The comparative effect of serotonin agonists on the presynaptic and somatodendritic autoreceptors of serotoninergic neurons

Experimenting on the slices of cortex and dorsal raphe nucleus of midbrain of rats which were incubated with 3H-hydroxytrypta-mine (3H-HT) studies showed the influence of series of serotonin agonists on the spontaneous and electrically stimulated release of 3H-HT from the slices. It was established that the serotonin in concentration of 10(-5) mol/l similarly inhibits the release of 3H-HT from the electrically stimulated slices of the brain cortex (78.6%) and on slices of the dorsal raphe nucleus of the midbrain (81.6%) had no effect on the spontaneous release of serotonin. The serotonin agonists in order of increasing ability to inhibit the electrically stimulated release of 3H-HT from the cortex slices is as follows: ipsapirone (0%), 8-OH-DPAT (23%), kampirone (26.5%), 1.2-PP (28.6%), kaplapirone (35.7%), buspirone (48%) and TFMPP (67%). On the ability to influence the release of 3H-HT from the electrically stimulated slices of the dorsal raphe nucleus of the midbrain of the rats serotonin agonists were in the following order: TEMPP (12.3%), kampirone (40%), 1.2-PP (42.9%), ipsapirone (52%), 8-OH-DPAT (54.1%), kampirone (57.2%) and buspirone (65.3%). It is suggested that the effect of both ipsapirone, kampirone and 8-OH-DPAT is greatly localized on the somato-dendritic synapses P1A-HT receptors, TEMPP is more on the terminal axons of HT-ergic neurones while kampirone, buspirone and active metabolite 1.2-PP act on the presynaptic and somatodendritic autoreceptors of serotonin.     

Regional Distribution of Extracellular 5-Hydroxytryptamine and 5-Hydroxyindoleacetic Acid in the Brain of Freely Moving Rats

The extracellular concentrations of 5-hydroxytryptamine (5-HT) and its metabolite 5-hydroxyindoleacetic acid (5-HIAA) have been determined in six brain areas of awake rats (frontal cortex, striatum, hypothalamus, hippocampus, inferior colliculus, and raphe nuclei) using intracerebral microdialysis. The extracellular levels of 5-HT showed no significant differences among the brain regions studied. The tissue levels of 5-HT and 5-HIAA as well as the extracellular concentration of 5-HIAA were significantly higher in raphe nuclei. The regional distribution of tissue and extracellular 5-HIAA were very similar, suggesting that extracellular 5-HIAA depends mainly on the output from the intracellular compartment. On the other hand, extracellular 5-HT and tissue 5-HT showed a different distribution pattern. The tissue/extracellular ratio for 5-HT ranged from 739 in frontal cortex to 2,882 in raphe, whereas it only amounted to 1.8-3.6 for 5-HIAA. The relationship between the present results and the density of 5-HT uptake sites in these areas is discussed.     

Participation of the brain serotoninergic system in creating the stress reactivity of the hypophyseal-adrenal axis

The concentration of corticosteroids in the blood of rats was shown to increase in response to the immobilization stress at an earlier age than the brain serotonin metabolism changes. The level of corticosteroids in blood increased in response to the intraperitoneal serotonin injection also earlier than the reaction to the serotonin injection in the brain lateral ventricle sets up. The increase of the reaction of hypophysial-suprarenal system to stress during the period from the 12th till the 16th day of postnatal development coincided with the changes in serotonin metabolism in the brain stem and the reaction to serotonin injection in the brain lateral ventricle. It is suggested that the system of serotonin brain neurons connected with the hypophysial-suprarenal complex matures later tran the serotonin receptors in the periphery; the reaction to immobilization may be realized at the early developmental stages without the participation of brain serotonin.     

The cytochrome P450 2D‐mediated formation of serotonin from 5‐methoxytryptamine in the brain in vivo: a microdialysis study

