Serotonergic Neurons In Raphe Nuclei Research Articles

Neuropeptide VF neurons promote sleep via the serotonergic raphe.

Although several sleep-regulating neuronal populations have been identified, little is known about how they interact with each other to control sleep/wake states. We previously identified neuropeptide VF (NPVF) and the hypothalamic neurons that produce it as a sleep-promoting system (Lee et al., 2017). Here we show using zebrafish that npvf-expressing neurons control sleep via the serotonergic raphe nuclei (RN), a hindbrain structure that is critical for sleep in both diurnal zebrafish and nocturnal mice. Using genetic labeling and calcium imaging, we show that npvf-expressing neurons innervate and can activate serotonergic RN neurons. We also demonstrate that chemogenetic or optogenetic stimulation of npvf-expressing neurons induces sleep in a manner that requires NPVF and serotonin in the RN. Finally, we provide genetic evidence that NPVF acts upstream of serotonin in the RN to maintain normal sleep levels. These findings reveal a novel hypothalamic-hindbrain neuronal circuit for sleep/wake control.

Serotonergic projections to the ventral respiratory column from raphe nuclei in rats

The ventral respiratory column (VRC) generates rhythmical respiration and is divided into four compartments: the Bötzinger complex (BC), pre-Bötzinger complex (PBC), rostral ventral respiratory group (rVRG), and caudal ventral respiratory group (cVRG). Serotonergic nerve fibers are densely distributed in the rostral to caudal VRC and serotonin would be one of the important modulators for the respiratory control in the VRC. In the present study, to elucidate detailed distribution of serotonergic neurons in raphe nuclei projecting to the various rostrocaudal levels of VRC, we performed combination of retrograde tracing technique by cholera toxin B subunit (CTB) with immunohistochemistry for tryptophan hydroxylase 2 (TPH2). The double-immunoreactive neurons with CTB and TPH2 were distributed in the both rostral and caudal raphe nuclei, i.e. dorsal raphe nucleus, raphe magnus nucleus, gigantocellular reticular nucleus alpha and ventral parts, lateral paragigantocellular nucleus, parapyramidal area, raphe obscurus nucleus, and raphe pallidus nucleus. The distributions of double-immunoreactive neurons were similar among injection groups of BC, PBC, anterior rVRG, and posterior rVRG/cVRG. In conclusion, serotonergic neurons in both rostral and caudal raphe nuclei projected throughout the VRC and these serotonergic projections may contribute to respiratory responses to various environmental and vital changes.

Evaluation of food intake and Fos expression in serotonergic neurons of raphe nuclei after intracerebroventricular injection of adrenaline in free-feeding rats

Previous studies indicate that the modification of adrenergic neurotransmission in median raphe nucleus (MRN) enhances or removes an inhibitory influence on food intake, possibly serotonergic, due to a presence of serotonin-producing neurons in that nucleus. Therefore, the aim of this study is evaluated whether the activity of neurons in the MRN and dorsal raphe nucleus (DRN) are affected by intracerebroventricular injection of adrenaline (AD) in free-feeding rats. Male Wistar rats with guide cannulae chronically implanted in the lateral ventricle were injected with AD followed by evaluation of ingestive behavioral parameters. Behavior was monitored and the amount of food ingested was assessed. The highest dose (20 nmol) of AD was the most effective dose in increasing food intake. Subsequently, AD 20 nmol was injected to study neuronal activity indicated by the presence of Fos protein and its co-localization with serotonergic neurons in the MRN and DRN of naive rats with or without access to food during the recording of behavior. The administration of AD 20 nmol increased Fos expression and double labeling with serotonergic neurons in the DRN in rats with access to food, but not in animals without access. No statistically significant changes in Fos expression were observed in the MRN in any of the experimental conditions tested. These results suggest that DRN serotonergic and non-serotonergic neurons are activated by post-prandial signals. In contrast, the absence of Fos expression in the MRN suggests that this nucleus does not participate in the circuit involved in the control of post-prandial satiety.

Nonexocytotic serotonin release tonically suppresses serotonergic neuron activity.

