Habenular modulation of raphe indoleamine metabolism

Anatomical and neurophysiological findings have demonstrated neuronal connections between the diencephalic habenular nuclei and brain stem serotonergic raphe nuclei. Therefore we examined some neurochemical consequences of habenular lesions. Sixteen hours and one week after bilateral lesions serotonin metabolism (as reflected by concentrations of its metabolite, 5-hydroxyindoleacetic acid) was significantly increased in the dorsal but not the median raphe nuclei. Unilateral lesions produced a proportionally smaller augmentation. Motron locomotor activity was enhanced during the light and dark illumination phases in lesioned animals but only attained statistical significance during the day.     

Regional distribution and cellular expression of tryptophan hydroxylase messenger RNA in postmortem human brainstem and pineal gland

The characterization and cellular localization of tryptophan hydroxylase mRNA in the human brainstem and pineal gland were investigated by using northern blot analysis and in situ hybridization histochemistry. Northern analysis of human pineal gland revealed the presence of two mRNA species that were absent in RNA isolated from human raphe. In situ hybridization experiments revealed very dense hybridization signal corresponding to tryptophan hydroxylase mRNA in cells throughout the pineal gland. In contrast, tryptophan hydroxylase mRNA was heterogeneously distributed in neurons in the dorsal and median raphe nuclei. Within the dorsal raphe, the ventrolateral and interfascicular subnuclei contained the greatest number of tryptophan hydroxylase mRNA-positive neurons. Also, the cellular concentration of tryptophan hydroxylase mRNA varied widely within the dorsal and median raphe. Comparison of the cellular concentration of tryptophan hydroxylase mRNA between the pineal gland and the raphe nuclei revealed an 11- and 46-fold greater average grain density of tryptophan hydroxylase mRNA positive cells in the pineal gland compared with the dorsal and median raphe, respectively. These findings are the first to demonstrate the cellular localization of tryptophan hydroxylase mRNA in the human brain and pineal gland as well as heterogeneity in the cellular concentration within and between these tissues.     

中缝核5-羟色胺能神经投射对嗅球调制效应的研究进展

中缝核5-羟色胺能神经元通过其广泛的神经投射影响大脑多方面的功能,包括抑郁和焦虑、睡眠-觉醒周期、奖赏、决策中的耐心以及性别取向等.背侧中缝核和中央中缝核的5-羟色胺能神经元对嗅球有密集的神经投射,从而调控嗅觉信息的初步表征和编码.近年来,随着电生理、光学成像及光遗传技术的应用,关于中缝核5-羟色胺能神经元对嗅球的调制作用研究不断出现,大量离体和在体实验证据表明中缝核5-羟色胺能神经元对嗅球及嗅觉相关行为有广泛的调制.本文从嗅球不同神经元类型角度,就中缝核5-羟色胺能神经投射对嗅球的调控作用及其神经机制研究进展进行了总结.     

Independent effects of cholinergic and serotonergic lesions on acetylcholine and serotonin release in the neocortex of the rat

Rats received a unilateral lesion of the nucleus basalis magnocellularis (NBM) by infusion of ibotenic acid. In addition, the dorsal raphe nucleus was lesioned by infusion of 5,7-dihydroxytryptamine (5,7-DHT). The release of acetylcholine (ACh), choline, serotonin (5-HT) and 5-hydroxyindoleacetic acid (5-HIAA) was measured in the frontal neocortex by means of microdialysis. Lesions of the NBM, but not the raphe nucleus, reduced the release of ACh significantly (–47%). The release of 5-HT and 5-HIAA was reduced by raphe lesions (–44% and –79%), but not by NBM lesions. In no case did the combined lesion affect neurotransmitter release more than a single lesion. These results suggest that serotonergic projections from the dorsal raphe nucleus are not involved in tonic inhibition of ACh release in the neocortex.     