The cytochrome P450 2D (CYP2D) mediates synthesis of serotonin from 5‐methoxytryptamine (5‐MT), shown in vitro for cDNA‐expressed CYP2D‐isoforms and liver and brain microsomes. We aimed to demonstrate this synthesis in the brain in vivo. We measured serotonin tissue content in brain regions after 5‐MT injection into the raphe nuclei (Model‐A), and its extracellular concentration in rat frontal cortex and striatum using an in vivo microdialysis (Model‐B) in male Wistar rats. Naïve rats served as control animals. 5‐MT injection into the raphe nuclei of PCPA‐(tryptophan hydroxylase inhibitor)‐pretreated rats increased the tissue concentration of serotonin (from 40 to 90% of the control value, respectively, in the striatum), while the CYP2D inhibitor quinine diminished serotonin level in some brain structures of those animals (Model‐A). 5‐MT given locally through a microdialysis probe markedly increased extracellular serotonin concentration in the frontal cortex and striatum (to 800 and 1000% of the basal level, respectively) and changed dopamine concentration (Model‐B). Quinine alone had no effect on serotonin concentration; however, given jointly with 5‐MT, it prevented the 5‐MT‐induced increase in cortical serotonin in naïve rats and in striatal serotonin in PCPA‐treated animals. These results indicate that the CYP2D‐catalyzed alternative pathway of serotonin synthesis from 5‐MT is relevant in the brain in vivo, and set a new target for the action of psychotropics.

     

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.     

Alpha-neo-endorphin-like immunoreactivity in the cat brain stem

This paper examines the distribution of fibers and cell bodies containing alpha-neo-endorphin in the cat brain stem by using an indirect immunoperoxidase technique. A high or moderate density of immunoreactive cell bodies was found in the superior central nucleus, nucleus incertus, dorsal tegmental nucleus, nucleus of the trapezoid body, and in the laminar spinal trigeminal nucleus, whereas a low density of such perikarya was observed in the inferior colliculus, nucleus praepositus hypoglossi, dorsal nucleus of the raphe, nucleus of the brachium of the inferior colliculus, and in the nucleus of the solitary tract. The highest density of immunoreactive fibers was found in the substantia nigra, dorsal motor nucleus of the vagus, nucleus coeruleus, lateral tegmental field, marginal nucleus of the brachium conjunctivum, and in the inferior and medial vestibular nuclei. These results indicate that alpha-neo-endorphin is widely distributed in the cat brain stem and suggest that the peptide could play an important role in several physiological functions, e.g., those involved in respiratory, cardiovascular, auditory, and motor mechanisms.     

Effects of pentadecapeptide BPC157 on regional serotonin synthesis in the rat brain: alpha-methyl-L-tryptophan autoradiographic measurements

A novel pentadecapeptide, BPC157, was recently reported to have a large spectrum of in vivo activities, from anti-ulcer to central action on the brain dopaminergic system. The mechanisms of these actions are not well understood. In this study, the evaluation of the effects of acute and repeated administration of BPC157 on serotonin (5-HT) synthesis in the rat brain is reported. The alpha-[14C]methyl-L-tryptophan (alpha-MTrp) autoradiographic method was used to measure regional 5-HT synthesis rates. In the first series of experiments, a single dose treatment of BPC157 (10 microg/kg) administered intraperitoneally 40 min before the alpha-MTrp tracer injection significantly reduced the regional rate of 5-HT synthesis in the dorsal thalamus, hippocampus, lateral geniculate body and hypothalamus. 5-HT synthesis rates in the substantia nigra reticulate and medial anterior olfactory nucleus in BPC157 treated rats were significantly higher than in the control rats. No significant change in the synthesis rate was observed in the raphe nuclei. In the second series of experiments, following a 7-day treatment with BPC157 (10 microg/kg; s.c.), a significant reduction in the 5-HT synthesis rate was observed in the dorsal raphe nucleus, and significant increases were observed in the substantia nigra, lateral caudate, accumbens nucleus and superior olive. This data suggests that BPC157, a gut peptide, influences brain 5-HT synthesis in rats, but we cannot determine, from this data, the mechanism of this action.     

Brain 5-HT synthesis in the Flinders Sensitive Line rat model of depression: an autoradiographic study

