and ADN. Stimulation of the ACE and PVH antidromically activated 9 and 7 units, respectively, in PB and approximately half of these also responded to buffer nerve stimulation. In the ipsilateral PB 56 of 207 and in the contralateral PB 11 of 103 units responded orthodromically to ACE stimulation, and 23 of 177 ipsilaterally and 2 of 103 contralaterally responded orthodromically to PVH stimulation with primarily excitatory responses and were located primarily in the ventrolateral portion of the PB and KF. Of these units approximately half also responded to buffer nerve stimulation. These results suggest an important role for PB-KF in mediating ascending and descending cardiovascular and respiratory control signals.
Hypothalamic connections with the parabrachial nuclei in the cat were studied at light and electron microscopic levels following wheat germ agglutinin-horseradish peroxidase injections into the parabrachial nuclei and electrolytic lesions in the hypothalamus. The greatest concentration of retrogradely labeled neurons occurred in the paraventricular nucleus. Labeled neurons were also seen within the preoptic, anterior, lateral, dorsomedial and ventromedial hypothalamic nuclei. Hypothalamic lesions resulted in the degeneration of terminals forming axosomatic and axodendritic synapses in the parabrachial nuclei, particularly its lateral division. These findings support the idea that hypothalamic connections to specific regions of the parabrachial nuclei may underlie the topographical functional organization demonstrated for these brainstem nuclei.
A retrograde labeling procedure utilizing fluorescent substances (Granular Blue, Nuclear Yellow and propidium iodide) was used to establish the presence of branching axons in the ascending raphe system of young rats. After injections in septum, medial thalamus and olfactory cortex, the number of double-labeled cells in various combinations was found to be relatively large in the dorsal raphe nucleus, whereas triple-labeled cells occurred more rarely. Each class of neurons, i.e. single-, double- and triple-labeled, were shown to have a predominant distribution within specific parts of the nucleus.
The connections of the subfornical organ (SFO) wer investigated by using the HRP technique. Injections into the SFO labeled neurons in the medial septum, but not in lateral septum nor in the diagonal band nucleus. Labeled cells were observed in the median preoptic nucleus, below the ependyma of the third ventricle, in the dorsal preoptic region near the anterior commissure, and diffusely throughout the medial preoptic and anterior bypothalamic areas.
Fibers were followed from the ventral stalk of the SFO. Precommissural fibers enter the median preoptic nucleus where many of them appear to terminate. Others continue on to the medial septum, the OVLT, the supraoptic nucleus, and the suprachiasmatic nucleus, HRP injections into the median preoptic nucleus labled many neurons in the SFO.
Postcommissural fibers reach the hypothalamus by descending along the walls of the ventricle in the subependymal space, by traveling in the columns of the fornix and the medial corticohypothalamic tract, or by passing through the paraventricular nucleus of the thalamus. Some postcommissural fibers turn rostrally and enter the median preoptic nucleus while others join precommissural fibers bound for the supraoptic nucleus. More caudally directed fibers appear to innervate the paraventricular nucleus of the hypothalamus and the medial preoptic and anterior hypothalamic areas. HRP injections into the paraventricular nucleus of the hypothalamus labeled neurons in the SFO.
These finding corroborate and extend previous work in describing neural connections between two brain regions that are important for fluid blance.
The efferent projections of the subfornical organ (SFO) of rats were traced using the autoradiographic method of following anterograde transport of labelled proteins through axons. The efferents of the SFO go to two different areas. The first is the anteroventral third ventricular area of the preoptic region and the second is the hypothalamus particularly the neurosecretory, magnocellular nuclei. Specifically, the apparent terminal fields in the first area are in the nucleus medianus of the medial preoptic area (NM), the organum vasculosum of the lamina terminalis (OVLT), and the anterior periventricular area (PeV). Many efferent fibers to this area emerge from the rostral SFO, pass anteriorly over the anterior commissure in the midline and either descend along the anterior border of the NM or enter the PeV dorsally just beneath the anterior commissure. The apparent terminal fields within the hypothalamus are in the anterior and tuberal supraoptic nuclei (SONa and SONt), the paraventricular nucleus (PVN) including its rostral accessory cluster, the nucleus circularis (NC), the dorsal perifornical area (PFd), and in both the lateral preoptic area and lateral hypothalamus adjacent to the SON. Many efferent fibers to the hypothalamus emerge from the rostral SFO and enter the columns of the fornix, diverge with the ventral stria medullari to disperse medially and laterally over the columns of the fornix and along their dorsal border at the anterior dorsal level of the columns trajectory through the hypothalamus. These findings are discussed in terms of the SFO's role within a neural network mediating water balance behaviorally and physiologically.
Evidence that lipolytic peptide B occurs in the ACTH/MSH‐cells of the pituitary and in the brain
- Loren I.