Article

The serotonin neurons in nucleus raphe dorsalis of adult rat: A light and electron microscope radioautographic study

May 1982
The Journal of Comparative Neurology 207(3):239-54

May 1982
207(3):239-54

DOI:10.1002/cne.902070305

Source
PubMed

Authors:

Alain Beaudet

Canadian Institutes of Health Research

The serotoninergic nerve cell body population of nucleus raphe dorsalis (RD) was identified by radioautography following cerebroventricular instillation of tritiated serotonin ([3H]5-HT) in adult rats pretreated with a monoamine oxidase inhibitor. Series of histological sections taken throughout the midbrain and upper pons exhibited a similar distribution and number of labeled nerve cell bodies in RD after prolonged administration of either 10-5 or 10-4M [3H]5-HT or 10-4M [3H]5-HT and 10-3M nonradioactive noradrenaline. This allowed systematic mapping and quantification of serotoninergic nerve cell bodies at various levels of the RD. Their extrapolated total number averaged 11,500. Twice as many unreactive (nonserotoninergic) neurons were present within the same region. In electron microscope radioautographs, the labeled cells were usually larger (17.9 micrometer mean diameter) than their unlabeled congeners (13.1 micrometer), but stereological sampling of their perikarial organelle content failed to reveal any difference in cytoplasmic composition. Few [3H]5-HT-labeled axonal varicosities were observed in RD and none were found in close apposition or in synaptic junction with labeled nerve cell bodies, dendrites, or unreactive perikarya. A detailed statistical analysis of silver grain distribution in both labeled and "unlabeled" nerve cell bodies, indicated that in the former, but not in the latter, dense bodies had a relatively high affinity for [3H]5-HT. Mitochondria and the cytoplasmic membrane were the only other organelles to show higher labeling indices in labeled than in unlabeled cells. Other sites of [3H]5-HT localization could be ascribed to artefactitious cross-linkage of the tracer by the fixative, since they had the same relative affinity in the two cell populations. These results provide new insights into the morphology and cytofunctional properties of the 5-HT neurons of rat RD.

Stereological investigation of 5-HT3 receptors in the substantia nigra and dorsal raphe nucleus in the rat

Article

Jan 2021
J CHEM NEUROANAT

Serotonin (5-HT) is a common neurotransmitter in mammals, playing a central role in the regulation of various processes such as sleep, perception, cognitive and autonomic functions in the nervous system. Previous studies have demonstrated that 5-HT type 3 (5-HT3) receptors are expressed in either or both the substantia nigra (SN) and the dorsal raphe nucleus (DRN) in humans, marmosets, rats and Syrian hamsters. Here, we quantify the distribution of 5-HT3 receptors across these regions in the adult rat. Fluorescent immunohistochemistry was performed on sections of rat brain covering the entire rostro-caudal extent of the SN and DRN with antibodies specific to the 5-HT3A receptor subunit, as well as others targeting the monoaminergic markers tyrosine hydroxylase (TH) and the 5-HT transporter (SERT). The number of 5-HT3A receptor-positive, TH-positive (n = 28,428 ± 888, Gundersen’s m = 1 coefficient of error [CE] = 0.05) and SERT-positive (n = 12,852 ± 462, CE = 0.06) cells were estimated in both the SN and the DRN using stereology. We found that 5-HT3A receptor-positive cells are present in the SNr (n = 1250 ± 64, CE = 0.24), but they did not co-localise with TH-positive cells, nor were they present in the SNc. In contrast, no 5-HT3A receptor-positive cells were found in the DRN. These results support the presence of 5-HT3 receptors in the SN, but not in the DRN, and do not support their expression on monoaminergic cells within these two brain areas.

Unraveling the serotonin saga: from discovery to weight regulation and beyond - a comprehensive scientific review

Article

Full-text available

Aug 2023

The prevalence of obesity is rapidly increasing worldwide, while the development of effective obesity therapies lags behind. Although new therapeutic targets to alleviate obesity are identified every day, and drug efficacy is improving, adverse side effects and increased health risks remain serious issues facing the weight-loss industry. Serotonin, also known as 5-HT, has been extensively studied in relation to appetite reduction and weight loss. As a result, dozens of upstream and downstream neural targets of 5-HT have been identified, revealing a multitude of neural circuits involved in mediating the anorexigenic effect of 5-HT. Despite the rise and fall of several 5-HT therapeutics in recent decades, the future of 5-HT as a therapeutic target for weight-loss therapy looks promising. This review focuses on the history of serotonin, the state of current central serotonin research, previous serotonergic therapies, and the future of serotonin for treating individuals with obesity.

Signalisation de l'insuline au sein du système sérotoninergique central : nexus de la comorbidité diabète-dépression

Thesis

Dec 2021

Sébastien Bullich

Les études épidémiologiques estiment que le risque de dépression majeure (DM) est plus élevé chez les patients diabétiques comparé à la population générale. Des études plus spécifiques mettent en lumière des corrélations entre la dégradation de certains paramètres métaboliques et les symptômes anxio-dépressifs chez l'humain. C'est notamment le cas pour l'insulino-résistance périphérique qui est positivement corrélée à la sévérité de la DM. En revanche, les conséquences de l'insulino-résistance centrale sur les troubles dépressifs n'ont jamais été étudiés de manière approfondie non seulement en clinique mais également chez l'animal de laboratoire. Compte tenu de la présence du récepteur à l'insuline dans le cerveau, une des hypothèses serait que cette hormone module directement (ou indirectement) l'activité des systèmes monoaminergiques et notamment celle des neurones sérotoninergiques (5-HT) majoritairement regroupé dans le noyau dorsal du raphé (NDR). En effet, si l'influence de l'insuline sur le système dopaminergique et le comportement alimentaire a déjà été montré, très peu d'études se sont intéressées à son impact sur le système 5-HT pourtant clé dans la physiopathologie de la DM. Au cours de ce travail de thèse nous avons pu montrer que le récepteur à l'insuline est présent sur les neurones 5-HT du NDR. Grâce à des techniques d'électrophysiologie ex- et in-vivo et de microdialyse intracérébrale réalisées sur modèle murin, nous avons caractérisé l'effet excitateur de l'insuline sur l'activité électrique des neurones 5-HT. Ces résultats nous ont amené à tester les effets comportementaux de l'insuline et à montrer les effets anxiolytiques de son injection intra-raphé et intra-nasale chez la souris saine. Dans un second temps, afin de se placer dans un contexte pathologique et de mieux comprendre l'impact de la perturbation de la signalisation de l'insuline sur l'humeur, nous avons étudié l'activité du système 5-HT et les comportements de type anxio-dépressifs dans des modèles murins de diabète de type 1 et 2 (DT1/DT2). Dans ces deux modèles, que ce soit dans un contexte d'insulinopénie (DT1) ou d'insulino-résistance (DT2), les souris présentent un phénotype anxieux et certains traits de la DM associés à un diminution de l'activité du système sérotoninergique du NDR. Enfin, nous avons tenté d'identifier l'implication de l'apeline, une adipokine connue pour ses propriétés insulino-sensibilisatrice sur les anomalies comportementales induites par un DT2. Nos résultats montrent que les souris présentant une invalidation génétique de l'apeline, sont plus susceptibles à développer une insulino-résistance en réponse à un régime alimentaire diabétogène et des troubles comportementaux. De manière intéressante le traitement par la metformine, un antidiabétique aux propriétés insulino-sensibilisatrice, ne permet pas l'amélioration des paramètres métaboliques de ces souris mutantes mais améliore leur état anxieux. Ainsi ce travail de thèse a permis de souligner l'existence d'interactions anatomiques et fonctionnelles entre le système insulinergique et sérotoninergique central ainsi que leur importance dans l'anxiété, un trouble psychiatrique souvent annonciateur d'un épisode dépressif. [...]

Cerebellin-2 regulates a serotonergic dorsal raphe circuit that controls compulsive behaviors

Article

Full-text available

Dec 2021

Cerebellin-1 (Cbln1) and cerebellin-2 (Cbln2) are secreted glycoproteins that are expressed in distinct subsets of neurons throughout the brain. Cbln1 and Cbln2 simultaneously bind to presynaptic neurexins and postsynaptic GluD1 and GluD2, thereby forming trans-synaptic adhesion complexes. Genetic associations link cerebellins, neurexins and GluD’s to neuropsychiatric disorders involving compulsive behaviors, such as Tourette syndrome, attention-deficit hyperactivity disorder (ADHD), and obsessive-compulsive disorder (OCD). Extensive evidence implicates dysfunction of serotonergic signaling in these neuropsychiatric disorders. Here, we report that constitutive Cbln2 KO mice, but not Cbln1 KO mice, display robust compulsive behaviors, including stereotypic pattern running, marble burying, explosive jumping, and excessive nest building, and exhibit decreased brain serotonin levels. Strikingly, treatment of Cbln2 KO mice with the serotonin precursor 5-hydroxytryptophan or the serotonin reuptake-inhibitor fluoxetine alleviated compulsive behaviors. Conditional deletion of Cbln2 both from dorsal raphe neurons and from presynaptic neurons synapsing onto dorsal raphe neurons reproduced the compulsive behaviors of Cbln2 KO mice. Finally, injection of recombinant Cbln2 protein into the dorsal raphe of Cbln2 KO mice largely reversed their compulsive behaviors. Taken together, our results show that Cbln2 controls compulsive behaviors by regulating serotonergic circuits in the dorsal raphe.

Oxytocin receptor expression in the midbrain dorsal raphe is dynamic across female reproduction in rats

Article

Full-text available

Jan 2021

Central oxytocin receptor (OTR) expression is extremely sensitive to circulating steroid hormones and OTRs influence many of the neurobehavioural adaptations associated with female reproduction (e.g., postpartum caregiving, aggression, cognition, affective responses). Changes in central OTR expression across female reproduction have often been studied, but almost all of such research has focused on the forebrain, ignoring hormone‐sensitive midbrain sites such as the serotonergic dorsal raphe (DR) that are also critical for postpartum behaviours. To investigate the effects of female reproductive state on OTRs in the DR, we first used autoradiography to examine OTR binding across four female reproductive states in laboratory rats: dioestrous virgin, pregnancy day 10, the day of parturition and postpartum day 7. OTR binding in the rostral DR (but not other DR subregions) was approximately 250% higher in parturient rats compared to dioestrous virgins and dropped back down to virgin levels by postpartum day 7. Given the chemical heterogeneity of the DR, we then examined OTR expression in the three most abundant neuronal phenotypes of the DR (i.e., serotonin, GABA and dopamine) in dioestrous virgins and recently parturient females. Using dual‐label immunohistochemistry and in situ hybridisation, we found that twice as many dopaminergic cells in the parturient rostral DR contained OTR immunoreactivity compared to that found in virgins. On the other hand, mothers had fewer rostral DR GABAergic cells expressing OTRs than did virgins. OTR expression in serotonin cells did not differ between the two groups. Overall, these results suggest that the rostral subregion of the midbrain DR is uniquely sensitive to oxytocin around the time of parturition, with subpopulations of cells that become more sensitive (i.e., dopamine), less sensitive (i.e., GABA) and show no change (i.e., serotonin) to this neuropeptide. This dynamic OTR signalling in the female DR may help drive the numerous behavioural changes across female reproduction that drive successful motherhood.

A Shift in the Activation of Serotonergic and Non-serotonergic Neurons in the Dorsal Raphe Lateral Wings Subnucleus Underlies the Panicolytic-Like Effect of Fluoxetine in Rats

Article

Full-text available

Sep 2019
MOL NEUROBIOL

A wealth of evidence indicates that the lateral wings subnucleus of the dorsal raphe nucleus (lwDR) is implicated in the processing of panic-associated stimuli. Escape expression in the elevated T-maze, considered a panic-related defensive behavior, markedly and selectively recruits non-serotonergic cells within this DR subregion and in the dorsal periaqueductal gray (dPAG), another key panic-associated area. However, whether anti-panic drugs may interfere with this pattern of neuronal activation is still unknown. In the present study, the effects of acute (10 mg/kg) or chronic fluoxetine (10 mg/kg/daily/21 days) treatment on the number of serotonergic and non-serotonergic cells induced by escape expression within the rat DR and PAG subnuclei were investigated by immunochemistry. The results showed that chronic, but not acute, treatment with fluoxetine impaired escape expression, indicating a panicolytic-like effect, and markedly decreased the number of non-serotonergic cells that were recruited in the lwDR and dPAG. The same treatment selectively increased the number of serotonergic neurons within the lwDR. Our immunochemistry analyses also revealed that the non-serotonergic cells recruited in the lwDR and dPAG by the escape expression were not nitrergic. Overall, our findings suggest that the anti-panic effect of chronic treatment with fluoxetine is mediated by stimulation of the lwDR-dPAG pathway that controls the expression of panic-associated escape behaviors.

The dorsal diencephalic conduction system in reward processing: Spotlight on the anatomy and functions of the habenular complex

Article

Apr 2018
BEHAV BRAIN RES

Marc Fakhoury

The dorsal diencephalic conduction system (DDC) is a highly conserved pathway in vertebrates that provides a route for the neural information to flow from forebrain to midbrain structures. It contains the bilaterally paired habenular nuclei along with two fiber tracts, the stria medullaris and the fasciculus retroflexus. The habenula is the principal player in mediating the dialogue between forebrain and midbrain regions, and functional abnormalities in this structure have often been attributed to pathologies like mood disorders and substance use disorder. Following Matsumoto and Hikosaka seminal work on the lateral habenula as a source of negative reward signals, the last decade has witnessed a great surge of interest in the role of the DDC in reward-related processes. However, despite significant progress in research, much work remains to unfold the behavioral functions of this intriguing, yet complex, pathway. This review describes the current state of knowledge on the DDC with respect to its anatomy, connectivity, and functions in reward and aversion processes.

Plausible Role of Stem Cell Types for Treating and Understanding the Pathophysiology of Depression

Article

Full-text available

Mar 2023

Abstract: Major Depressive Disorder (MDD), colloquially known as depression, is a debilitating condition affecting an estimated 3.8% of the population globally, of which 5.0% are adults and 5.7% are above the age of 60. MDD is differentiated from common mood changes and short-lived emotional responses due to subtle alterations in gray and white matter, including the frontal lobe, hippocampus, temporal lobe, thalamus, striatum, and amygdala. It can be detrimental to a person's overall health if it occurs with moderate or severe intensity. It can render a person suffering terribly to perform inadequately in their personal, professional, and social lives. Depression, at its peak, can lead to suicidal thoughts and ideation. Antidepressants manage clinical depression and function by modulating the serotonin, norepinephrine, and dopamine neurotransmitter levels in the brain. Patients with MDD positively respond to antidepressants, but 10-30% do not recuperate or have a partial response accompanied by poor life quality, suicidal ideation, self-injurious behavior, and an increased relapse rate. Recent research shows that mesenchymal stem cells and iPSCs may be responsible for lowering depression by producing more neurons with increased cortical connections. This narrative review discusses the plausible functions of various stem cell types in treating and understanding depression pathophysiology.