Differential Effects of Ipsapirone on 5-Hydroxytryptamine Release in the Dorsal and Median Raphe Neuronal Pathways

Abstract: Serotonergic neurons of the dorsal and median raphe nuclei are morphologically dissimilar. Recent results challenge previous evidence indicating a greater inhibition of dorsal raphe neurons after 5-hydroxytryptamine_1A (5-HT_1A) autoreceptor activation. As both nuclei innervate different forebrain territories, this issue is critical to understanding the changes in brain function induced by anxiolytic and antidepressant drugs. Using microdialysis, we examined the modifications of 5-HT release induced by the selective 5-HT_1A agonist ipsapirone in both neuronal pathways. Maximal and minimal basal 5-HT values (in the presence of 1 µ M citalopram) were 45.0 ± 4.8 fmol/fraction in the median raphe nucleus and 8.4 ± 0.4 fmol/fraction in the dorsal hippocampus. Ipsapirone (0.3, 3, and 10 mg/kg s.c.) reduced dose-dependently 5-HT in the two raphe nuclei and four forebrain areas. Maximal reductions (to ∼25% of predrug values) were observed in cortex and striatum and in median raphe nucleus. The effects were more moderate in dorsal and ventral hippocampus (to 66 and 50% of baseline, respectively). These results are consistent with a higher sensitivity of dorsal raphe neurons to 5-HT_1A autoreceptor activation. Yet the differential reduction of 5-HT release in the median raphe nucleus and hippocampus suggests the presence of complex mechanisms of control of 5-HT release in these neurons.     

Glutamic Acid Decarboxylase Activity of the Preoptic Area and Hypothalamus Is Influenced by the Serotonergic System

The effect of the serotonergic system on glutamic acid decarboxylase (GAD) activity of the preoptic area and the hypothalamus was studied in female rats on the day of proestrus. A circadian rhythm of GAD activity was observed with higher values in rats killed at 1130 h than in rats killed at 1500 h. In rats bearing lesions of the median raphe nucleus (MRn), a nucleus that sends 5-hydroxytryptamine nerve terminals to the areas under study decreased GAD activity. On the contrary, electrochemical stimulation of the MRn enhanced GAD activity in intact rats killed at 1500 h, but not in those killed at 1130 h, an effect that was prevented by the injection of the 5-hydroxytryptamine antagonist, methysergide. Furthermore, the injection of 5-hydroxytryptamine into the third ventricle, either in intact rats in the afternoon or in MRn-lesioned rats in the morning, also increased GAD activity. The results of the present study suggest that activation of the serotonergic system increases GAD activity in the preoptic area and hypothalamus.     

Preferential Potentiation of the Effects of Serotonin Uptake Inhibitors by 5-HT1A Receptor Antagonists in the Dorsal Raphe Pathway: Role of Somatodendritic Autoreceptors

Abstract: 5-HT_1A autoreceptor antagonists enhance the effects of antidepressants by preventing a negative feedback of serotonin (5-HT) at somatodendritic level. The maximal elevations of extracellular concentration of 5-HT (5-HT_ext) induced by the 5-HT uptake inhibitor paroxetine in forebrain were potentiated by the 5-HT_1A antagonist WAY-100635 (1 mg/kg s.c.) in a regionally dependent manner (striatum > frontal cortex > dorsal hippocampus). Paroxetine (3 mg/kg s.c.) decreased forebrain 5-HT_ext during local blockade of uptake. This reduction was greater in striatum and frontal cortex than in dorsal hippocampus and was counteracted by the local and systemic administration of WAY-100635. The perfusion of 50 µmol/L citalopram in the dorsal or median raphe nucleus reduced 5-HT_ext in frontal cortex or dorsal hippocampus to 40 and 65% of baseline, respectively. The reduction of cortical 5-HT_ext induced by perfusion of citalopram in midbrain raphe was fully reversed by WAY-100635 (1 mg/kg s.c.). Together, these data suggest that dorsal raphe neurons projecting to striatum and frontal cortex are more sensitive to self-inhibition mediated by 5-HT_1A autoreceptors than median raphe neurons projecting to the hippocampus. Therefore, potentiation by 5-HT_1A antagonists occurs preferentially in forebrain areas innervated by serotonergic neurons of the dorsal raphe nucleus.     