The firing activity of serotonergic neurons in raphe nuclei is regulated by negative feedback exerted by extracellular serotonin (5-HT)o acting through somatodendritic 5-HT1A autoreceptors. The steady-state [5-HT]o, sensed by 5-HT1A autoreceptors, is determined by the balance between the rates of 5-HT release and reuptake. Although it is well established that reuptake of 5-HTo is mediated by 5-HT transporters (SERT), the release mechanism has remained unclear. It is also unclear how selective 5-HT reuptake inhibitor (SSRI) antidepressants increase the [5-HT]o in raphe nuclei and suppress serotonergic neuron activity, thereby potentially diminishing their own therapeutic effect. Using an electrophysiological approach in a slice preparation, we show that, in the dorsal raphe nucleus (DRN), continuous nonexocytotic 5-HT release is responsible for suppression of phenylephrine-facilitated serotonergic neuron firing under basal conditions as well as for autoinhibition induced by SSRI application. By using 5-HT1A autoreceptor-activated G protein-gated inwardly rectifying potassium channels of patched serotonergic neurons as 5-HTo sensors, we show substantial nonexocytotic 5-HT release under conditions of abolished firing activity, Ca(2+) influx, vesicular monoamine transporter 2-mediated vesicular accumulation of 5-HT, and SERT-mediated 5-HT transport. Our results reveal a cytosolic origin of 5-HTo in the DRN and suggest that 5-HTo may be supplied by simple diffusion across the plasma membrane, primarily from the dense network of neurites of serotonergic neurons surrounding the cell bodies. These findings indicate that the serotonergic system does not function as a sum of independently acting neurons but as a highly interdependent neuronal network, characterized by a shared neurotransmitter pool and the regulation of firing activity by an interneuronal, yet activity-independent, nonexocytotic mechanism.

Alteration in anxiety‐related behaviors and reduction of serotonergic neurons in raphe nuclei in adult rats prenatally exposed to ethanol

It is known that the developing serotonergic system is one of the targets of ethanol teratogenicity. Because serotonin has multiple functions in both mature and immature brains, disturbance of the serotonergic system by ethanol exposure in utero can be cause of a wide range of psychiatric problems in adulthood. In the present study, we observed serotonergic neurons in the midbrain raphe nuclei and anxiety-like behaviors which would be affected by an altered serotonergic system in adult rats prenatally exposed to ethanol. Pregnant rats were fed a liquid diet containing 2.5-5.0% (w/v) ethanol on gestational days 10-21. Their offspring were examined at 60-70 days of age. A significant decrease in the number of serotonergic cells in the midbrain raphe nuclei was shown in prenatally ethanol-exposed offspring. In an open field test, they spent more time in a central area compared to controls. Also in an elevated plus maze test, prenatally ethanol-exposed offspring spent more time on the open arms than controls. These behavioral results suggested that prenatally ethanol-exposed rats were less sensitive to anxiety. However, 44% of prenatally ethanol-exposed offspring exhibited freezing behavior on the open arms of the elevated plus maze, causing strong anxiety, compared with 0% in intact control and 12.5% in isocaloric sucrose-fed control groups. These findings suggest that prenatal ethanol exposure decreases both susceptibility and resistance of anxiety. Insufficient serotonergic actions caused by reduced serotonergic neurons in the raphe nuclei might contribute to the alterations in anxiety-related behaviors observed in our prenatally ethanol-exposed rats.

Variation in tryptophan hydroxylase-2 gene is not associated to male completed suicide in Estonian population

Dysfunction of the central serotonergic system has been related to a spectrum of psychiatric disorders, including suicidal behavior. Tryptophan hydroxylase isoform 2 (TPH2) is the rate-limiting enzyme in the biosynthetic pathway of serotonin, being expressed in serotonergic neurons of raphe nuclei. We investigated genetic variation in TPH2 gene in two samples of male subjects: 288 suicide completers and 327 volunteers, in order to reveal any associations between 14 single nucleotide polymorphisms and completed suicide. No associations were revealed neither on allelic nor haplotype level. Our finding does not support the hypothesis of TPH2 being a susceptibility factor for completed suicide in males of Estonian origin.