Constitutive 5-HT2C receptor knock-out facilitates fear extinction through altered activity of a dorsal raphe-bed nucleus of the stria terminalis pathway

Article

Full-text available

Nov 2022

Serotonin 2C receptors (5-HT2CRs) are widely distributed throughout the brain and are strongly implicated in the pathophysiology of anxiety disorders such as post-traumatic stress disorder (PTSD). Although in recent years, a considerable amount of evidence supports 5-HT2CRs facilitating effect on anxiety behavior, the involvement in learned fear responses and fear extinction is rather unexplored. Here, we used a 5-HT2CR knock-out mouse line (2CKO) to gain new insights into the involvement of 5-HT2CRs in the neuronal fear circuitry. Using a cued fear conditioning paradigm, our results revealed that global loss of 5-HT2CRs exclusively accelerates fear extinction, without affecting fear acquisition and fear expression. To investigate the neuronal substrates underlying the extinction enhancing effect, we mapped the immediate-early gene product cFos, a marker for neuronal activity, in the dorsal raphe nucleus (DRN), amygdala and bed nucleus of the stria terminalis (BNST). Surprisingly, besides extinction-associated changes, our results revealed alterations in neuronal activity even under basal home cage conditions in specific subregions of the DRN and the BNST in 2CKO mice. Neuronal activity in the dorsal BNST was shifted in an extinction-supporting direction due to 5-HT2CR knock-out. Finally, the assessment of DRN-BNST connectivity using antero- and retrograde tracing techniques uncovered a discrete serotonergic pathway projecting from the most caudal subregion of the DRN (DRC) to the anterodorsal portion of the BNST (BNSTad). This serotonergic DRC-BNSTad pathway showed increased neuronal activity in 2CKO mice. Thus, our results provide new insights for the fear extinction network by revealing a specific serotonergic DRC-BNSTad pathway underlying a 5-HT2CR-sensitive mechanism with high significance in the treatment of PTSD.

Neuromodulatory organization in the developing rat somatosensory cortex

Preprint

Full-text available

Nov 2022

The vast majority of cortical synapses are found in the neuropil which is implicated in multiple and diverse functions underlying brain computation. Unraveling the organizing principles of the cortical neuropil requires an intricate characterization of synaptic connections established by excitatory and inhibitory axon terminals, of intrinsic and extrinsic origin and from ascending projections that govern the function of cortical microcircuits through the release of neuromodulators either through point-to-point chemical synapses or diffuse volume transmission (VT). Even though neuromodulatory release has been studied for almost a century it is still not clear if one modality prevails upon the other. The hindlimb representation of the somatosensory cortex (HLS1) of two-week old Wistar rats has served as a model system to dissect the microcircuitry of neurons and their synaptic connections. In the present study, we quantified the fiber length per cortical volume and the density of varicosities for cholinergic, catecholaminergic and serotonergic neuromodulatory systems in the cortical neuropil using immunocytochemical staining and stereological techniques. Acquired data were integrated into a novel computational framework to reconcile the specific modalities and predict the effects of neuromodulatory release in shaping neocortical network activity. We predict that acetylcholine (ACh), dopamine (DA), serotonin (5-HT) release desynchronizes cortical activity by inhibiting slow oscillations (delta range), and that 5-HT triggers faster oscillations (theta). Moreover, we found that high levels (>40%) of neuromodulatory VT are sufficient to induce network desynchronization, but also that combining volume release with synaptic inputs leads to more robust and stable effects, meaning that lower levels of VT are needed to achieve the same outcome (10%).

Electrophysiological Characteristics of Dorsal Raphe Nucleus in Tail Suspension Test

Article

Full-text available

May 2022

The dorsal raphe nucleus (DRN) is a major source of serotonin in the central nervous system, which is closely related to depression-like behaviors and is modulated by local GABAergic interneurons. Although serotonin neurons are known to be activated by struggling behavior in tail suspension test (TST), the exact electrophysiological characteristics are still unclear. Here, we combined in vivo electrode recording and behavioral test to explore the mice neuron electrophysiology in DRN during TST and observed that gamma oscillation was related to despair-like behaviors whereas burst fraction was crucial for survival-like behaviors. We reported the identification of a subpopulation of DRN neurons which change their firing rates when mice get into and during TST immobile states. Both increase (putative despair units, D units for short) and decrease (putative survival units, S units for short) in firing rate were observed. Furthermore, using optogenetics to identify parvalbumin-positive (PV+) and serotonin transporter-positive (SERT+) neurons, we found that SERT+ neurons were almost S units. Interestingly, those that have been identified PV+ neurons include ~20% of D units and ~50% of S units. These results suggest that electrophysiological characteristics incorporated in despair-like behavior studies can provide new insight into the study of anti-depression targets, and GABAergic interneuron is a complex key hub to the coding and regulation of local neural network.

Functional Interaction Between GABAergic Neurons in the Ventral Tegmental Area and Serotonergic Neurons in the Dorsal Raphe Nucleus

Article

Full-text available

May 2022

GABAergic neurons in the ventral tegmental area (VTA) have brain-wide projections and are involved in multiple behavioral and physiological functions. Here, we revealed the responsiveness of Gad67+ neurons in VTA (VTAGad67+) to various neurotransmitters involved in the regulation of sleep/wakefulness by slice patch clamp recording. Among the substances tested, a cholinergic agonist activated, but serotonin, dopamine and histamine inhibited these neurons. Dense VTAGad67+ neuronal projections were observed in brain areas regulating sleep/wakefulness, including the central amygdala (CeA), dorsal raphe nucleus (DRN), and locus coeruleus (LC). Using a combination of electrophysiology and optogenetic studies, we showed that VTAGad67+ neurons inhibited all neurons recorded in the DRN, but did not inhibit randomly recorded neurons in the CeA and LC. Further examination revealed that the serotonergic neurons in the DRN (DRN5–HT) were monosynaptically innervated and inhibited by VTAGad67+ neurons. All recorded DRN5–HT neurons received inhibitory input from VTAGad67+ neurons, while only one quarter of them received inhibitory input from local GABAergic neurons. Gad67+ neurons in the DRN (DRNGad67+) also received monosynaptic inhibitory input from VTAGad67+ neurons. Taken together, we found that VTAGad67+ neurons were integrated in many inputs, and their output inhibits DRN5–HT neurons, which may regulate physiological functions including sleep/wakefulness.

Rewiring of the Serotonin System in Major Depression

Article

Full-text available

Dec 2021

Serotonin is a key neurotransmitter that is implicated in a wide variety of behavioral and cognitive phenotypes. Originating in the raphe nuclei, 5-HT neurons project widely to innervate many brain regions implicated in the functions. During the development of the brain, as serotonin axons project and innervate brain regions, there is evidence that 5-HT plays key roles in wiring the developing brain, both by modulating 5-HT innervation and by influencing synaptic organization within corticolimbic structures. These actions are mediated by 14 different 5-HT receptors, with region- and cell-specific patterns of expression. More recently, the role of the 5-HT system in synaptic re-organization during adulthood has been suggested. The 5-HT neurons have the unusual capacity to regrow and reinnervate brain regions following insults such as brain injury, chronic stress, or altered development that result in disconnection of the 5-HT system and often cause depression, anxiety, and cognitive impairment. Chronic treatment with antidepressants that amplify 5-HT action, such as selective serotonin reuptake inhibitors (SSRIs), appears to accelerate the rewiring of the 5-HT system by mechanisms that may be critical to the behavioral and cognitive improvements induced in these models. In this review, we survey the possible 5-HT receptor mechanisms that could mediate 5-HT rewiring and assess the evidence that 5-HT-mediated brain rewiring is impacting recovery from mental illness. By amplifying 5-HT-induced rewiring processes using SSRIs and selective 5-HT agonists, more rapid and effective treatments for injury-induced mental illness or cognitive impairment may be achieved.

Résistance à l’Insuline du Système Sérotoninergique : Implication dans les Troubles de l’Humeur Comorbides au Diabete De Type 2

Thesis

Jun 2021

Hugo Martin

Les études épidémiologiques mettent en évidence que les patients atteint de diabète de type 2 (DT2) ont deux fois plus de risque de souffrir de dépression majeure (DM), un trouble mental caractérisé par une tristesse intense et/ou une anhédonie. Plus précisément, les données de la littérature indiquent que la résistance à l’insuline, qui est la caractéristique majeure du DT2, est positivement corrélée à la sévérité des symptômes dépressifs. Etant donné le rôle essentiel que joue la neurotransmission sérotoninergique (5-HT) dans la physiopathologie de la DM, nous avons émis l’hypothèse que la résistance à l’insuline sélective de ce système neuronal est responsable des troubles de l’humeur associés au DT2. En ce sens, des altérations du système 5-HT ont été observées dans un modèle de troubles émotionnels associés à un DT2. En effet, le régime obésogène utilisé dans ce modèle induit une altération significative des propriétés électrophysiologiques des neurones 5-HT du Raphé dorsal (DR) ainsi qu’une diminution des taux de sérotonine. Cette étude vise également à déterminer l’action de l’insuline cérébrale sur le comportement émotionnel et le système sérotoninergique. En utilisant une approche par électrophysiologie ex-vivo, nous avons pu observer que l’insuline modulait positivement l’activité des neurones 5-HT du RD. Nous avons également pu mettre en évidence un effet de type anxiolytique de l’action de l’insuline sur le cerveau par voie intranasale. Cet effet est accompagné de diminution des taux de sérotonine (5-HT) tissulaires dans certaines structures cérébrales impliquées dans la régulation de l’anxiété. Enfin, nous avons utilisé un modèle transgénique grâce auquel nous avons invalidé sélectivement le récepteur à l’insuline dans les neurones 5-HT. L’étude comportementale a permis de mettre en évidence une diminution du comportement de type anxieux chez ces animaux, associé à une diminution de l’activité des neurones sérotoninergiques du RD. Ensemble ces données suggèrent que l’insuline peut moduler le comportement émotionnel notamment via le système sérotoninergique. Ces éléments pourront contribuer à la possible découverte de nouveaux traitements et à la prise en charge des troubles de l’humeur chez les patients atteint de DT2.

Integrating the Monoamine and Cytokine Hypotheses of Depression: Is histamine the missing link?

Article

Jul 2021

Psychiatric diseases, like depression, largely affect the central nervous system (CNS). While the underlying neuropathology of depressive illness remains to be elucidated, several hypotheses have been proposed as molecular underpinnings for major depressive disorder, including the monoamine hypothesis and the cytokine hypotheses. The monoamine hypothesis has been largely supported by the pharmaceuticals that target monoamine neurotransmitters as a treatment for depression. However, these antidepressants have come under scrutiny due to their limited clinical efficacy, side effects and delayed onset of action. The more recent, cytokine hypothesis of depression is supported by the ability of immune-active agents to induce ‘sickness behavior’ akin to that seen with depression. However, treatments that more selectively target inflammation have yielded inconsistent anti-depressive results. As such, neither of these hypotheses can fully explain depressive illness pathology, implying that the underlying neuropathological mechanisms may encompass aspects of both theories. The goal of the current review is to integrate these two well-studied hypotheses and to propose a role for histamine as a potential unifying factor that links monoamines to cytokines. Additionally, we will focus on stress-induced depression, to provide an updated perspective of depressive illness research and thereby identify new potential targets for the treatment of major depressive disorder.

Serotonergic regulation of the dopaminergic system: Implications for reward-related functions

Article

Jun 2021
NEUROSCI BIOBEHAV R

Serotonin is a critical neuromodulator involved in development and behavior. Its role in reward is however still debated. Here, we first review classical studies involving electrical stimulation protocols and pharmacological approaches. Contradictory results on the serotonergic’ involvement in reward emerge from these studies. These differences might be ascribable to either the diversity of cellular types within the raphe nuclei or/and the specific projection pathways of serotonergic neurons. We continue to review more recent work, using optogenetic approaches to activate serotonergic cells in the Raphe to VTA pathway. From these studies, it appears that activation of this pathway can lead to reinforcement learning mediated through the excitation of dopaminergic neurons by serotonergic neurons co-transmitting glutamate. Finally, given the importance of serotonin during development on adult emotion, the effect of abnormal early-life levels of serotonin on the dopaminergic system will also be discussed. Understanding the interaction between the serotonergic and dopaminergic systems during development and adulthood is critical to gain insight into the specific facets of neuropsychiatric disorders.

Reciprocal Lateral Hypothalamic and Raphe GABAergic Projections Promote Wakefulness

Article

Full-text available

Apr 2021
J NEUROSCI

The lateral hypothalamus (LH), together with multiple neuromodulatory systems of the brain, such as the dorsal raphe nucleus (DR), is implicated in arousal, yet interactions between these systems are just beginning to be explored. Using a combination of viral tracing, circuit mapping, electrophysiological recordings from identified neurons, and combinatorial optogenetics in mice, we show that GABAergic neurons in the LH selectively inhibit GABAergic neurons in the DR, resulting in increased firing of a substantial fraction of its neurons that ultimately promotes arousal. These DR GABA neurons are wake active and project to multiple brain areas involved in the control of arousal, including the LH, where their specific activation potently influences local network activity leading to arousal from sleep. Our results show how mutual inhibitory projections between the LH and the DR promote wakefulness and suggest a complex arousal control by intimate interactions between long-range connections and local circuit dynamics. SIGNIFICANCE STATEMENT: Multiple brain systems including the lateral hypothalamus and raphe serotonergic system are involved in the regulation of the sleep/wake cycle, yet the interaction between these systems have remained elusive. Here we show that mutual disinhibition mediated by long range inhibitory projections between these brain areas can promote wakefulness. The main importance of this work relies in revealing the interaction between a brain area involved in autonomic regulation and another in controlling higher brain functions including reward, patience, mood and sensory coding.

Disinhibition mediated by reciprocal hypothalamic and raphé GABAergic projections promotes wakefulness

Preprint

Full-text available

Nov 2020

The lateral hypothalamus (LH), together with multiple neuromodulatory systems of the brain, such as the dorsal raphé nucleus (DR), is implicated in arousal, yet interactions between these systems are just beginning to be explored. Using a combination of viral tracing, circuit mapping, electrophysiological recordings from identified neurons and combinatorial optogenetics, we show that GABAergic neurons in the LH selectively inhibit GABAergic neurons in the DR resulting in disinhibition of a substantial fraction of its neurons that ultimately promotes arousal. These DR GABAergic neurons are wake active and project to multiple brain areas involved in the control of arousal including the LH, where their specific activation potently influences local network activity leading to arousal from sleep. Our results show how mutual inhibitory projections between the LH and the DR promote wakefulness through disinhibitory mechanisms and suggest a complex arousal control by intimate interactions between long-range connections and local circuit dynamics.

MC4R Signaling in Dorsal Raphe Nucleus Controls Feeding, Anxiety, and Depression

Article

Full-text available

Oct 2020

Major depressive disorder is associated with weight loss and decreased appetite; however, the signaling that connects these conditions is unclear. Here, we show that MC4R signaling in the dorsal raphe nucleus (DRN) affects feeding, anxiety, and depression. DRN infusion of α-MSH decreases DRN neuronal activation and feeding. DRN MC4R is expressed in GABAergic PRCP-producing neurons. DRN selective knockdown of PRCP (PrcpDRNKD), an enzyme inactivating α-MSH, decreases feeding and DRN neuronal activation. Interestingly, PrcpDRNKD mice present lower DRN serotonin levels and depressive-like behavior. Similarly, PRCP-ablated MC4R mice (PrcpMC4RKO) show metabolic and behavioral phenotypes comparable to those of PrcpDRNKD mice. Selective PRCP re-expression in DRN MC4R neurons of PrcpMC4RKO mice partially reverses feeding, while fully restoring mood behaviors. Chemogenetic inhibition of DRN MC4R neurons induces anxiety, depression, and reduced feeding, whereas chemogenetic activation reverses these effects. Our results indicate that MC4R signaling in DRN plays a role in feeding, anxiety, and depression.