Alterations of serotonin (5-HT) levels and serotonergic transmission have been associated with depression. 5-HT synthesis is an important factor of serotonergic neurotransmission that may also be altered in depression. Many studies of the relationships between brain serotonergic functions and affective disorders have been performed in different animal models. In this study, brain regional 5-HT synthesis was examined using the alpha-[(14)C]methyl-L-tryptophan (alpha-MTrp) autoradiographic method in a genetic rat model of depression, Flinders Sensitive Line (FSL) rats, and was compared to both the Flinders Resistant Line (FRL) rats and the control Sprague-Dawley (SD) rats. The plasma concentration of free tryptophan in the FSL rats was not significantly different (p > 0.05; ANOVA and post-hoc Bonferroni correction) when compared to that of the FRL and SD rats. The FSL rats had significantly lower 5-HT synthesis (one sample two-tailed t-test on the ratio) than both the FRL and SD rats (the mean ratios were 0.78 +/- 0.12 and 0.73 +/- 0.15, respectively). Overall, the 5-HT synthesis in the FRL rats was not significantly different (p > 0.05) from that in the SD rats (one sample two-tailed t-test on the ratio and the mean ratio was 0.93 +/- 0.13). Studies of individual brain structures, such as the raphe nuclei and their many terminal areas, including the nucleus accumbens, cingulate and frontal cortex, hippocampus, amygdala, and thalamus revealed significant reductions (typically 25-50%) in 5-HT synthesis in the FSL rats compared to the non-depressive FRL and SD rats. These results suggest that significantly reduced 5-HT synthesis in the raphe nuclei and limbic areas in FSL rats may contribute to their depressive features.     

The Sequential Alterations of Endoneurial Cholesterol and Fatty Acid in Wallerian Degeneration and Regeneration

Abstract: The rate of tryptophan hydroxylation in vivo was estimated in discrete rat brain nuclei by measuring L-5-hydroxytryptophan (5-HTP) accumulated after pharmacological blockade of ^L-5-hydroxytryptophan decarboxylase by NSD 1015, using a sensitive radioenzymatic microassay. Endogenous serotonin, a major contaminant in this assay, was quantitatively removed by cationexchange chromatography prior to analysis. In non-treated animals, endogenous 5-HTP could be detected in small but measurable amounts. Following NSD 1015, accumulation occurred linearly for at least 30 min. At this time the recorded figures were two to six times higher when compared to values obtained in the same discrete structure from non-treated animals. This allows an accurate estimation of the rate of tryptophan hydroxylation in vivo in small fragments of grossly dissected brain regions (e.g. cortex) as well as in discrete nuclei containing either serotoninergic (5-HT) cell bodies (brain stem raphe nuclei) or 5-HT-terminals (e.g. catecholaminergic group A l, A2, A6.,. etc). Parachlorophenylalanine drastically reduced the rate of tryptophan hydroxylation in vivo in both terminal regions and raphe nuclei, with similar figures, 3 h or 3 days after injection. Chloral hydrate anaesthesia was attended by a transient decrease which appeared delayed in the raphe nuclei. Finally, pargyline pretreatment led to an 80% decrease in the forebrain, while no significant change appeared in the raphe nuclei. Thus, as illustrated by these few pharmacological manipulations, this method allows the study of the regulation of tryptophan hydroxylation in vivo with an improved anatomical resolution. Investigations can be carried out in the various raphe nuclei and their corresponding terminals in discrete brain areas simultaneously.     

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.     

Immunohistochemical study on the distribution of serotonin-containing cell bodies in the brain stem of the dog

The distribution of serotonin-containing neurons in the brain of the dog was studied by use of PAP immunohistochemistry. The lower brain stem was endowed with extensively scattered serotonergic cell bodies, a large portion of which was located in the raphe nuclei. At the same time, prominent distribution of serotonergic neurons in lateral areas outside the raphe nuclei was also demonstrated. Our observations on the brain stem were, in principle, consistent with those on rats, cats and monkeys, with only minor differences.     

Brain prolactin is involved in stress-induced REM sleep rebound

REM sleep rebound is a common behavioural response to some stressors and represents an adaptive coping strategy. Animals submitted to multiple, intermittent, footshock stress (FS) sessions during 96 h of REM sleep deprivation (REMSD) display increased REM sleep rebound (when compared to the only REMSD ones, without FS), which is correlated to high plasma prolactin levels. To investigate whether brain prolactin plays a role in stress-induced REM sleep rebound two experiments were carried out. In experiment 1, rats were either not sleep-deprived (NSD) or submitted to 96 h of REMSD associated or not to FS and brains were evaluated for PRL immunoreactivity (PRL-ir) and determination of PRL concentrations in the lateral hypothalamus and dorsal raphe nucleus. In experiment 2, rats were implanted with cannulas in the dorsal raphe nucleus for prolactin infusion and were sleep-recorded. REMSD associated with FS increased PRL-ir and content in the lateral hypothalamus and all manipulations increased prolactin content in the dorsal raphe nucleus compared to the NSD group. Prolactin infusion in the dorsal raphe nucleus increased the time and length of REM sleep episodes 3 h after the infusion until the end of the light phase of the day cycle. Based on these results we concluded that brain prolactin is a major mediator of stress-induced REMS. The effect of PRL infusion in the dorsal raphe nucleus is discussed in light of the existence of a bidirectional relationship between this hormone and serotonin as regulators of stress-induced REM sleep rebound.     