中缝背核5-羟色胺能神经元在睡眠调节中的作用研究

目的：研究中缝背核(DRN)5-羟色胺(5-HT)能神经元在睡眠中的调节作用。方法：运用脑立体定位、核团微量注射和多导睡眠描记(PSG),观察DRN 5-HT能神经元对大鼠睡眠的影响。结果：DRN微量注射谷氨酸钠(L-Glu),大鼠睡眠减少,特别是深慢波睡眠(SWS2)明显减少,觉醒(W)增加;DRN微量注射海人酸(KA)和对氯苯丙氨酸(PCPA),大鼠SWS2和异相睡眠(PS)增加,W减少。结论：DRN 5-HT能神经元参与睡眠的调节,兴奋DRN 5-HT能神经元睡眠时间减少,抑制DRN 5-HT能神经元则具有促进睡眠的作用。     

The Distribution of Serotonergic Nerves in Microencephalic Rats Treated Prenatally with Methylazoxymethanol

Prenatal exposure of pregnant rats to methylazoxymethanol acetate (MAM) induces microencephaly in the offspring. In the present study of these microencephalic rats (MAM rats) we used quantitative autoradiography to investigate [³H] paroxetine binding sites, which are a selective marker of serotonin (5-HT) transporters (5-HTT). The binding in the accumbens, cortex, hippocampus, and dorsolateral thalamus was significantly increased in MAM rats, compared to the control rats, while there was a significant decrease in the dorsal raphe nucleus of the MAM rats. The levels of 5-HTT mRNA in the dorsal raphe nuclei were analyzed by in situ hybridization, which revealed a significant decrease in 5-HTT mRNA-positive neurons in the MAM rats compared to the control rats. The results imply serotonergic hyperinnervation in the cerebral hemispheres of MAM rats, while a target-dependent secondary degeneration of 5-HT neurons might be induced in the dorsal raphe nuclei of MAM rats.     

Galanin Alters GABAergic Neurotransmission in the Dorsal Raphe Nucleus

The neuropeptide galanin and its three receptor subtypes (Gal R1-3) are highly expressed in the dorsal raphe nucleus (DRN), a region of the brain that contains a large population of serotonergic neurons. Galanin is co-expressed with serotonin in approximately 40% of the DRN neurons, and galanin and GALR2 expression are elevated by antidepressants like the SSRI fluoxetine, suggesting an interaction between serotonin and galanin. The present study examines the effect of galanin (Gal 1–29), a pan ligand for GalR (1–3) and the GalR2/GalR3-selective ligand, Gal 2–11, on the electrophysiological properties of DRN serotonergic neurons in a slice preparation. We recorded from cells in the DRN with electrophysiological characteristics consistent with those of serotonergic neurons that exhibit high input resistance, large after-hyperpolarizations and long spike duration as defined by Aghajanian and Vandermaelen. Both Gal 1–29 and Gal 2–11 decreased the amplitudes pharmacologically-isolated GABAergic inhibitory postsynaptic potentials (IPSPs) in these putative serotonergic neurons. Furthermore, based on paired pulse facilitation studies, we show that Gal 1–29 likely decreases GABA release through a presynaptic mechanism, whereas Gal 2–11 may act postsynaptically. These findings may enhance understanding of the cellular mechanisms underlying the effects of antidepressant treatments on galanin and galanin receptors in DRN. Special issue article in honor of Dr. Frode Fonnum.     

p-Chlorophenylalanine Changes Serotonin Transporter mRNA Levels and Expression of the Gene Product