Sensitivity of functional connectivity to periaqueductal gray localization, with implications for identifying disease-related changes in chronic visceral pain: A MAPP Research Network neuroimaging study

Article

Full-text available

Sep 2020

Previous studies examining the resting-state functional connectivity of the periaqueductal gray (PAG) in chronic visceral pain have localized PAG coordinates derived from BOLD responses to provoked acute pain. These coordinates appear to be several millimeters anterior of the anatomical location of the PAG. Therefore, we aimed to determine whether measures of PAG functional connectivity are sensitive to the localization technique, and if the localization approach has an impact on detecting disease-related differences in chronic visceral pain patients. We examined structural and resting-state functional MRI (rs-fMRI) images from 209 participants in the Multidisciplinary Approach to the Study of Chronic Pelvic Pain (MAPP) Research Network study. We applied three different localization techniques to define a region-of-interest (ROI) for the PAG: 1) a ROI previously-published as a Montreal Neurological Institute (MNI) coordinate surrounded by a 3 mm radius sphere (MNI-sphere), 2) a ROI that was hand-traced over the PAG in a MNI template brain (MNI-trace), and 3) a ROI that was hand-drawn over the PAG in structural images from 30 individual participants (participant-trace). We compared the correlation among the rs-fMRI signals from these PAG ROIs, as well as the functional connectivity of these ROIs with the whole brain. First, we found important non-uniformities in brainstem rs-fMRI signals, as rs-fMRI signals from the MNI-trace ROI were significantly more similar to the participant-trace ROI than to the MNI-sphere ROI. We then found that choice of ROI also impacts whole-brain functional connectivity, as measures of PAG functional connectivity throughout the brain were more similar between MNI-trace and participant-trace compared to MNI-sphere and participant-trace. Finally, we found that ROI choice impacts detection of disease-related differences, as functional connectivity differences between pelvic pain patients and healthy controls were much more apparent using the MNI-trace ROI compared to the MNI-sphere ROI. These results indicate that the ROI used to localize the PAG is critical, especially when examining brain functional connectivity changes in chronic visceral pain patients.

New insights into limbic system roles in the myocardial effects of hypertension

Article

Jan 2017

There is a strong association between abnormalities in central nervous system structure and function and increased risk for the development of hypertension. Comprehensive clinical studies and research using animal models, including brain imaging studies, have revealed effects of insula metabolites on hypertension associated with OSA. Alterations in the limbic system of the central nervous system are closely linked both to hypertension status and to myocardial changes that result from pressure overload. While therapy, including continuous positive airway pressure, have shown excellent results, new studies delineating mechanisms of action are needed. This review article summarizes current concepts regarding central nervous system connections to hypertension and cardiac pathology associated with obstructive sleep apnea (OSA). The limbic system provides one such novel mechanism for resistant hypertension and related cardiac diseases that may be amenable to treatment in the setting of OSA.

Whole-Brain Longitudinal Profiling of Serotonergic Reuptake Inhibition

Preprint

Full-text available

Aug 2020

The serotonergic system is widely implicated in affect regulation, and a common target for psychopharmacological interventions. Selective Serotonin Reuptake Inhibitors (SSRIs) are the foremost drug class for treating depression, as well as anxiety, phobia and other affective disorders. However, the functional mechanisms determining SSRI efficacy remain elusive, hindering the targeted further development of serotonergic system interventions. Assays for longitudinal whole-brain interrogation of the serotonergic system are unavailable, yet such techniques are essential for identifying differential intervention effects across projection areas. We present a novel longitudinal opto-fMRI assay suitable for imaging longitudinal drug treatment effects on the mouse serotonergic system --- within-subject and with sub-millimetre spatial resolution. We apply this assay to a longitudinal fluoxetine treatment, and document reliable segmentation of brain-wide treatment effects, including identification of a brainstem cluster with a highly significant longitudinal trajectory, constituting a novel neurophenotype for psychopharmacological interventions. We differentiate serotonergic neuron activation from projection area activation, and offer brain-wide fMRI evidence for the prominent autoinhibition down-regulation theory of SSRI effects. Further, we show that given the sensitivity of the assay, SSRI treatment produces no persistent effects after treatment cessation in healthy subjects.

Modulation of extracellular noradrenaline in rat cortex by selective serotonin reuptake inhibitors (SSRIs)

Thesis

Jan 1998

Zoë A Hughes

This research project has investigated the modulation of central noradrenergic function in rat cortex by antidepressant drugs. Particular attention has been paid to the type of antidepressants known as the selective serotonin (5-HT) reuptake inhibitors (SSRIs). The therapeutic effects of these drugs are presently attributed exclusively to their inhibition of neuronal uptake of 5-HT. The aim of this work was to investigate whether the SSRIs, fluoxetine and citalopram, modify central noradrenergic function and, if so, was their site of action on noradrenergic neurones? In order to achieve this two techniques were used: the first used microdialysis to measure the concentration of extracellular noradrenaline in vivo, the second looked at the inhibition of [3H]noradrenaline uptake into rat cortical synaptosomes in vitro. In this thesis changes in the concentration of extracellular noradrenaline caused by local infusion of antidepressants in the frontal cortex of rats, were monitored using microdialysis. Fluoxetine and citalopram, as well as the noradrenaline uptake inhibitor, desipramine, increased noradrenaline efflux. To investigate whether inhibition of noradrenaline uptake could contribute to this increase in efflux, the effects of these drugs on synaptosomal [3H]noradrenaline uptake were studied. Because these drugs inhibit [3H]noradrenaline uptake, it is likely that inhibition of noradrenaline uptake contributes to the increase in noradrenaline efflux in vivo. In view of this finding, the possible site(s) of action of these drugs were investigated. An uptake site targeted by SSRIs could be located on serotonergic neurones. The 5-HT neurotoxin, 5,7-dihydroxytryptamine (5,7-DHT), was used as a tool to investigate this possibility. Because a selective lesion of 5-HT neurones did not affect the inhibition of [3H]noradrenaline uptake by any of the drugs, it is unlikely that fluoxetine, citalopram or desipramine were acting at a site on serotonergic neurones. The second possible site of action investigated was one located on noradrenergic neurones. Pretreatment of rats with the noradrenergic neurotoxin, N-(2-chloroethyl)-N-etiyl-2-broinobenzylamine (DSP-4), modified the effects of these antidepressants on noradrenergic function. Low concentrations of desipramine inhibited a smaller proportion of [3H]noradrenaline uptake than in control rats. In contrast, a greater proportion of uptake in the cortex of DSP-4 lesioned rats was sensitive to inhibition by low concentrations of fluoxetine in vitro. The effects of these drugs on noradrenaline efflux in vivo were also changed after DSP-4 pretreatment. The increase in noradrenaline efflux induced by desipramine was greater in DSP-4 treated rats, whereas the fluoxetine-induced increase, apparent in control rats, was not evident after DSP-4. The effects of citalopram were unaffected by DSP-4 treatment, suggesting that SSRIs do not all have the same pharmacological profile and that fluoxetine, but not citalopram, could act at a site on noradrenergic neurones. One additional, but important finding to emerge from work described in this thesis was that, although DSP-4 treatment caused a 70% reduction in tissue noradrenaline content, the concentration of extracellular noradrenaline was increased nearly 2-fold. Results suggest that neurones which survive DSP-4 treatment have a greater rate of noradrenaline release, moreover, released noradrenaline is taken up via low affinity uptake sites. Overall, the experiments in this thesis have exposed marked effects of SSRIs on central noradrenergic function which could contribute to the efficacy of antidepressants hitherto regarded as acting selectively on serotonergic neurones.

Separable gain control of ongoing and evoked activity in the visual cortex by serotonergic input

Article

Full-text available

Apr 2020

Controlling gain of cortical activity is essential to modulate weights between internal ongoing communication and external sensory drive. Here, we show that serotonergic input has separable suppressive effects on the gain of ongoing and evoked visual activity. We combined optogenetic stimulation of the dorsal raphe nucleus (DRN) with wide-field calcium imaging, extracellular recordings, and iontophoresis of serotonin (5-HT) receptor antagonists in the mouse visual cortex. 5-HT1A receptors promote divisive suppression of spontaneous activity, while 5-HT2A receptors act divisively on visual response gain and largely account for normalization of population responses over a range of visual contrasts in awake and anesthetized states. Thus, 5-HT input provides balanced but distinct suppressive effects on ongoing and evoked activity components across neuronal populations. Imbalanced 5-HT1A/2A activation, either through receptor-specific drug intake, genetically predisposed irregular 5-HT receptor density, or change in sensory bombardment may enhance internal broadcasts and reduce sensory drive and vice versa.

Separable gain control of ongoing and evoked activity in the visual cortex by serotonergic input

Article

Full-text available

Apr 2020

Separable gain control of ongoing and evoked activity in the visual cortex by serotonergic input

Article

Full-text available

Apr 2020
eLife

Neuronal vulnerability in Parkinson disease: Should the focus be on axons and synaptic terminals?

Article

Sep 2019

While current effective therapies are available for the symptomatic control of PD, treatments to halt the progressive neurodegeneration still do not exist. Loss of dopamine neurons in the SNc and dopamine terminals in the striatum drive the motor features of PD. Multiple lines of research point to several pathways which may contribute to dopaminergic neurodegeneration. These pathways include extensive axonal arborization, mitochondrial dysfunction, dopamine's biochemical properties, abnormal protein accumulation of α‐synuclein, defective autophagy and lysosomal degradation, and synaptic impairment. Thus, understanding the essential features and mechanisms of dopaminergic neuronal vulnerability is a major scientific challenge and highlights an outstanding need for fostering effective therapies against neurodegeneration in PD. This article, which arose from the Movement Disorders 2018 Conference, discusses and reviews the possible mechanisms underlying neuronal vulnerability and potential therapeutic approaches in PD. © 2019 International Parkinson and Movement Disorder Society

Serotonergic neurons in the dorsal raphe nucleus mediate the arousal-promoting effect of orexin during isoflurane anesthesia in male rats

Article

Full-text available

Mar 2019
NEUROPEPTIDES

Previous studies have demonstrated that the activation of orexinergic neurons facilitates the recovery of animals from general anesthesia. Moreover, serotonergic neurons that receive projections from orexin neurons have also been shown to participate in sleep-wakefulness regulation. In the present study, we aimed to explore whether orexinergic neurons facilitate emergence from isoflurane anesthesia in rats by activating serotonergic neurons. Orexin A (30 or 100 pmol), orexin B (30 or 100 pmol), and their respective antagonists SB-334867 and TCS-OX2–29 (5 or 20 μg) were microinjected into the dorsal raphe nucleus (DRN) of rats, and their effects on induction and emergence times were analyzed. Electroencephalogram (EEG) changes were also recorded and analyzed to illuminate the effect of orexin injection into the DRN on cortical excitability under isoflurane anesthesia. Activation of serotonergic neurons was detected via immunohistochemical analysis of c-Fos expression following orexin administration. Our results indicated that injection of neither orexins nor orexin antagonists into the rat DRN exerted an impact on induction time, whereas orexin-A injection (100 pmol) enhanced arousal when compared with the saline group. In contrast, administration of orexin receptor type 1 antagonist SB-334867 (20 μg) prolonged emergence time from isoflurane anesthesia. Microinjection of orexin-A induced an arousal pattern on EEG, and decreased the burst suppression ratio under isoflurane anesthesia. Isoflurane anesthesia inhibited the activity of serotonergic neurons, as shown by decrease in the number of c-Fos-immunoreactive serotonergic neurons when compared with the sham group. This inhibitory effect was partially reversed by administration of orexin-A. Taken together, our findings suggest that orexinergic signals facilitate emergence from isoflurane anesthesia, at least partially, by reversing the effects of isoflurane on serotonergic neurons of the DRN.

Ultrastructure of the dorsal claustrum in cat. II. Synaptic organization

Article

May 2019
ACTA HISTOCHEM

Are periaqueductal grey and dorsal raphe the foundation of appetitive and aversive control? A comprehensive review

Article

Feb 2019

Many see the periaqueductal gray (PAG) as a region responsible for the downstream control of defensive reactions. Here we provide a detailed review of anatomical and functional data on the different parts of the PAG together with the dorsal raphe, which completes the circle of periaqueductal nuclei. Based on anatomical features, we propose a new subdivision of the periaqueductal gray that accounts for the distinct characteristics of the area. We provide a comprehensive functional view of the periaqueductal gray, going beyond simple panic and escape to integrate data on fear, anxiety, and depression. Importantly, we conclude that this periaqueductal cluster of nuclei is broadly involved in motivated behavior controlling not only aversive but also appetitive behavior and with some involvement in more complex motivational processes such as approach-avoidance conflict resolution. In sum, these highly conserved nuclei surrounding the aqueduct appear to be the simplest, foundational, elements of integrated motivated goal-directed control of all types.

Serotonin actions within the prelimbic cortex induce anxiolysis mediated by serotonin 1a receptors

Article

Dec 2018
J PSYCHOPHARMACOL

Background Serotonin plays an important role in the regulation of anxiety, acting through complex modulatory mechanisms within distinct brain structures. Serotonin can act through complex negative feedback mechanisms controlling the neuronal activity of serotonergic circuits and downstream physiologic and behavioral responses. Administration of serotonin or the serotonin 1A receptor agonist, (±)-8-hydroxy-2-(dipropylamino)tetralin (8-OH-DPAT), into the prefrontal cortex, inhibits anxiety-like responses. The prelimbic area of the prefrontal cortex regulates serotonergic neurons within the dorsal raphe nucleus and is involved in modulating anxiety-like behavioral responses. Aims This study aimed to investigate the serotonergic role within the prelimbic area on anxiety- and panic-related defensive behavioral responses. Methods We investigated the effects of serotonin within the prelimbic area on inhibitory avoidance and escape behaviors in the elevated T-maze. We also extended the investigation to serotonin 1A, 2A, and 2C receptors. Results Intra-prelimbic area injection of serotonin or 8-OH-DPAT induced anxiolytic effects without affecting escape behaviors. Previous administration of the serotonin 1A receptor antagonist, WAY-100635, into the prelimbic area counteracted the anxiolytic effects of serotonin. Neither the serotonin 2A nor the serotonin 2C receptor preferential agonists, (±)-2,5-dimethoxy-4-iodoamphetamine (DOI) and 6-chloro-2-(1-piperazinyl) pyrazine (MK-212), respectively, affected behavioral responses in the elevated T-maze. Conclusion Facilitation of serotonergic signaling within the prelimbic area of rats induced an anxiolytic effect in the elevated T-maze test, which was mediated by local serotonin 1A receptors. This inhibition of anxiety-like defensive behavioral responses may be mediated by prelimbic area projections to neural systems controlling anxiety, such as the dorsal raphe nucleus or basolateral amygdala.

Subtraction and division of visual cortical population responses by the serotonergic system

Preprint

Oct 2018

Normalization is a fundamental operation throughout neuronal systems to adjust dynamic range. In the visual cortex various cell circuits have been identified that provide the substrate for such a canonical function, but how these circuits are orchestrated remains unclear. Here we suggest the serotonergic (5-HT) system as another player involved in normalization. 5-HT receptors of different classes are co-distributed across different cortical cell types, but their individual contribution to cortical population responses is unknown. We combined wide-field calcium imaging of primary visual cortex (V1) with optogenetic stimulation of 5-HT neurons in mice dorsal raphe nucleus (DRN) — the major hub for widespread release of serotonin across cortex — in combination with selective 5-HT receptor blockers. While inhibitory (5-HT1A) receptors accounted for subtractive suppression of spontaneous activity, depolarizing (5-HT2A) receptors promoted divisive suppression of response gain. Added linearly, these components led to normalization of population responses over a range of visual contrasts.