Temporal Distribution of [3H]-Imipramine Binding in Rat Brain Regions is Not Changed by Chronic Methamphetamine

Specific binding of [³H]-imipramine in the rat suprachiasmatic nuclei, occipital cortex and caudate putamen underwent significant and replicable changes throughout 24 hr under a light-dark cycle or under constant conditions. Daily variations were also found in the medial and dorsal raphe nuclei and the lateral hypothalamus. Methamphetamine, a psychoactive drug with marked effect on circadian rhythms in physiological and hormonal parameters and adrenergic receptors, did not have any significant effect on imipramine binding rhythms in eight discrete brain regions. Thus a drug known to reduce serotoninergic neurotransmission did not change characteristics of the modulatory binding site related to serotonin uptake.     

Effect of specific serotonergic lesions on cholinergic neurons in the hippocampus,cortex and striatum

Local injection of 5, 7-dihydroxytryptamine into the median raphe nucleus of rats pretreated with desipramine decreases the serotonin content of the hippocampus and cortex. The turnover of acetylcholine, as measured by the rate of decline of acetylcholine content after hemicholinium-3, the rate of decline of acetylcholine content after hemicholinium-3, is not affected in the hippocampus or the striatum, but is increased in the cortex by such treatment. Local injection of 5, 7-dihydroxytryptamine into the dorsal raphe nucleus of desipramine-treated rats decreases the serotonin content of the hippocampus, cortex, and striatum. The turnover of acetylcholine is increased in the hippocampus and cortex, but not affected in the striatum. Thus, serotonergic neurons from the median raphe nucleus appear to tonically inhibit cholinergic neurons in the cortex, and serotonergic neurons from the dorsal raphe nucleus appear to tonically inhibit cholinergic neurons in the hippocampus and cortex. These serotonergic neurons do not appear to act tonically on striatal cholinergic neurons.     

Identification of serotonin-modulated protein fraction and study of its role in passive avoidance reaction in rats]

The effect 10(-3) M serotonin and norepinephrine solutions application on the brain occipital cortex on discrete water soluble protein fractions content was investigated in narcotized rats. It was revealed that application of serotonin solution caused repeated decrease in N6 fraction content. Immunoglobulins against N6 fraction being injected into the lateral ventricle of rat brain increased the number of photocell crossing and the presence probability of rats at the dark compartment entrance region in passive avoidance schedule. The conclusion of selective serotonin participation in regulation of brain cortex discrete proteins turnover and of these proteins involvement in searching behavior was made.     

Habenula--a new target for treatment of intractable depression

Despite substantial advancement in psychopharmacological and electro-magnetic treatments over the last decades on the depression patients, there are non-responders remain with a chronic disease and high suicidal risk yet. Deep brain stimulation (DBS) is now being experimentally to treat the intractable depression and yielded an impressive therapeutic benefit, and especially few adverse effect occurred. The beneficial action of DBS is closely related to the stimulation sit. And the efficacy of high frequency stimulation of lateral habenula is one of the best choice. In depression, the concentration of 5-HT released by the raphe nuclei is decreased. It's due to mainly the overactivation of the lateral habenula. High frequency stimulation of lateral habenula impairs the activation of lateral habenula, and the inhibitory effect of lateral habenula on raphe nuclei is decreased. Then, the 5-HT concentration released by the raphe nuclei is increased, the pathological changes of depression is eliminated. The lateral habenula could be a promising novel target for BDS in the cases of intractable depression.     

Pharmacological modification of the serotonergic transmitter system and beta-N-acetylhexosaminidase activity in rats

The enzyme activity and activation energy of plasma beta-N-acetylhexosaminidase (Hex) was determined in rats whose serotonergic system had been pharmacologically altered. In the group of animals treated with 5-hydroxytryptophan, in the different dissected brain regions (brain stem, cortex and hippocampus) significantly higher levels of serotonin and 5-hydroxyindolacetic acid were found, and significantly lower in the group treated with p-chlorophenylalanine, than in the control group. In the total number of animals studied (n = 21), a statistically significant correlation was found between the plasma concentration of 5-hydroxyindolacetic acid and the levels of this metabolite in the different brain regions (p < 0.001). No significant differences were found for the activity of Hex in the plasma, or for its activation energy, which is a marker of its isoenzyme composition, among the three groups of animals. The results obtained using our experimental model in rats do not confirm the hypothesis of other authors who suggest that the Hex responds secondary to increases or decreases of serotonin turnover, and could be a biological test to monitor the serotonin status in psychiatric patients.     