Abstract: After a single intraperitoneal injection of the irreversible tryptophan hydroxylase inhibitor p -chlorophenylalanine (PCPA; 300 mg/kg), there was a rapid down-regulation of serotonin (5-HT) transporter mRNA levels in cell bodies. This change was significant at 1 and 2 days after PCPA administration within the ventromedial but not the dorsomedial portion of the dorsal raphe nucleus. Seven days after PCPA treatment, 5-HT transporter mRNA levels were significantly elevated compared with controls in both regions of the dorsal raphe nucleus. PCPA administration produced no change in the [³H]-citalopram binding and synaptosomal [³H]5-HT uptake in terminal regions at 2 and 7 days after treatment but significantly reduced both these parameters by ∼20% in the hippocampus and in cerebral cortex 14 days after PCPA administration. The striatum showed a lower sensitivity to this effect. No significant changes were observed in the levels of [³H]citalopram binding to 5-HT cell bodies in the dorsal raphe nucleus. In the same animals used for 5-HT transporter mRNA level measurements, levels of tryptophan hydroxylase mRNA in neurons of the ventromedial and dorsomedial portions of the dorsal raphe nucleus were increased 2 days after PCPA administration and fell to control levels 7 days after injection in the ventromedial region but not in the dorsomedial portion of the dorsal raphe nucleus, where they remained significantly higher than controls. Altogether, these results show that changes in 5-HT transporter mRNA are not temporally related to changes in 5-HT transporter protein levels. In addition, our results suggest that the 5-HT transporter and tryptophan hydroxylase genes are regulated by different mechanisms. We also provide further evidence that dorsal raphe 5-HT neurons are differentially regulated by drugs, depending on their location.     

Modulation of the hippocampal alpha-adrenoceptor population by lesion of the serotonergic raphe-hippocampal pathway in rats

Electrolytic lesion of the ascending serotonergic fibers in the median raphe nucleus or in both the median raphe nucleus and dorsal raphe nucleus caused after 18 days more than 80% depletion of serotonin in the hippocampus and frontal cortex, respectively, without affecting norepinephrine and acetylcholine contents. alpha 1-Adrenoceptor binding of (3H) WB-4104 was increased in the hippocampus but not in the frontal cortex. Scatchard analysis revealed that the increase in (3H) WB-4101 binding in the lesioned hippocampus was the result of an elevated density of alpha 1-adrenergic receptors of about 65%. This phenomenon began 8 days postlesion and persisted for at least 90 days postlesion. Similar qualitative and quantitative results were obtained following chemical lesion of the serotonergic cells of origin in the median raphe nucleus with 5,7-dihydroxytryptamine. Selectivity of the phenomenon was further demonstrated as or beta-adrenoceptor binding with (3H) dihydroalprenolol and cholinergic muscarinic receptor binding with (3H) dexetimide were not significantly affected in the hippocampus. By comparison, when norepinephrine in the hippocampus was depleted by more than 90% by bilateral lesion of the ascending noradrenergic fibers with 6-hydroxydopamine (18 days), the alpha 1-adrenoceptor number was significantly increased by only about 20% while the beta-adrenoceptor number was enhance by 40%. The area-selective increase in alpha 1-adrenoceptor number in the hippocampus in the presence of unchanged norepinephrine content and in the absence of serotonin probably signifies that serotonin actively participates in the modulation of the noradrenergic receptor population.     

In vivo interaction between serotonin and galanin receptors types 1 and 2 in the dorsal raphe: implication for limbic seizures