Potentiation of Glutamatergic Synaptic Transmission Onto Dorsal Raphe Serotonergic Neurons in the Valproic Acid Model of Autism

Article

Full-text available

Oct 2018

Autism spectrum disorder (ASD) is characterized by social and communicative impairments and increased repetitive behaviors. These symptoms are often comorbid with increased anxiety. Prenatal exposure to valproic acid (VPA), an anti-seizure and mood stabilizer medication, is a major environmental risk factor of ASD. Given the important role of the serotonergic (5-HT) system in anxiety, we examined the impact of prenatal VPA exposure on the function of dorsal raphe nucleus (DRn) 5-HT neurons. We found that male rats prenatally exposed to VPA exhibited increased anxiety-like behaviors revealed by a decreased time spent on the open arms of the elevated plus maze. Prenatal VPA exposed rats also exhibited a stereotypic behavior as indicated by excessive self-grooming in a novel environment. These behavioral phenotypes were associated with increased electrical activity of putative DRn 5-HT neurons recorded in vitro. Examination of underlying mechanisms revealed that prenatal VPA exposure increased excitation/inhibition ratio in synapses onto these neurons. The effect was mainly mediated by enhanced glutamate but not GABA release. We found reduced paired-pulse ratio (PPR) of evoked excitatory postsynaptic currents (EPSCs) and increased frequency but not amplitude of miniature EPSCs in VPA exposed rats. On the other hand, presynaptic GABA release did not change in VPA exposed rats, as the PPR of evoked inhibitory postsynaptic currents was unaltered. Furthermore, the spike-timing-dependent long-term potentiation at the glutamatergic synapses was occluded, indicating glutamatergic synaptic transmission is maximized. Lastly, VPA exposure did not alter the intrinsic membrane properties of DRn 5-HT neurons. Taken together, these results indicate that prenatal VPA exposure profoundly enhances glutamatergic synaptic transmission in the DRn and increases spontaneous firing in DRn 5-HT neurons, which could lead to increased serotonergic tone and underlie the increased anxiety and stereotypy after prenatal VPA exposure.

Molecular guidance of serotonin raphe neurons during development

Thesis

Sep 2016

Teng Teng

In mice, serotonin (5-HT) midbrain neurons are born from embryonic day 10 to 12, and start extending axons, shortly after neurogenesis, both rostrally to the telencephalon and caudally to the brainstem. These projections are highly collateralized but with some degree of topographic organization. In the telencephalon, the pattern of 5-HT innervation arising from the dorsal (B7, B6) or the medial (B5-B8) nuclei differs. However, there are no systematic detailed developmental studies in mice, which are the most extensively used model, in particular for genetic studies. Such data are important to gather in order to analyze the effects of mouse mutations on defined molecular pathway of serotonin neurons. Moreover the guidance molecules that direct these 5-HT raphe neurons to different targets are not known. We performed several studies of 5-HT innervation aimed at detecting how the dorsal and median raphe nuclei are targeted to different forebrain regions during development. We investigated the role of ephrinA-EphA signaling in selective targeting. Our results demonstrate that EphA5 mRNA is selectively expressed in distinct subpopulation of serotonin raphe neurons. Particularly, EphA5 exhibited the highest level in dorsal raphe serotonin neurons (B7). The results of in vitro explant cultures and in vivo electroporation analyses indicated that the ligands of EphA5 (ephrinA5 and ephrinA3) act as repellent factors for the serotonergic axon growth cones. Anterograde tracing in the ephrinA5 -/- mice showed mistargeting of dorsal raphe neurons projections, including the serotonergic projection. Particularly, our analysis of tracing studies show that targeting of the dorsal and median raphe axons to different layers of the olfactory bulb is altered in the ephrinA5 KO. However we do not know at what developmental stage these alterations occur, in particular whether this reflects an alteration in the orientation of ascending fiber tracts, or whether this reflects late developmental maturation when raphe axons collateralize and branch in specific target regions. We have taken advantage a new morphological method, which allows analyzing immunocytochemical labeling in 3_D. 5-HT immunolabeling, in whole brain serotonergic projection in 3_D. Our findings show that serotonergic fibers projecting to olfactory bulb require a special timing to enter the target. The expression pattern of ephrinA5 suggests that ephrinA5 can be one of the factors that modulate this timing. Overall, our results show for the first time the implication a guidance molecule for the region-specific and time-specific targeting of serotonin raphe neurons and has implications for the anatomo-functional parsing of raphe cell groups.

Serotonin transporter inhibition during neonatal period induces sex-dependent effects on mitochondrial bioenergetics in the rat brainstem

Article

May 2018

The serotonin reuptake is mainly regulated by the serotonin transporters (SERTs), which are abundantly found in the raphe nuclei, located in the brainstem. Previous studies have shown that dysfunction in the SERT has been associated with several disorders, including depression and cardiovascular diseases. In this manuscript, we aimed to investigate how gender and the treatment with a serotonin selective reuptake inhibitor (SSRI) could affect mitochondrial bioenergetics and oxidative stress in the brainstem of male and female rats. Fluoxetine, our chosen SSRI, was used during the neonatal period (i.e. from post‐natal day 1 to post‐natal day 21 ‐ PND1 to PND21) in both male and female animals. Thereafter, experiments were conducted in adult rats (60 days old). Our results demonstrate that during lactation, fluoxetine treatment modulates the mitochondrial bioenergetics in a sex‐dependent manner, such as improving male mitochondrial function and female antioxidant capacity. This article is protected by copyright. All rights reserved.

TRK-fused Gene Protein Localization Is Prominent in Serotonergic and Noradrenergic Cell Groups, and Some Lower Motor Neurons in the Corticospinal Tract of the Rat Brainstem

Article

Full-text available

May 2018

The TRK-fused gene (TFG) is reported to be involved in the regulation of cell size, apoptosis, cell growth, ER-Golgi protein secretion, NF-κβ pathway signaling, the ubiquitin-proteasome system, and pancreatic β-cell mass and function. TFG mutations were reported in some neurodegenerative diseases affecting sensory and motor functions. However, the function of TFG in the nervous system and how TFG mutations lead to neurodegeneration remain unclear. In this study, we employed double immunohistochemistry to investigate the details of TFG localization patterns in monoaminergic and cholinergic neurons in the brainstem. Intense TFG immunoreactivity was observed in the dorsal raphe nucleus, the locus coeruleus, and the ventral horn of the spinal cord. TFG immunoreactivity was observed in some serotonergic neurons in all B1–B9 cell groups, and some noradrenergic neurons in all A1–A7 cell groups in the rat brainstem, while no immunoreactivity was observed in the dopaminergic neurons in A8–A10 cell groups. TFG immunoreactivity was observed in all ChAT-positive motor nuclei in the lower corticospinal tract of the rat brainstem.

Aggression modulator: Understanding the multifaceted role of the dorsal raphe nucleus

Article

Feb 2024
BIOESSAYS

Aggressive behavior is instinctively driven behavior that helps animals to survive and reproduce and is closely related to multiple behavioral and physiological processes. The dorsal raphe nucleus (DRN) is an evolutionarily conserved midbrain structure that regulates aggressive behavior by integrating diverse brain inputs. The DRN consists predominantly of serotonergic (5‐HT:5‐hydroxytryptamine) neurons and decreased 5‐HT activity was classically thought to increase aggression. However, recent studies challenge this 5‐HT deficiency model, revealing a more complex role for the DRN 5‐HT system in aggression. Furthermore, emerging evidence has shown that non‐5‐HT populations in the DRN and specific neural circuits contribute to the escalation of aggressive behavior. This review argues that the DRN serves as a multifaceted modulator of aggression, acting not only via 5‐HT but also via other neurotransmitters and neural pathways, as well as different subsets of 5‐HT neurons. In addition, we discuss the contribution of DRN neurons in the behavioral and physiological aspects implicated in aggressive behavior, such as arousal, reward, and impulsivity, to further our understanding of DRN‐mediated aggression modulation.

Wiring and Volume Transmission: An Overview of the Dual Modality for Serotonin Neurotransmission

Article

Full-text available

Nov 2023

Serotonin is a neurotransmitter involved in the modulation of a multitude of physiological and behavioral processes. In spite of the relatively reduced number of serotonin-producing neurons present in the mammalian CNS, a complex long-range projection system provides profuse innervation to the whole brain. Heterogeneity of serotonin receptors, grouped in seven families, and their spatiotemporal expression pattern account for its widespread impact. Although neuronal communication occurs primarily at tiny gaps called synapses, wiring transmission, another mechanism based on extrasynaptic diffusion of neuroactive molecules and referred to as volume transmission, has been described. While wiring transmission is a rapid and specific one-to-one modality of communication, volume transmission is a broader and slower mode in which a single element can simultaneously act on several different targets in a one-to-many mode. Some experimental evidence regarding ultrastructural features, extrasynaptic localization of receptors and transporters, and serotonin–glia interactions collected over the past four decades supports the existence of a serotonergic system of a dual modality of neurotransmission, in which wiring and volume transmission coexist. To date, in spite of the radical difference in the two modalities, limited information is available on the way they are coordinated to mediate the specific activities in which serotonin participates. Understanding how wiring and volume transmission modalities contribute to serotonergic neurotransmission is of utmost relevance for the comprehension of serotonin functions in both physiological and pathological conditions.

Techniques for in vivo serotonin detection in the brain: State of the art

Article

Full-text available

Jun 2023
J NEUROCHEM

Neuronal circuits in the brain that utilize the neurotransmitter serotonin are essential to the regulation of mood and emotional expression. Disruptions in serotonin signaling underlie neuropsychiatric conditions such as depression and anxiety. However, the cellular mechanisms that regulate serotonergic signaling in the brain in healthy and diseased states remain to be better understood. In particular, as more is learned about serotonin in the brain, we recognize an urgent need to develop techniques capable of mapping its complex spatiotemporal dynamics in awake, behaving animals. Notably, analytical methods to detect serotonin in situ, including tomography, are widely used but still recognized as limited in terms of their spatiotemporal resolution, their methodological caveats, and their technical limitations when cross‐referenced with behavioral studies. To overcome such limitations, genetically encoded serotonin indicators were developed, leading to the introduction of novel imaging modalities that enable researchers to achieve remarkable spatiotemporal resolution in the study of serotonergic circuits in preclinical models of neuropsychiatric disorders. These novel approaches, while remarkably powerful, are also not without limitations. Here, we review the current techniques for detecting and quantifying serotonin in vivo within the brain and discuss how novel approaches such as genetically encoded serotonin indicators will lead to new insights into the roles of serotonergic circuits in health and disease. image

Stem Cell Types for Treating Depression

Article

Full-text available

Mar 2023

Transcranial ultrasound stimulation relieves depression in mice with chronic restraint stress

Article

Full-text available

Aug 2022

Objective: Exhaustion of Serotonin (5-hydroxytryptamine, 5-HT) is a typical cause of the depression disorder's development and progression, including depression-like behaviors. Transcranial ultrasound stimulation (TUS) is an emerging non-invasive neuromodulation technique treating various neurodegenerative diseases. This study aims to investigate whether TUS ameliorates depression-like behaviors by restoring 5-HT levels. Methods: The depression model mice are established by chronic restraint stress (CRS). Ultrasound waves (FF = 1.1MHz, PRF = 1000 Hz, TBD = 0.5 ms, SD = 1 s, ISI = 1 s, and DC = 50%) were delivered into the dorsal raphe nucleus (DRN) for 30 min per day for two weeks. Depression-like behavior changes are evaluated with the sucrose preference and tail suspension tests. Liquid chromatography-mass spectrometry is performed to quantitatively detect the concentration of 5-HT in the DRN to explore its potential mechanism. The effectiveness and safety of TUS assessed by c-Fos immunofluorescence and hematoxylin and eosin (HE) staining, respectively. Results: Three weeks after CRS, 22 depressive mice models were screened by sucrose preference index (SPI). After two weeks of ultrasound stimulation of the DRN (DRN-TUS) in depressive mice, the SPI was increased (p = 0.1527) and the tail suspension immobility duration was significantly decreased (p = 0.0038) compared with the non-stimulated group. In addition, TUS significantly enhances the c-Fos (p = 0.05) positive cells' expression and the 5-HT level (p = 0.0079) in the DRN. Importantly, HE staining shows no brain tissue damage. Conclusion: These results indicate that DRN-TUS has safely and effectively improved depression-like behaviors including anhedonia and hopelessness, potentially by reversing the depletion of 5-TH. Significance: TUS may provide a new perspective on depression therapy, possibly through restoring monoamine levels.

Dorsal Raphe Nucleus Serotoninergic Neurons Mediate Morphine Rewarding Effect and Conditioned Place Preference

Article

Nov 2021
NEUROSCIENCE

Morphine rewarding properties are the main reasons for drug-craving in behaviors occurring during morphine addiction. It has been suggested that morphine addiction relies on signals to the mesolimbic dopamine system, although the mechanisms outside the dopaminergic system are still unclear. Notably, the role of the dorsal raphe nucleus (DRN) serotoninergic (5-hydroxytryptamine, 5-HT) system remains unexplored. Using in vivo electrophysiological and optogenetic approaches, we found that morphine treatment increased DRN 5-TH neurons firing rate and optogenetic activation of DRN 5-HT neurons induced a rewarding effect, indicating that morphine reward is related to DRN 5-HT neurons. Accordingly, optogenetic inhibition of DRN 5-HT neurons following morphine injection reversed conditioned place preference (CPP) during chronic morphine treatment. These findings aid our understanding of the new functions of the DRN 5-HT neurons for morphine rewarding effect and provide a potential approach for the treatment of morphine addiction.

monoamines as brain state switches

Article

Full-text available

Oct 2021

Sayed A Azizi

Etude du rôle des altérations des cycles éveil/sommeil dans la progression de la maladie de Parkinson et de l'atrophie multisystématisée