Acute and Chronic D-Fenfluramine Treatments Have Different Effects on Serotonin Synthesis Rates in the Rat Brain: An Autoradiographic Study

The effects of acute and chronic treatments with D-fenfluramine on the regional rates of serotonin (5-hydroxy-tryptamine; 5-HT) synthesis were investigated using the -[¹⁴C]methyl-L-tryptophan (-[¹⁴C]MTrp) autoradiographic method. In the first series of experiments, acute D-fenfluramine treatment (5 mg/kg; i.p.) given 20 min before the tracer injection significantly (p < 0.05) decreased 5-HT synthesis in the dorsal raphe, and significantly (p < 0.05) increased the rates in the cerebral cortices and caudate nucleus, when compared to the rates in the control rats (saline treated). In a second series of experiments, following a 7-day treatment with D-fenfluramine (5 mg/kg/day; i.p.), a significant (p < 0.05) decrease of 5-HT synthesis, in the dorsal raphe was observed, and significant (p < 0.05) increases were observed in the hypothalamus, the dorsal thalamus, the medial and lateral geniculate body and some brain stem regions (locus ceruleus, inferior and superior colliculus). No significant changes were observed in the cerebral cortices.     

Regulation of catecholamine synthesis in rat brain synaptosomes

Abstract— Catecholamine synthesis in synaptosomal preparations of rat striatum, cortex and brain stem was investigated. The striatum had much higher activity than either the cortex or brain stem. Equilibration of labelled tyrosine between tissue and incubation medium was completed within 2 min. The apparent K_m of tyrosine hydroxylase (EC 1.14.3a) and of the overall catecholamine synthetic pathway were both approximately 5 ± 10^?6m for tyrosine. The following amines were found to inhibit striatal dopamine synthesis: dopamine, 25% inhibition at 5 ± 10^?7m ; noradrenaline, 25% inhibition at 5 ± 10^?6m ;and serotonin, 30% inhibition at 10^?5m . The catecholamine-induced inhibition of synthesis was antagonized by pre-incubation with cocaine. Increasing the potassium concentration from 5 to 55 mm caused a release of amines into the medium which was accompanied by a 40% increase in dopamine synthesis, when synthesis was measured during the first 5 min of exposure to elevated potassium. These results indicate that synaptosomal catecholamine synthesis is inhibited by increases in intra-synaptosomal amine levels, and that short-term exposure to depolarizing concentrations of potassium can increase synthesis.     

Developmental changes in the brain-stem serotonergic nuclei of teleost fish and neural plasticity

Summary 1. During early ontogeny, the serotonergic neurons in the brain stem of the three-spined stickleback shows a temporal and spatial developmental pattern that closely resembles that of amniotes.2. However, in the adult fish, only the midline nuclei of the rostral group (dorsal and median raphe nuclei) and the dorsal lateral tegmental nucleus are consistently serotonin-immunoreactive (5-HTir), whereas the groups of the upper and lower rhombencephalon (raphe pontis, raphe magnus, and raphe pallidus/obscurus nuclei) are variable and, when present, contain relatively small numbers of 5-HTir neurons.3. Using specific antisera against tryptophan 5-hydroxylase and aromaticl-amino acid decarboxylase, we have shown that the lateral B9 group and the groups of the upper and lower rhombencephalon are consistently present in adult sticklebacks. The results are discussed in relation to other known instances of neurotransmitter plasticity or transient neurotransmitter expression in teleost fish.4. While there are several instances of transient expression of neurotransmitter markers by discrete neuronal populations, there is so far no evidence of changes from one neurotransmitter phenotype to another in the brain of teleost fish. However, there are indications of plasticity of expression of catecholamines and indoleamines, and their respective synthesizing enzymes, as reflected in age-dependent changes and variation between individuals of different physiological status.5. As the brain grows continuously in teleost fish, and new neurons are added from proliferative regions, synaptic connections may be expected to undergo remodeling in all brain regions throughout life. Thus, the teleostean brain may be considered a suitable model for experimental studies of different aspects of neural plasticity.