The neuropeptide galanin suppresses seizure activity in the hippocampus by inhibiting glutamatergic neurotransmission. Galanin may also modulate limbic seizures through interaction with other neurotransmitters in neuronal populations that project to the hippocampus. We examined the role of galanin receptors types 1 and 2 in the dorsal raphe (DR) in the regulation of serotonergic transmission and limbic seizures. Infusion of a mixed agonist of galanin receptors types 1 and 2 [galanin (1-29)] into the DR augmented the severity of limbic seizures in both rats and wild-type mice and concurrently reduced serotonin concentration in the DR and hippocampus as measured by immunofluorescence or HPLC. In contrast, injection of the galanin receptor type 2 agonist galanin (2-11) mitigated the severity of seizures in both species and increased serotonin concentration in both areas. Injection of both galanin fragments into the DR of galanin receptor type 1 knockout mice exerted anticonvulsant effects. Both the proconvulsant activity of galanin (1-29) and seizure suppression by galanin (2-11) were abolished in serotonin-depleted animals. Our data indicate that, in the DR, galanin receptors types 1 and 2 modulate serotonergic transmission in a negative and a positive fashion, respectively, and that these effects translate into either facilitation or inhibition of limbic seizures.     

Optogenetic Recruitment of Dorsal Raphe Serotonergic Neurons Acutely Decreases Mechanosensory Responsivity in Behaving Mice

The inhibition of sensory responsivity is considered a core serotonin function, yet this hypothesis lacks direct support due to methodological obstacles. We adapted an optogenetic approach to induce acute, robust and specific firing of dorsal raphe serotonergic neurons. In vitro, the responsiveness of individual dorsal raphe serotonergic neurons to trains of light pulses varied with frequency and intensity as well as between cells, and the photostimulation protocol was therefore adjusted to maximize their overall output rate. In vivo, the photoactivation of dorsal raphe serotonergic neurons gave rise to a prominent light-evoked field response that displayed some sensitivity to a 5-HT1A agonist, consistent with autoreceptor inhibition of raphe neurons. In behaving mice, the photostimulation of dorsal raphe serotonergic neurons produced a rapid and reversible decrease in the animals'' responses to plantar stimulation, providing a new level of evidence that serotonin gates sensory-driven responses.     

Cortical and septal responses to dorsal raphe nucleus stimulation in the rat: long-term clomipramine actions.

In cerebral cortex and lateral septal nuclei different serotonergic receptor subtypes coexist, thus a different action on neuronal firing may be expected depending on the receptor activated. Dorsal raphe nucleus stimulation produced an increased rate of firing in cortical layer V, and in lateral septal nuclei. However, firing rate in cortical layer VI remained unchanged after stimulating the dorsal raphe nucleus. Clomipramine is a tricyclic which exerts its main actions on serotonergic receptors, and long-term treatment with this antidepressant produced a selective increased firing rate in lateral septal neurons, but not in cortical neurons. From an electrophysiological point of view, it is concluded that the excitatory actions on firing rate elicited by dorsal raphe nucleus stimulation or clomipramine treatment are mediated by 5-HT2 receptor subtype activation which is likely to be acting as a 5-HT1A modulator in such places where both receptor subtypes coexist.     

Synthesis of Serotonin in Traumatized Rat Brain

Abstract: Previous studies have demonstrated that focal freezing lesions in rats cause a widespread decrease of cortical glucose use in the lesioned hemisphere and this was interpreted as a reflection of depression of cortical activity. The serotonergic neurotransmitter system was implicated in these alterations when it was shown that (1) cortical serotonin metabolism was increased widely in focally injured brain and (2) inhibition of serotonin synthesis prevented the development of cortical hypometabolism. In the present studies we applied an autoradiographic method that uses the accumulation of the ¹⁴C-labeled analogue of serotonin α-methylserotonin to assess changes in the rate of serotonin synthesis in injured brain. The results confirmed that 3 days after the lesion was made, at the time of greatest depression of glucose use, serotonin synthesis was significantly increased in cortical areas throughout the injured hemisphere. The increase was also seen in the dorsal hippocampus and area CA3, as well as in the medial geniculate and dorsal raphe, but not in any other subcortical structures including median raphe. Present results suggest that the functional changes in the cortex of the lesioned hemisphere are associated with an increased rate of serotonin synthesis mediated by activation of the dorsal raphe. We also documented by α-[¹⁴C]aminoisobutyric acid autoradiography that there was increased permeability of the blood-brain barrier, but this was restricted to the rim of the lesion.