Thesis

Full-text available

Mar 2021

Marine Persillet

La maladie de Parkinson (MP) et l’atrophie multisystématisée (AMS) sont des synucléinopathies caractérisées par la perte des neurones dopaminergiques (DA) dans la substance noire pars compacta (SNc) et par la présence d’inclusions cytoplasmiques, appelés corps de Lewy (LB) et inclusions cytoplasmiques gliales (GCI) respectivement et constitués notamment d’alpha-synucléine (a-syn) mal conformée. En plus des troubles moteurs, les patients MP et AMS présentent de nombreux symptômes non-moteurs dont les altérations des cycles éveil/sommeil qui peuvent apparaitre de manière précoce et qui sont même considérées comme prédictifs du développement de synucléinopathies. Le but de ma thèse est donc d’étudier le possible lien entre la progression de la neurodégénérescence, la progression de la pathologie liée à l’a-syn et l’apparition d’altérations des cycles éveil/sommeil.Tout d’abord, comme la perte des neurones DA est une caractéristique des synucleinopathies, nous avons étudié le sommeil en condition de déplétion de la dopamine. Le sommeil des souris a été enregistré après injection de réserpine et d’alpha méthyltyrosine qui induisent respectivement la forte réduction des stocks de monoamines, incluant la dopamine, et l’inhibition de la tyrosine hydroxylase et donc la synthèse de la dopamine.Alors que les modèles basés sur l’injection de neurotoxines présentent à la fois les symptômes moteurs et non-moteurs de la MP, ils ne reproduisent pas le processus du développement de la neurodégénérescence requis pour ce projet. Nous avons donc utilisés des modèles la dégénérescence de la MP développés récemment appelés souris LB et singe LB. Ces modèles sont adaptés pour étudier de possibles altérations des cycles éveil/sommeil comme la pathologie progresse lentement et qu’elle est liée à l’a-¬syn. Les souris sauvages ont reçues une injection dans la SNc de fractions LB contenant de l’a-syn pathologique. L’injection de ces fractions, extraites du cerveau de patients MP, induisent une perte progressive des neurones DA. Les souris contrôles sont injectées avec du sucrose. Ensuite, un appareil permettant l’enregistrement de l’activité neuronale corticale (électroencéphalographie, EEG) et la contraction des muscles du cou (électromyographie, EMG) est implanté sur les souris pour discriminer les différents stades des cycles éveil/sommeil : éveil, sommeil avec mouvements oculaires non-rapides (sommeil NREM), et sommeil avec mouvements oculaires rapides (sommeil REM). Les primates non-humains ont été injectés dans le striatum avec des fractions LB (singes LB), ou du sucrose. Ils ont ensuite été appareillés pour l’enregistrement EEG, EMG, et pour l’enregistrement de l’activité électrique des yeux (électrooculographie, EOG) pour distinguer l’éveil, le sommeil NREM (sommeil lent et profond) et le sommeil REM.Concernant l’AMS, le modèle utilisé est une souris transgénique qui surexprime l’a-syn humaine sous le contrôle d’un promoteur glial (souris PLP). Ce modèle présente des troubles des fonctions autonomes, un critère clé dans le diagnostic de l’AMS. Là aussi la maladie progresse lentement et est liée à l’a-syn. Comme avec les souris LB, un appareil d’enregistrement est implanté aux souris PLP et aux souris contrôles pour enregistrer les phases d’éveil, de sommeil NREM et de sommeil REM.Enfin, comme le sommeil a un rôle dans l’élimination protéique, nous avons étudier son rôle dans la pathologie liée à l’a-syn. Des expériences de microdialyse ont été réalisées pour quantifier l’a-syn dans le LCR de souris LB et de souris contrôles en conditions normales ou après privation de sommeil.Le travail réalisé est particulièrement pertinent car comprendre si les altérations des cycles éveil/sommeil peuvent servir au niveau expérimental de marqueur de la progression de la pathologie pourrait permettre une détection précoce et amener à de nouvelles stratégies thérapeutiques pour ralentir sa progression.

Bridging the gap between single receptor type activity and whole‐brain dynamics

Article

Full-text available

Apr 2021
FEBS J

What is the effect of activating a single modulatory neuronal receptor type on entire brain network dynamics? Can such effect be isolated at all? These are important questions because characterizing elementary neuronal processes that influence network activity across the given anatomical backbone is fundamental to guide theories of brain function. Here, we introduce the concept of the cortical ‘receptome’ taking into account the distribution and densities of expression of different modulatory receptor types across the brain's anatomical connectivity matrix. By modelling whole‐brain dynamics in silico, we suggest a bidirectional coupling between modulatory neurotransmission and neuronal connectivity hardware exemplified by the impact of single serotonergic (5‐HT) receptor types on cortical dynamics. As experimental support of this concept, we show how optogenetic tools enable specific activation of a single 5‐HT receptor type across the cortex as well as in vivo measurement of its distinct effects on cortical processing. Altogether, we demonstrate how the structural neuronal connectivity backbone and its modulation by a single neurotransmitter system allow access to a rich repertoire of different brain states that are fundamental for flexible behaviour. We further propose that irregular receptor expression patterns—genetically predisposed or acquired during a lifetime—may predispose for neuropsychiatric disorders like addiction, depression and anxiety along with distinct changes in brain state. Our long‐term vision is that such diseases could be treated through rationally targeted therapeutic interventions of high specificity to eventually recover natural transitions of brain states.

Behavioral arrest and a characteristic slow waveform are hallmark responses to selective 5-HT2A receptor activation

Article

Full-text available

Jan 2021

Perception, emotion, and mood are powerfully modulated by serotonin receptor (5-HTR) agonists including hallucinogens. The 5-HT2AR subtype has been shown to be central to hallucinogen action, yet the precise mechanisms mediating the response to 5-HT2AR activation remain unclear. Hallucinogens induce the head twitch response (HTR) in rodents, which is the most commonly used behavioral readout of hallucinogen pharmacology. While the HTR provides a key behavioral signature, less is known about the meso level changes that are induced by 5-HT2AR activation. In response to administration of the potent and highly selective 5-HT2AR agonist 25I-NBOH in mice, we observe a disorganization of behavior which includes frequent episodes of behavioral arrest that consistently precede the HTR by a precise interval. By combining behavioral analysis with electroencephalogram (EEG) recordings we describe a characteristic pattern composed of two distinctive EEG waveforms, Phase 1 and Phase 2, that map onto behavioral arrest and the HTR respectively, with the same temporal separation. Phase 1, which underlies behavioral arrest, is a 3.5–4.5 Hz waveform, while Phase 2 is slower at 2.5–3.2 Hz. Nicotine pretreatment, considered an integral component of ritualistic hallucinogen practices, attenuates 25I-NBOH induced HTR and Phase 2 waveforms, yet increases behavioral arrest and Phase 1 waveforms. Our results suggest that in addition to the HTR, behavioral arrest and characteristic meso level slow waveforms are key hallmarks of the response to 5-HT2AR activation. Increased understanding of the response to serotonergic hallucinogens may provide mechanistic insights into perception and hallucinations, as well as regulation of mood.

Monoamines: Dopamine, Norepinephrine, and Serotonin, Beyond Modulation, “Switches” That Alter the State of Target Networks

Article

Full-text available

Dec 2020

Sayed A Azizi

How do monoamines influence the perceptual and behavioral aspects of brain function? A library of information regarding the genetic, molecular, cellular, and function of monoamines in the nervous system and other organs has accumulated. We briefly review monoamines’ anatomy and physiology and discuss their effects on the target neurons and circuits. Monoaminergic cells in the brain stem receive inputs from sensory, limbic, and prefrontal areas and project extensively to the forebrain and hindbrain. We review selected studies on molecular, cellular, and electrophysiological effects of monoamines on the brain’s target areas. The idea is that monoamines, by reversibly modulating the “primary” information processing circuits, regulate and switch the functions of brain networks and can reversibly alter the “brain states,” such as consciousness, emotions, and movements. Monoamines, as the drivers of normal motor and sensory brain operations, including housekeeping, play essential roles in pathogenesis of neuropsychiatric diseases.

Embracing diversity in the 5-HT neuronal system

Article

Apr 2019
NAT REV NEUROSCI

Neurons that synthesize and release 5-hydroxytryptamine (5-HT; serotonin) express a core set of genes that establish and maintain this neurotransmitter phenotype and distinguish these neurons from other brain cells. Beyond a shared 5-HTergic phenotype, these neurons display divergent cellular properties in relation to anatomy, morphology, hodology, electrophysiology and gene expression, including differential expression of molecules supporting co-transmission of additional neurotransmitters. This diversity suggests that functionally heterogeneous subtypes of 5-HT neurons exist, but linking subsets of these neurons to particular functions has been technically challenging. We discuss recent data from molecular genetic, genomic and functional methods that, when coupled with classical findings, yield a reframing of the 5-HT neuronal system as a conglomeration of diverse subsystems with potential to inspire novel, more targeted therapies for clinically distinct 5-HT-related disorders.

Serotonergic Neurons in Vertebrate and Invertebrate Model Organisms (Rodents, Zebrafish, Drosophila melanogaster, Aplysia californica, Caenorhabditis elegans)

Chapter

Jan 2019

Etude de l'hypothermie induite par la méthionine sulfoximine chez le rat : intervention des systèmes sérotoninergiques

Thesis

Nov 1991

Madeleine Ginefri-Gayet

L'injection systémique de la méthionine sulfoximine (MSO), chez le rat, entraîne une encéphalopathie lentement progressive caractérisée par un désordre du mouvement (ataxie) suivi quelques heures plus tard de convulsions toniques et cloniques. Ce modèle est utilisé pour la recherche des mécanismes métaboliques liés aux convulsions. Le but du travail que nous présentons est d'étudier les modalités de la variation de la température interne du rat sous l'action de la MSO, et, de tenter de tirer des conclusions en ce qui concerne les voies neuronales dans le système nerveux central qui peuvent être impliquées dans cette variation.

The distribution of enkephalin-immunoreactive cell bodies in the rat central nervous system

Article

Full-text available

Jul 1977
NEUROSCI LETT

With the indirect immunofluorescence technique the distribution of methionine-enkephalin-immunoreactive cell bodies was studied in the central nervous system of rats pretreated with colchicine. The antiserum used did cross-react to 10% with leucine-enkephalin but to less than 0.1% with alpha-, beta-, and gamma-endorphine. Cell bodies with a specific immunofluorescence were observed in the tel-, di-, mes- and rhombencephalon and in the spinal cord.

Ascending projections of the locus coeruleus in the rat. II. Autoradiographic study

Article

Full-text available

Jun 1977
BRAIN RES

The ascending projections of the locus coeruleus were studied using an autoradiographic method. The major projection of locus coeruleus neurons ascends in a dorsal pathway traversing the midbrain tegmentum in a position ventrolateral to the periaqueductal gray. At the caudal diencephalon the locus coeruleus axons descend to enter the medial forebrain bundle at a caudal tuberal hypothalamic level. They are jointed in the medial forebrain bundle by a much smaller locus coeruleus projection which takes a ventral course through the midbrain tegmentum and enters the medial forebrain bundle via the mammillary peduncle and ventral tegmental area. Terminal projections are evident in the midbrain to the periaqueductal gray, tegmentum and raphe nuclei. There are widespread projections to the dorsal thalamus. The heaviest of these are to the intralaminar nuclei, the anteroventral and anteromedial nuclei, the dorsal lateral geniculate and the paraventricular nucleus. In the hypothalamus the largest projections are to the lateral hypothalamic area, periventricular nucleus, supraoptic nucleus and paraventricular nucleus. As the locus coeruleus projection ascends in the medial forebrain bundle, fibers leave it to traverse the lateral hypothalamus and zona incerta and enter the internal capsule, the ventral amygdaloid bundle and ansa peduncularis. These appear to terminate in the amygdaloid complex and, via the external capsule, in the lateral and dorsal neocortex. At the level of the septum 4 projections are evident. One group of fibers enters the stria medullaris to terminate in the paraventricular nucleus and habenular nuclei. A second group joins the stria terminalis to terminate in the anygdaloid complex. The third group turns into the diagonal band and medial septum; some fibers terminate in the septal nuclei and others continue into the fornix to termimate in hippocampus. A large component continues around the corpus callosum into the cingulum to terminate in the cingulate and adjacent neocortex, the subiculum and hippocampus. The remaining fibers continue rostrally in the medial forebrain bundle to terminate in olfactory forebrain and frontal neocortex. Commissural projections arise at 4 locations. The first decussation occurs in the dorsal tegmentum just below the central gray rostral to the locus coeruleus. The crossing fibers enter the contralateral dorsal bundle. A second group of fibers leaves the ipsilateral dorsal pathway, crosses in the posterior commissure and enters the contralateral dorsal pathway at the level. The third commissural projection arises more rostrally and crosses in the dorsal supraoptic commissure to enter the contralateral medial forebrain bundle. The fourth commissural projection is through the anterior commissure. The termination of the contralateral projection appears similar to that of the ipsilateral projection.

Cerebellar Dentate Nucleus

Book

Jan 1977

Victoria Chan-Palay

Further mapping of central 5-hydroxytryptamine neurons: Studies with neurotoxic dihydroxytryptamine

Article

Jan 1974

Catecholamine-containing neurons in the brain stem of the cat and their role in waking

Article

Jan 1969

Barbara E Jones

The organization of the ascending catecholamine neuron systems in the rat brain as revealed by the glyxylic acid fluorescent method

Article

Jan 1974
Acta Physiol Scand Suppl

Acidic Glycolipoprotein Granules (Lysosomes) as Probable Binding Sites of Biogenic Amines

Article

Dec 1964
Progr Brain Res

Harold Koenig

This chapter focuses on a special class of cytoplasmic granules as a potential binding site for biogenic amines. These granules display a highly selective affinity for intravitally administered basic dyes and metalic ions, suggesting that they are negatively charged in vivo. The chapter proposes that lysosomes may be the binding sites of biogenic amines on the following grounds: (1) Lysosomes are polyanionic lipoprotein granules with demonstrated ability to segregate cationic substances, such as basic dyes and metallic cations. (2) Lysosomal enzymes and biogenic amines share the following features: (a) they are particulate; (b) they show structural latency, disruptive procedures releasing them into solution in active state; and (c) ionic bonds have been implicated in their binding to particles.

Morphological Correlates for Transmitter Synthesis, Transport, Release, Uptake and Catabolism: A Study of Serotonin Neurons in the Nucleus Paragigantocellularis Lateralis

Article

Jan 1978

Victoria Chan-Palay

The aim of this report is to gather available evidence from the literature and to provide fresh evidence from new experiments for the morphological intraneuronal correlates for transmitter synthesis, packaging, transport, uptake, and metabolism in the mammalian central nervous system. A large body of literature is available for the catecholamines (norepinephrine and dopamine) and gamma-amino butyric acid and they have been the subject of recent extensive reviews (Bloom, 1973; Roberts et al., 1976). The present report will explore the indoleamine-serotonin systems. The pathways for the biosynthesis and catabolism of serotonin (5HT) and related indoleamines are well known and aptly discussed by Cooper et al., 1974, pp. 175–199. The essential steps are summarized in Table 1. The essential amino acid tryptophan is converted enzymatically by tryptophan hydroxylase, the rate limiting enzyme, to 5-hydroxytryptophan which undergoes decarboxylation to form 5-hydroxytryptamine or serotonin (5HT). These initial steps represent the synthesis of 5HT; once formed, 5HT is stored, transported or released by the indole amine neuron in the course of its activity. 5HT is metabolized by monoamine oxidases in the presence of aldehyde dehydrogenase to form 5-hydroxyindole acetic acid. This report will review the findings of several pertinent studies using cytochemical or immunological methods in order to discover the intracellular organelles in indoleamine neurons which relate to the synthesis, transport, uptake, and catabolism of 5HT.

Immunofluorescent studies on the localization of tyrosine hydroxylase and dopamine-β-hydroxylase in the Mes-, Di-, And telencephalon of the rat using unperfused fresh frozen sections

Article

Jan 1979

Distributions of tyrosine hydroxylase (TH) and dopamine-β-hydroxylase (DBH) in the rat brain were examined using immunofluorescent staining on unperfused fresh frozen sections. TH-positive and DBH-negative dopaminergic neurons were observed in the anterior hypothalamic nucleus. In other hypothalamic nuclei such as ventromedialis, dorsomedialis, and posterior hypothalamic nuclei, a few dopaminergic neurons were also observed. Our immunofluorescent results with unperfused fresh frozen sections indicated that TH or DBH was mainly localized in the cell bodies of catecholaminergic neurons and that small amounts of the enzymes were distributed in the nerve terminals. Although it had been reported that the amine enzymes existed in nerve terminals as well as in the cell bodies by using paraformaldehyde perfused sections, the fluorescence observed in nerve terminals might be produced not only by the amine enzymes but also by amines themselves.

Fluorescenzhistochemische und neurohistologische Untersuchungen, am Nucleus raphe dorsalis der Ratte 1)

Article

Dec 1980
ACTA HISTOCHEM

The nucleus raphes dorsalis of the adult rat was investigated by means of fluorescence histochemical and neurohistological methods. After application of Falck-Hillarp-technique characteristic 5-HT-fluorescence could be seen in many, albeit not all of the cells of this nucleus. After using the rapid Golgi impregnation technique, 3 neuron types could be demonstrated: type-1-neurone (polygonal neurons with somatic spines); typ-2-Neurons (fusiform neurons), and type-3-neurons (pyriform neurons). The type-1- and type-2-neurons are, due to their different axon projection mode, considered to be efferent raphe neurons. The type-3-neurons are thought to be raphe interneurons. The type-1-neurons are inferred to represent the 5-HT producing neurons of the nucleus raphe dorsalis.

Association of serotonin-binding protein with projections of the midbrain raphe nuclei

Article

Jan 1975
BRAIN RES

The Rat Brain. A Stereotaxic Atlas

Article

Oct 1964

Dahlstrom A, Fuxe K. Evidence for existence of monoamine-containing neurons in central nervous system. I. Demonstration of monoamines in cell bodies of brain stem neurons. Acta Physiol Scand 232(Suppl): 1-55

Article

Nov 1963
Acta Physiol Scand Suppl

Regional Distribution of Circulating Blood During Submersion Asphyxia in the Duck

Article

Sep 1964
Acta Physiol Scand

Kjell Johansen

Regional blood flow distribution in ducks was estimated from fractional distribution of Rb86CI. A comparison was made between normally breathing ducks and submerged ducks. The data document that conspicuous changes in regional blood flow take place upon submersion. The skin, skeletal muscle, and organs of the gastrointestinal system showed a marked decrease in activity in the submerged condition down to an average of 3.9 % for the gizzard, 29.4 % for skin excised from the thorax and 11.0 % for gastrocnemius muscle. Surprising exceptions to this were displayed by tissues in the cranial part of the animals. Thus both skin and muscle from the head region showed an increase in activity during submersion. The same was true for the excised eye. The esophagus similarly showed an increased activity in the submerged animals. The myocardium from both atria and ventricles showed a striking increase amounting to 4.1 times higher activity in the left ventricular myocardium in the submerged animals. The change in vasomotor constrictor tone taking place upon submersion is thus highly selective and possibly segmentally oriented giving an increase in peripheral resistance to most organs posterior to the heart while the most cranial tissues seem to be subjected to a general decrease in vasoconstrictor tone with an increased blood flow. Activity in the kidneys was markedly consistent and 9.1 % of the value found in normally breathing animals. The adrenals showed an interesting increase in activity on the average as high as 4.9 times the value in normally breathing animals.

Electron microscopic observation on monoamine-containing brainstem neurons in normal and drug-treated rats

Article

Dec 1965
Anat Rec

Nicholas J. Lenn

Brain stem nuclei whose neurons contain monoamines have been surveyed by Dahlström and Fuxe ('64), using a fluorescence histochemical method. There was a diffuse fluorescence of the perikarya in these regions, with increases in catechol amine and 5-hydroxytryptamine fluorescence after administration of dihydroxyphenylanine and pargyline, respectively. The perikarya of the locus coeruleus, nucleus dorsalis raphes and the region lateral to the nucleus interpeduncularis where numerous monoamine-containing neurons are located, have been examined by electronmicroscopy and found not to be distinctive, except for small numbers of granular vesicles scattered in the cytoplasm. When three normal rats were compared with three given pargyline, no significant increase in the number of granular vesicles occurred in the nucleus dorsalis raphes. Thus the small number of granular vesicles does not seem to correlate with the diffuse fluorescence of these perikarya, nor does the small change in the number of granular vesicles seem to correlate with the profound increase in fluorescence after pargyline administration. It is therefore concluded that the monoamine contained in these perikarya is not located exclusively, if at all, in vesicles either of the granular or nongranular type.

Combined immunocytochemistry and autoradiography after in vivo injections of monoclonal antibody to Substance P and 3H-serotonin:

Article

Mar 1979

Victoria Chan-Palay

A novel approach is presented for combining immunocytochemistry and autoradiography after in vivo injections of monoclonal antibody to Substance P and 3H-serotonin. These experiments provide convincing evidence that the soma and processes of a single neuron can contain serotonin and immunoreactivity with another putative peptide transmitter Substance P in a single permanent preparation suitable for study with light and electron microscopy. The serotonin-Substance P cells have uptake systems for low molarity 3H-serotonin that are not affected by reserpine treatment. They are sensitive to serotonin uptake inhibitors and monoamine oxidase inhibitors and the cells are destroyed by 5,6-dihydroxytryptamine. They contain endogenous stores of serotonin detectable by the Flack-Hillarp technique and microspectrofluorimetry and immunofluorescence by antibody to serotonin. Their Substance P content is identified by specific binding with monoclonal antibody, and by animal antisera against Substance P. The raphe pallidus neurons are a mixed population — some cells have a majority of Substance P immunoreactivity with little detectable serotonin, others have considerable quantities of both Substance P immunoreactivity and serotonin, and still others have only serotonin but are innervated by Substance P immunoreactive processes. These subtle differences in neuronal content of the two compounds are made more obvious by avoiding the use of colchicine, a drug known to inhibit fast axoplasmic transport. Such differences may be an expression of the dynamic or cyclic functions of neurons with multiple putative transmitter substances. Fluctuating levels of one or both substances may depend upon parameters of rhythm, demands for one or the other mediator during specific types or phases of activity. The fundamental scheme of simultaneous injection of multiple labels, one radioactive, another a specific characterized monoclonal antibody for the detection of multiple, separate transmitter systems opens vast avenues for future investigation.

Neurotensin-containing cell bodies, fibers and nerve terminals in the brain stem of the rat: Immunohistochemical mapping

Article

Jun 1979
BRAIN RES

Neurotensin immunoreactive perikarya, fibers and nerve terminals, visualized by the indirect immunohistofluorescent method in colchicine-pretreated animals, are localized in many discrete regions of the rat brain stem. Cell body groups are found in the inner aspect of the substantia gelatinosa of the caudal trigeminal nuclear complex, the nucleus of the solitary tract, the parabrachial nuclei, the locus coeruleus, the dorsal raphénucleus, the periaqueductal gray matter, and the ventral tegmental area of Tsai. These areas of cell body density are accompanied by concentrations of fibers and terminals, while the occasional positive perikaryon noted in the dorsal cochlear nucleus is accompanied by only sparse fluorescent fiber/terminal patterns. Other brain stem regions, such as the floor of the fourth ventricle and aspects of the caudal ventrolateral reticular formation, possess substantial numbers of fibers and terminals that are not accompanied by cell bodies. Many aspects of this distribution coincide with the brain stem distribution of the enkephalin pentapeptides, though significant differences in localization are also evident. Interactions of neurotensin with other neurotransmitter candidates are also suggested by its presence in areas enriched in norepinephrine, dopamine, serotonin, and substance P. Certain neurotensin localizations suggest an association of the peptide with functional brain systems preferentially involving these regions. In particular periaqueductal gray and substantia gelatinosa neurotensin synapses are plausible sites for the analgesia elicited after intercisternal injection of low doses of neurotensin.

Immunohistochemical mapping of enkephalin containing cell bodies, fibers and nerve terminals in the brain stem of the rat

Article

May 1979
BRAIN RES

Enkephalin immunoreactive perikarya, fibers and nerve terminals, visualized by the indirect immunohistofluorescent method in colchicine-pretreated animals, are localized in many discrete regions of the rat brain stem. These specific immunohistofluorescent patterns are similar after staining with selective primary antisera directed against either methione-enkephalin or leucine-enkephalin. Cell bodies are found in the substantia gelatinosa and interpolaris zones of the trigeminal nuclear complex, the nucleus of the solitary tract, in the vicinity of the nucleus raphe´magnus, in the dorsal cochlear, medial vestibular, and paraolivary nuclei and, dorsal to this last region, in the parabrachial nuclei and the dorsal tegmental nucleus of Gudden, in the periaqueductal gray matter and interpeduncular nucleus and along the borders of the lateral lemniscus and medial geniculate. In some areas, such as the parabrachial region, nucleus of the solitary tract and substantia gelatinosa of the trigeminal nucleus, these perikarya are associated with densities of fibers and terminals. Other regions, such as the dorsal cochlear nucleus and the vicinity of the nucleus raphe´magnus, contain cell bodies associated with low densities of processes and terminals. In still other nuclei, such as the nucleus of the facial nerve and the locus coeruleus, fiber and terminal densities without associated cell bodies are evident. Many of these enkephalin localizations can be rationalized on the basis of known actions of opiate drugs and the brain stem distribution of opiate receptors.

Serotonin nerve terminals in adult rat neocortex* 1

Article

Jan 1976
BRAIN RES

Axonal processes which take up and retain exogenous tritiated serotonin ([3H]5-HT) have been demonstrated in the fronto-parietal cortex of adult rats, by means of high resolution radioautography. Prolonged topical superfusion with relatively high concentrations of [3H]5-HT allowed detection of a maximal number of axonal reactions in the upper 3 layers of cortex. Comparison of results obtained from normal rats and animals pretreated with 6-hydroxydopamine or bearing midbrain raphe lesions established the specificity of this labeling. All reactive axons seemed to arise from the serotonin nerve cell bodies in groups B7 and B8 of Dahlström and Fuxe15.

Distribution of serotonin-immunoreactivity in the central nervous system of the rat—Cell bodies and terminals

Article

Feb 1981
NEUROSCIENCE

Harry W M Steinbusch

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.

Vasoactive intestinal polypeptide (VIP) in mouse and rat brain: An immunocytochemical study

Article

Apr 1980
BRAIN RES

Immunoperoxidase technique and light microscopy were used to investigate the distribution of vasoactive intestinal polypeptide (VIP) in mouse and rat brain. Both 50 μm unmounted cryostat and 6 μm deparaffinized sections were studied in coronal or sagittal plane. At least 4 different major VIP systems were found: (1) an intracerebral cortical system; (2) one innervating the central amygdala and nucleus of the stria terminalis; (3) a pathway originating in the suprachiasmatic nucleus of the hypothalamus; and (4) another originating in the central grey of the midbrain. Specific cell body staining was seen in the limbic and neocortex, in the basal-caudal portion of the suprachiasmatic nucleus of the hypothalamus, and in the central grey of the midbrain. Heavy terminal field patterns were noted in the suprachiasmatic nucleus, central amygdaloid nucleus, bed nucleus of the stria terminalis and nucleus accumbens. Fiber density was moderate in the tuberculum olfactorium, anterior hypothalamus including the medial preoptic area, mediobasal hypothalamus (especially dorsomedial region), periventricular thalamus, lateral lemniscal system, parabrachial nucleus, nucleus solitarius, and area postrema. Fibers could be traced dorsally from the suprachiasmatic nucleus to the dorsomedial and paraventricular nuclei of the hypothalamus and the periventricular nucleus of the thalamus. Scattered cell bodies and fibers were found in a number of other forebrain and brain stem areas with only a rare fiber seen in median eminence.

Horseradish peroxidase tracing of the lateral habenular-midbrain raphe nuclei connexions in the rat

Article

Sep 1976
BRAIN RES BULL

Connections of the habenular complex to the nuclei of the midline in the midbrain (interpeduncularis, medianus raphe, and dorsalis raphe) have been studied classically by anterograde degeneration in the monkey, the cat, and marsupials. Passing fibers from the medial septal nucleus and lateral preoptic area, however, have also been demonstrated which can complicate interpretation of these results. In this paper the habenular projections were studied in the rat by the retrograde axonal transport of horseradish peroxidase (HRP). After HRP injections in the medianus raphe nucleus labelled neurons appeared in the lateral habenular nucleus and parafascicular nucleus. Labelled neurons were also found in the lateral habenular nucleus after injections in either the dorsalis raphe nucleus or the caudal central gray substance. The habenular projections were always bilateral. There were no labelled neurons in the medial habenular nucleus after HRP injections in the medianus raphe nucleus, dorsalis raphe nucleus, or central gray. These data stress the lateral habenular influences upon the raphe nuclei, especially on the dorsalis raphe neurons which have usually been thought of as functionally related to other brainstem structures. The present results suggest also that in the rat the lateral habenular nucleus might be the link between basal forbrain inputs and the limbic midbrain area. Thus, the raphe nuclei of the midbrain appear to be crucial regions for integrating two descending circuits: first, a limbic (through septum) circuit, and, second, a basal forebrain (through lateral habenular-preoptic area) circuit.

Estimation of Nuclear Population from Microtome Sections

Article

Feb 1946

M ABERCROMBIE

Occurrence of dopamine-containing neurons in the midbrain raphe nuclei of the rat

Article

Aug 1978
NEUROSCI LETT

Formaldehyde-induced fluorescence histochemistry, by the use of the improved filter system, revealed that dorsal and median raphe nuclei contain in varying numbers blue-green fluorescent neurons among a large number of yellow fluorescent serotonin-containing neurons. Pharmacological treatments indicated the presence of catecholamine in the blue-green fluorescent neurons. Moreover, microspectrofluorometry identified the catecholamine therein as dopamine, based on an excitation maximum shift characteristic of dopamine fluorescence caused by HCl vapor. Significance of these dopaminergic neurons in the raphe nuclei is suggested.

Monoamine-synthesizing enzymes in central dopaminergic, noradrenergic and serotonergic neurons. Immunocytochemical localization by light and electron microscopy

Article

Aug 1976

The monoamine-synthesizing enzymes tyrosine hydroxylase (TH), dopamine-beta-hydroxylase (DBH) and tryptophan hydroxylase (TrH) were immunocytochemical localized in dopaminergic, noradrenergic and serotonergic neurons of rat brain by light and electron microscopy. In dopaminergic and serotonergic neurons, the respective synthesizing enzymes. TH and TrH, were distributed throughout the cytoplasm of the neuronal perikarya, dendrites, axons and terminals. The most selective accumulation of reaction product for the specific enzyme was associated: (a) in perikarya with endoplasmic reticulum, Golgi apparatus and microtubules, (b) in processes with microtubules, and (c) in terminals with dense granules or clear vesicles. The labeled terminals were characterized by their content of labeled organelles and the absence of synaptic junctions. In noradrenergic neurons, both TH and DBH were localized in the perikarya, similar to TH in dopamine neurons. TH and DBH differed in their localization within proximal axons and dendrites in that TH was associated with microtubules but DBH was not. These results provide ultrastructural evidence to suggest that monoamines may be: (a) synthesized by enzymes which are associated with different organelles depending on the portion of the neuron and the type of enzyme; (b) synthesized in both axons and dendrites and (c) released from terminals without postsynaptic membrane specializations.

Immunohistochemical studies on the localization and distribution of monoamine neuron systems in the rat brain. I. Tyrosine hydroxylase in the mes and diencephalon

Article

Jan 1977
Med Biol

The localization and distribution of tyrosine hydroxylase (TH), the first enzyme in the catecholamine synthesis, in the mes- and diencephalon has been studied with the indirect immunofluorescence technique of Coons and collaborators. Principally, TH was present in neuron systems with a distribution similar to known dopamine, noradrenaline and adrenaline systems. The present data, taken together with published and some unpublished results, indicate that all parts of most central dopamine neurons, i.e. cell body, dendrites, axon and nerve terminals, appear strongly fluorescent. The adrenaline neurons also appeared strongly fluorescent, except for their axons, which only exhibited a weak fluorescence. Only cell bodies of noradrenaline neurons were strongly fluorescent, whereas the nerve terminals and axons showed a weak or moderate fluorescence intensity. The fine noradrenaline nerve terminals in some areas, such as the thalamus, were invisible or, under favourable conditions, weakly fluorescent. Therefore, in the present study we are mainly dealing with the dopamine neurons of the upper brain stem. Our results demonstrate a widespread occurrence of TH-positive neuron systems in the mes- and diencephalon. The different mesencephalic dopamine systems and their ascending projections were visualized. Numerous TH-positive cell bodies were present along the ventricle system extending from the aqueductus cerebri to the most cranial periventricular parts of the third ventricle. The caudal part of these neurons, consisting of very small cell bodies, belong to the dorsal periventricular system described by Lindvall and Björklund. Several TH-positive cell bodies were also observed in the inferior collicle of young animals. In the superficial layers of the inferior collicles TH positive nerve terminals were seen. At the hypothalamic level the A11 to A14 cell groups as well as some additonal cell bodies and extensive nerve terminal plexuses appeared strongly fluorescent. The differences in the intensity of the TH-related immunofluorescence between various brain regions and between various neuron systems may well reflect differences in enzyme levels between the various catacholamine systems rather than be due to the existence of different types of TH.

A Guide to the Synaptic Analysis of the Neuropil

Article

Feb 1976

A morphological analysis of the organization of the gray matter in the central nervous system depends on the discovery of consistent repetitive patterns. Without these, the gray matter remains a chaotic jungle. An hypothesis derived from the study of a few simple regions has been developed to serve as a guide in finding these patterns. It states that all nerve fibers and terminals arising from a particular group of nerve cells, or, more precisely, a particular nerve cell type, display similar axoplasmic configurations despite variations in size and shape of the terminations. This hypothesis is reminiscent of the so-called Dale's principle that a nerve cell makes use of the same transmitter at all of its branches or terminations. These apparent rules of uniformity or congruity merely reflect the functional integrity of the nerve cell and the role of its parts in the nervous system. But as an hypothesis, it needs to be tested, and it needs to be tested anew in each region, since exceptions to the assumed rule can be expected. It is therefore proposed as the first working hypothesis in each new region. If it should prove to be true in general, it will facilitate and rationalize the analysis of the gray matter, as it has already done in the cerebellar cortex and the deep cerebellar nuclei. If it should prove to be false in a few regions, the analysis will become more difficult, and additional modes of marking nerve endings will have to be used. Experimental methods for identifying nerve terminals can be translated from the light microscopic to the electron microscopic level, but there are significant drawbacks at both levels: lack of precision, destruction of fibers of passage, and rapid evolution of the degenerative process may greatly restrict their usefulness. Labeling with tritiated amino acids or transmitters, or with horseradish peroxidase, provide new methods for tracing interneuronal connections at the electron microscopic level. These have the advantages of high specificity, nondestructiveness and a physiological mode of selective marking. However, they do not offer a solution to the problem of short-range connections. For these, careful reconstructions of serial sections may prove necessary, as Sjöstrand (1974) has demonstrated in a remarkable paper on the retina. The aim of all these methods is to discover patterns of synaptic connectivity in order to map the cellular organization of the nervous system. In the foregoing, nothing was said about synapses other than those articulating axons with somata or dendrites and their appendages. Clearly the same principles of recognition apply to axo-axonal and dendro-dendritic synapses. Although the synapses that have been considered here are chemical synapses, the same questions regarding the identity of the partners in electrotonic junctions must be asked as well.

Electron microscope autoradiographic evidence for specific transneuronal transport in the mouse accessory olfactory bulb

Article

Sep 1978
BRAIN RES

The distribution of radioactive material was examined autoradiographically 8 h after application of [3H] proline to the vomeronasal organ in mice. Labelled material was transported along the axons of the vomeronasal nerves to their terminals in the glomerular layer of the accessory olfactory bulb (AOB). A lesser but consistent amount of radioactivity was found in the external plexiform layer (EPL) of the AOB. Electron microscopic autoradiography was used to determine which of the components of the EPL contained this labelled material. The method of proportional grain counts showed that the highest concentration of silver grains lay over the mitral cell dendrites, which are the elements immediately postsynaptic to the vomeronasal nerve axons. However, a fairly high proportion of grains also lay over the peripheral processes of granule cells. By application of a method of 'crossfire analysis' (which is explained in detail) it was possible to show that the observed grain distribution is best explained by the assumption that the radioactive material is confined to mitral cells, and the labelling over granule cell processes is due to crossfire from these sources. Im one animal at 5 days after [3H]proline administration label was found to have extended from mitral cells to granule cells, suggesting that the transsynaptically transported radioactive material, which was confined to the mitral cells at 8 h, may have become further redistributed at longer survivals. In a control experiment, [3H]proline was applied directly to the surface of the AOB. This gave rise to a completely different distribution of radioactivity in the EPL: radioactive material was present in all tissue components.

Radioautographic characterization of serotonin-accumulating nerve cell group in adult rat hypothalamus

Article

Feb 1979
BRAIN RES

Intensely labeled nerve cell bodies were identified by radioautography within the pars ventralis of nucleus dorsomedialis hypothalami (hdv), following intraventricular perfusion with 10(-5) or 10(-4) M tritiated serotonin [3H]5-HT in adult rats pretreated with a monoamine oxidase inhibitor. This selective reaction, which involved approximately 1000 neurons on each side of the third ventricle, was unaltered by concomitant administration of 10(-3) M non-radioactive norepinephrine, and was absent after intraventricular injection of 10(-5) or 10(-4) M tritiated norepinephrine. The 3H-labeled 5-HT nerve cell bodies were loosely grouped within the inner and caudal half of the hdv, and appeared morphologically similar to the unreactive neurons among which they were interspersed. Within the same region, numerous labeled axonal varicosities were also detected, which were never found in synaptic contact with the reactive cells. If the 3H-labeled 5-HT neurons contain endogenous 5-HT, they might constitute an intrinsic source of 5-HT innervation in the adult rat hypothalamus.

The Raphe nuclei of the rabbit brain stem

Article

Sep 1979

The raphe nuclei of the rabbit brain stem were found in the midline and adjacent reticular formation of the medulla, pons, and mesencephalon. Nuclei raphe obscurus, pallidus, and magnus were located in the medulla. Nucleus raphe pontis and the caudal portion of nuclei raphe dorsalis and centralis superior were present in the pons. The rostral portion of nuclei raphe dorsalis and centralis superior, and nuclei linearis caudalis and intermedius were present in the msencephalon. Wings of neurons extended from the midline clusters of raphe neurons into the adjacent reticular formation. These wings of neurons contained serotonergic perikarya which were cytoarchitecturally indistinguishable from the midline neurons. A detailed localization of these nuclei is presented in atlas form. These raphe nuclei contained heterogeneous populations of neurons which varied in the size, shape and density of the cell bodies. In addition, the dendritic branching, specific orientation of dendrites, and appearance of spines were distinct for each of the raphe nuclei. Individual raphe nuclei often contained several subpopulations of neurons characterized by unique spatial configuration and orientation. The main morphological similarities of the raphe nuclei are location in or adjacent to the midline, the presence of serotonergic cell bodies in all raphe nuclei except the linear nuclei, and heterogeneous cell populations.

Evidence against a neurotoxic action of halogenated amphetamines on serotoninergic B9 cells

Article

Jun 1978
BRAIN RES

Serotonin nerve terminals in the locus coeruleus of adult rat: A radioautographic study

Article

May 1978
BRAIN RES

Serotonin (5-HT) nerve terminals in the locus coeruleus (LC) of adult rat were visualized by high-resolution radioautography, in order to examine their distribution, fine structural features and intimate relationships with norepinephrine neurons. In animals pretreated with a monoamine oxidase inhibitor, prolonged intraventricular perfusion of 10(-4) M [3H]5-HT resulted in a specific identification of most if not all 5-HT axonal varicosities in LC. These terminals were equally distributed between the dorsal and ventral divisions of the nucleus. Their density was approximated at 10(7) per cu.mm within the middle third of the LC. In electron microscope radioautographs, the labeled 5-HT varicosities averaged 0.9 micron in diameter. They all exhibited a distinctive storage organelle, in the form of microvesicles and microcanaliculi (15-25 nm in diameter) partly filled with electron-dense material and usually aggregated in association with several large dense-core vesicles. While this finding of intrinsic morphological characteristics appeared compatible with a special cellular origin or regional differentiation, it was also suggestive of particular functional properties and/or mode of action. In a sample comprised of some 500 sectional profiles from labeled 5-HT varicosities in LC, a small proportion only (less than 10%) exhibited morphologically defined synaptic junctions. These rare contacts were invariably made with dendritic processes and never observed on the noradrenergic perikarya. It is therefore concluded that, in the LC, non-synaptic as well as synaptic mechanisms might be involved in the modulation and transneuronal regulation of norepinephrine neurons by 5-HT afferents.

Serotonin and Substance P Coexist in Neurons of the Rat's Central Nervous System

Article

Apr 1978

5-Hydroxytryptamine (serotonin)-containing neurons in the rat's medullary raphe and interfascicularis hypoglossi cell groups were identified by means of autoradiography following prolonged intraventricular administration of 5-hydroxy[(3)H]tryptamine, fluorescence histochemistry for the demonstration of endogenous 5-hydroxytryptamine, and microspectrofluorimetric analysis of excitation and emission spectra. Immunocytochemical methods (the unlabeled primary antibody-peroxidase antiperoxidase and indirect immunofluorescence methods) were applied with antisera to substance P in order to localize immunoreactivity in these medullary neurons. It was demonstrated that the raphe nuclei and the interfascicularis hypoglossi nucleus are heterogeneous cell groups that contain: (i) Neurons that display both an uptake-storage capacity for 5-hydroxy[(3)H]tryptamine and a formaldehyde-induced fluorescence with spectral characteristics identical to those of the 5-hydroxytryptamine fluorophor. These cells exhibit high to low fluorescence intensities without detectable substance P-like immunoreactivity. (ii) Neurons with various 5-hydroxytryptamine fluorescence intensities and intense to low degrees of substance P-like immunoreactivity. (iii) Neurons with various degrees of substance P-like immunoreactivity without detectable 5-hydroxytryptamine fluorescence or 5-hydroxy[(3)H]tryptamine uptake and storage capacity. These results indicate that some neurons contain high or low levels of only 5-hydroxytryptamine or substance P, whereas other neurons contain both 5-hydroxytryptamine and substance P in various proportions. The present findings demonstrate the presence of two putative transmitters, a biogenic amine and a polypeptide, within the same neuron in the mammalian central nervous system.

Immunohistochemical evidence of Substance P-like immunoreactivity in some 5-hydroxytryptamine-containing neurons of the central nervous system

Article

Feb 1978
NEUROSCIENCE

With the indirect immunofluorescence technique of Coons and collaborators a possible coexistence of 5-hydroxytryptamine (5-HT) and substance P in neurons of the lower medulla oblongata was explored. Antisera to 5-HT and to dopadecarboxylase (aromaticl-aminoacid decarboxylase), an enzyme probably present in immunologically indistinguishable forms both in catecholamine and 5-HT neurons, were used as markers for 5-HT neurons and an antiserum raised to synthetic substance P conjugated with bovine serum albumin for substance P-containing neurons. Five or 10 μm thick, consecutive sections were stained with the three antisera. Numerous cell somata in nucleus raphe magnus, nucleus raphe obscurus, nucleus raphe pallidus, pars α of the nucleus reticularis gigantocellularis and nucleus interfascicularis hypoglossi contained both substance P-like immunoreactivity and 5-HT (and dopadecarboxylase) immunoreactive material. After intraventricular or intracisternal injections of 5,6- or 5,7-dihydroxytryptamine, two neurotoxins assumed to cause degeneration mainly of 5-HT neurons, enlarged substance P and 5-HT (and dopadecarboxylase) positive fibres were seen in, around and lateral to the olivary complex. Furthermore, in these rats both substance P and 5-HT positive nerve terminals in the ventral horns of the spinal cord disappeared.

A brainstem atlas of catecholaminergic neurons and serotonergic perikarya in a Pygmy Primate (Cebuella pygmaea)

Article

Feb 1978

The present paper provides a brainstem atlas showing the distribution of catecholaminergic cells and processes, as well as serotonergic perikarya, in the pygmy marmoset. The findings revealed by the Falck and Hillarp histofluorescence method conform in essential details to what has been described in rodents. These and other comparative data indicate that a prototypical pattern of organization of aminergic systems has been retained in the evolution of primates.

The efferent connections of the nucleus raphe centralis superior in the rat as revealed by autoradiography. Brain Res 166: 1-8

Article

May 1979
BRAIN RES

By chronically implanting a glass micropipette filled with tritiated leucine in the raphe centralis superior of the rat, the projection of this nucleus was traced by radioautography. The majority of the ascending projections were located within the ventral tegmental area and, further rostrally, the median forebrain bundle. Along the course of this bundle numerous fibers branched successively into the mammillary peduncle, the fasciculus retroflexus, the stria medullaris, the fornix and the cingulum. The most significant projections included the ones to the interpeduncular nucleus, the mammillary bodies, the habenular nuclei and the hippocampus. No projections were detected in the striatum, the cortex piriformis or the amygdala. Descending projections diffused to the pontine reticular formation and central gray through the medial and the dorsal longitudinal bundles. In addition widespread projections were also seen in nuclei located near the raphe centralis superior: raphe nuclei, dorsal and ventral tegmental nuclei.

GABA-accumulating neurons in the nucleus raphe dorsalis and periaqueductal gray in the rat: A biochemical and radioautographic study

Article

Aug 1979
BRAIN RES

The possibility of a GABAergic innervation of the nucleus raphe dorsalis (NRD) has been investigated by using the following approaches: (i) the identification of the principal neuronal groups afferent to the NDR by using horseradish peroxidase retrograde transport, (ii) the determination of glutamate decarboxylase activity (GAD) in the NRD after lesioning these groups or their putative pathways, and (iii) the radioautographic identification of terminals axons and nerve cells accumulating intraventricularly injected [3H]GABA. The hypothesis of a local GABAergic network is supported by the failure to obtain important changes in GAD after lesions of NRD afferents and the presence in this nucleus of terminals, fibers and nerve cell bodies accumulating [3H]GABA. It appears that these GABA-accumulating neurons could represent a portion of aperiventricular GABAergic system in the periaqueductal gray and the pontine ventricular gray.

High resolution radioautographic identification of [3H]GABA labeled neurons in the rat nucleus raphe dorsalis

Article

Dec 1979
NEUROSCI LETT

Following intraventricular administration of [3H]GABA into the lateral ventricle of the rat radioautography demonstrates the presence of labeled cell bodies and fibers within the nucleus raphe dorsalis (NRD). Pharmacological controls suggest that labeled perikarya whose diameter does not exceed 10 micrometer and which are not probably more than ten in the entire NRD could be GABAergic. The effects of 5,7-dihydroxytryptamine (5,7-DHT) pretreatment support the hypothesis that labeled fibers could belong at least to two populations: ones being probably serotonergic and the others being possibly GABAergic. These results are in good agreement with the existence of an intrinsic GABAergic system within the rat NRD.

Atlas of the distribution of monoamine nerve cell bodies in the brain stem of the cat

Article

Jun 1978

The distribution and morphological characteristics of monoamine (MA)-containing neuronal somata in the brain stem of kittens and of adult cats were studied by means of the Falck-Hillarp histofluorescence method. This investigation has shown, among other things, that in the midbrain of the cat the catecholamine (CA) perikarya are chiefly confined to the pars compacta of the substantia nigra, the ventromedial tegmental area, the nucleus linearis rostralis and the nucleus parabrachialis pigmentosus. Numerous CA neurons are also present in the dorsolateral part of the pontine tegmentum but also within the nucleus subcoeruleus, in nuclei lemnisci lateralis dorsalis and in nuclei parabrachialis lateralis and medialis. In the medulla, a few CA neuronal somata are lying near the hypoglossal nucleus whereas a larger number of CA cell bodies occur at the level of nucleus reticularis lateralis and in nucleus paragigantocellularis lateralis. On the other hand, most of the serotonin (5-HT) perikarya are confined to the raphe nuclei of the brain stem: nuclei raphe dorsalis, centralis superior, raphe pontis, raphe magnus, raphe pallidus and raphe obscurus. Some 5-HT neuronal somata are also found lateral to the pyramidal tract and to the inferior olivary complex. The various similarities and differences in respect to the pattern of the topographical distribution of MA neurons in the brain stem of the cat as compared to that of other mammals are discussed.

Developmental organization of raphe serotonin neuron groups in the rat

Article

Oct 1978

The pre- and early postnatal development of serotonin neurons in the rat brainstem was studied using the fluorescence histochemical method. The technique utilized does not require drug pretreatment to visualize an intense serotonin fluorophore localized in neuronal perikarya, dendrites, and axons. All the serotonin neuron groups develop as bilateral nuclei which extend from the midbrain through the medulla. Six of the nine groups undergo a midline fusion from embryonic day 18 (E 18) through postnatal day 6 (P 6) in a rostrocaudal gradient. Cells of the nucleus raphe dorsalis fuse first (by P 1), whereas the serotonin neurons located in nucleus raphe pallidus do not fuse until P 6. This gradient is comparable to the one described for the first observable fluorescence in the serotonin neurons groups. After final cell division, the serotonin neurons undergo a primary migration from the ventricular zone along the midline, where they are situated during embryogenesis, and a secondary migration extending into postnatal life which concludes with fusion in the midline. The bilateral origins of the serotonin cell groups are maintained in the adult. This is expressed by the apparent ipsilateral projections of some of the raphe neurons determined recently in our laboratory utilizing autoradiographic and horseradish peroxidase techniques.

Do raphe nuclei of the reticular formation have a neurosecretory or vascular sensor function?

Article

Jun 1975

Golgi analysis of the various subnuclei of the raphe complex of the brain stem's reticular core reveals an apparently special neurovascular relationship for two of them. Cells of the n. raphe pontis and n. linealis rostralis which have been recognized for some time as forming paired systems are now seen to lie closely applied along the surface of raphe vessels which ascend from the basilar artery on each side of midline. Both cells and dendrites of these two neuronal ensembles ascend along the vessels, the dendrites frequently appearing to bear specialized expansions en passage or terminally, with which to relate to the vascular walls. Histochemical and electron microscopic techniques have been used in efforts to determine the actual nature of this relationship. It has been suggested that the role of the two raphe systems might include: (i) neurosecretion, releasing some active principle into the underlying vascular system; (ii) chemosensor function, responding to specific circulating substances, (iii) mechanoreceptor function, responding to changes in tone and/or diameter of the vessel wall. Introductory studies appear to make the first unlikely, although all possibilities need further evaluation. If these nuclei should be shown to fulfill the second or third roles, it would make it less likely that raphe neurons themselves represent the original triggr for the onset of slow wave sleep, but rather, act in response to a distant pacemaker.

Serotonergic afferents to the dorsal raphe nucleus: Evidence from HRP and synaptosomal uptake studies

Article

Feb 1977
BRAIN RES

Afferent connections of the serotonin (5-HT)-containing dorsal raphe nucleus were investigated in the rat utilizing the horseradish peroxidase (HRP) retrograde cell labeling technique. Small quantities (0.1-0.5 mul) of HRP solutions were infused into the dorsal raphe, and the brains were examined 19-72 h later for retrograde transport of the enzyme. Intrinsic connections within the dorsal raphe nucleus were revealed by this mapping technique, as was an input to the dorsal raphe from another serotonergic cell group, the median raphe nucleus. Little evidence was found for projections from other, more remote, brain sites. A serotonergic innervation of the dorsal raphe was also demonstrated by the presence of high affinity uptake of [3H]5-HT (Km=0.17 muM) into synaptosomal suspensions of the dorsal raphe nucleus. Synaptosomal uptake of [3H]5-HT was blocked by selective destruction of serotonergic axon terminals induced by the intraventricular injection of 200 mug of 5,7-dihydroxytryptamine following desipramine HCl pretreatment, but not by destruction of catecholaminergic axon terminals induced by intraventricularly injected 6-hydroxydopamine (2 X 250 mug). The uptake of [3H]-5-HT by synaptosomes of the dorsal raphe was comparable to that of striatal and hypothalamic synaptosomes, and markedly greater than that of synaptosomes from the cerebellum or nearby dorsal central gray or midbrain reticular formation, indicating the presence of a relatively dense serotonergic innervation. These data together indicate that neurons in the dorsal raphe nucleus receive a prominent serotonergic input that is derived, at least in part, from other neurons within the dorsal nucleus and from a neighboring raphe nucleus.

Habenular and other midbrain raphe afferents demonstrated by a modified retrograde tracing technique

Article

Mar 1977
BRAIN RES

Afferents to th midbrain dorsal and median raphe nuclei in the rat were studied by means of the horseradish peroxidase (HRP) retrograde transport method. The HRP was given by means of a modified iontophoretic delivery technique. This technique permitted an efficient and localized deposition of a high concentration of HRP into the raphe nuclei. Afferents to the raphe as determined by this method could be categorized into 2 classes; those exclusively to the raphe and those also positive for adjacent reticular formation. The most striking afferent area to the raphe, both in terms of selectivity and density, was the lateral habenula. This result is in accord with previous studies using degeneration methods which indicate an habenular projection to the raphe area. There were afferents exclusively positive for the dorsal raphe nucleus emanating from the nucleus of the solitary tract. Most other raphe afferent areas were also positive for the reticular formation (e.g;, prefrontal cortex, medial forebrain bundle, preoptic nuclei, and reticular formation). The existence of a major afferent system from the lateral habenula to the midbrain raphe is consistent with the concept of a "dorsal pathway" which might be responsible for relaying information from forebrain limbic structures to the "midbrain limbic areas".

A simplified method of 'hypothetical grain' analysis of electron microscope autoradiographs

Article

Apr 1977

An improved method for the analysis of high resolution electron microscope autoradiographs is described which is simple and quick to perform. The method uses a transparent screen superimposed over the autoradiograph prints to provide information on the distribution of grains over neighboring structures, produced by radioactive disintegrations occurring in different regions of the section. This "cross-scatter" information is used to estimate the radioactivity in different structures without the need to make any assumptions about the size, shape and arrangement of the structures within the autoradiographs being analyzed. The method allows activity in membranes and other linear structures to be determined and also the accuracy attributable to the activity values obtained.

Intracisternal neurotoxins and monoamine neurons innervating the spinal cord: Acute and chronic effects on cell and axon counts and nerve terminal densities

Article

Jul 1977
Histochemistry

The total number of catecholamine (CA) and 5-hydroxy-trydroxy-tryptamine (5-HT) containing nerve cell bodies in groups A1-7 and B1-3 were counted using Falck-Hillarp fluorescence histochemistry and found to be about 5000 and about 6300 respectively. The total number of CA axons in the white matter of the spinal cord was found to be about 4300 at the cervical level and about 3300 at the lumbar level as revealed in cryostat sections. Together with previous studies our quantitative results suggest that individual cell bodies in locus coeruleus send axon collaterals to the spinal cord, cortex cerebri, cortex cerebelli and probably also other areas of the brain. The decrease in number of CA axons in spinal cord occurred approximately at the level where the CA nerve terminal innervation of the sympathetic lateral column ends. Single CA axons were observed to give off multiple branches at right angles, suggesting that single CA axons innervate several different levels of the grey matter. A transient loss of visible CA cell bodies in groups A1-7 was observed after intracisternal injection of 25–50 μg 6-hydroxydopamine (6-OH-DA). A permanent loss of visible 5-HT cell bodies in groups B1-3 was observed after 25 μg 5,6-dihydroxy-tryptamine (5,6-HT) given intracisternally, a transient decrease was obtained when the same dose of 5,6-HT was given intraventricularly. CA nerve terminals regenerated in a cranio-caudal direction along the motoneuron columns after lesioning with 6-OH-DA given intracisternally. The reappearing CA nerve terminals seemed to outline the longitudinally oriented dendritic bundles of the motoneurons and was exclusively found in these areas of the ventral horn. Thus, no substantial CA nerve terminal reinnervation of zona intermedia or the dorsal horn was observed. About 10% of the CA axons running in white matter were left after intracisternal 6-OH-DA and found at all levels of the cord. The number of remaining CA axons did not increase with time. After 10–50 μg 5,6-HT given intracisternally a return of 5-HT nerve terminals was only observed in cervical segments. There was a 60–90% decrease in number of axons that accumulated 5-HT after an acute transverse lesion and no recovery of the number of such accumulations was observed during the following six months. It was concluded that regeneration of CA nerve terminals in the spinal cord can occur after lesioning with 6-OH-DA given intracisternally. Regeneration of 5-HT nerve terminals following intracisternal 5,6-HT is very restricted probably because of the permanent loss of 5-HT cell bodies obtained by the intracisternal route of administration as opposed to the intraventricular route. It was suggested that the reappearing CA nerve terminals were derived from locus coeruleus and that they were specifically guided by the motoneuron columns.

Indoleamine neurons and their processes in the normal rat brain and in chronic diet-induced thiamine deficiency demonstrated by uptake of3H-Serotonin

Article

Dec 1977

Victoria Chan-Palay

Indoleamine structures in the brains of normal, pair-fed control, and thiamine deficient rats are selectively localized by autoradiography following monoamine oxidase inhibition and simultaneous intraventricular infusion of 3H-5HT (10−5 m) and cold d–1 norepinephrine (10−4 m). The locations of indoleamine neurons and their plexuses are comparable in the normal and control animals and are enumerated and mapped in detail. In addition to numerous labeled neurons in the raphe and reticular formation of pons and medulla, there are indoleamine neurons in several hypothalamic nuclei, in the mammillary, habenular, and interpeduncular nuclei. Labeled axonal plexuses occur in almost all areas of the brain but certain regions in spinal cord, medulla, midbrain, diencephalon, basal ganglia, and hippocampus are particularly rich. Thiamine deficiency causes lack of label in almost all indoleamine neurons and their processes in the midbrain and medulla; only a few dystrophic cells with hypertrophied axons are seen. In the diencephalon more indoleamine neurons and their axonal plexuses are visualized in these uptake studies. The periventricular regions of spinal cord, medulla, midbrain and diencephalon, the mammillary nuclei, habenular nuclei, and cerebellum – areas richly innervated by indoleamine axons in the normal brain – are the most severely affected. The ventricular and leptomeningeal indoleamine structures are preserved, however, and individual axons may hypertrophy. It remains to be resolved whether these changes reflect disturbances in the ability of certain indoleamine neurons to take up and retain 3H-5HT or true neuronal degeneration.

Monoamine-containing cell bodies in the Squirrel monkey brain

Article

Feb 1974

Fifteen catecholamine-containing cell groups and eight indoleamine-containing cell groups are present in the brain of the squirrel monkey. Most of the catecholamine-containing cell groups (12) are similar to catecholamine-containing cell groups previously described in the rat. However, three catecholamine-containing cell groups not previously noted are found in the squirrel monkey brain. The indoleamine-containing cell groups are found within, or adjacent to, the raphe nuclei. Differences between the localization of indoleamine-containing cell bodies in the brain of the rat and the squirrel monkey are minor.

Fluorescence histochemistry of monoamine containing cell bodies in the brain stem of the squirrel monkey (Saimiri sciureus). III. Serotonin containing groups

Article

Feb 1974

The distribution and cytological characteristics of the serotonin-containing cell bodies in the brain stem of the squirrel monkey (Saimiri sciureus) are described using the fluorescence histochemical technique of Falck and Hillarp for the demonstration of monoamines. Eight groups of serotonin-containing neurons were found, located predominantly in the midline raphe region of the brain stem within defined nuclei, such as the nucleus raphe obscurus, nucleus raphe pallidus, nucleus raphe magnus, nucleus raphe pontis, nucleus raphe dorsalis and nucleus centralis superior. Portions of some of the groups also extend laterally in the brain stem, particularly at the levels of the upper pons and midbrain. The appearance and distribution of the serotonin-containing groups of cell bodies in the squirrel monkey were found to be somewhat similar to that described for the rat, but differed in at least one major respect from that reported for the cat.

Histochemical Fluorescence of Raphe Neurons: Selective Enhancement by Tryptophan

Article

Jul 1971

The histochemical fluorescence of those neurons in brainstem raphe nuclei which are presumed to contain serotonin is selectively and stereospecifically enhanced by L-tryptophan at doses that also produce an elevation in the concentration of serotonin. However, contrary to our assumptions, the increase in raphe fluorescence is not prevented by p-chlorophenylalanine, an inhibitor of serotonin synthesis. These results suggest that under some conditions derivatives of tryptophan other than, or in addition to, serotonin may be of significance in raphe neurons.

Serotonin-containing neuronal perikarya and terminals: Differential effects of P-chlorophenylalanine

Article

Jun 1973
BRAIN RES

Previous studies have shown that p-cholorophenylalanine (PCPA), an irreversible inhibitor of serotonin (5-HT) synthesis at the tryptophan hydroxylase step, fails to prevent the l-tryptophan-induced increase in the fluorescence of 5-HT-containing neuronal perikarya of the brain stem raphe nuclei, although there is a marked depletion in whole brain 5-HT concentration. In an effort to explain this apparent discrepancy, the effect of PCPA upon the histofluorescence of raphe cell terminals in the midbrain and forebrain of rats was examined. Demonstration of the fluorescence of raphe cell terminals was aided in part by pretreatment with l-tryptophan in the presence of a monoamine oxidase inhibitor. PCPA was found to produce a marked decrease in the fluorescence of raphe terminals but, in agreement with previous results, not in the perikarya. Parallel biochemical studies showed that 5-HT was depleted by PCPA (with or without l-tryptophan loading) to a much greater extent in the forebrain (a region rich in raphe terminals), than in a block of tissue containing the raphe cell perikarya. These results indicate that the synthesis of 5-HT is more susceptible to inhibition by PCPA in the terminals than in the perikarya of raphe neurons, perhaps due to a continued synthesis of new tryptophan hydroxylase enzyme in the latter site.

The serotonin neurons in nucleus raphe dorsalis of adult rat: A light and electron microscope radioautographic study

Abstract

No full-text available

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