Serotonin receptors antagonistically modulate Caenorhabditis elegans longevity

The neurotransmitter serotonin has been implicated in affecting the variation of longevity in natural Drosophila populations and age-related diseases in mammals. Based on these observations, it has been predicted that serotonin signal, perhaps at levels of serotonin biosynthesis, may control lifespan. Here, we investigated a variety of mutations in serotonin-signal genes, including serotonin biosynthesis genes, a serotonin transporter gene, and serotonin receptor genes. Despite this prediction, mutations in the serotonin biosynthesis genes had little or modest effects on lifespan, while the mod-5 mutation with increased availability of serotonin caused a modest life-shortening effect. In contrast, a deletion mutation of the ser-1 serotonin receptor gene increased longevity by up to 46%, likely through the insulin/insulin-like growth factor 1 pathway. This result suggests an interaction between the serotonin pathway and the insulin/insulin-like growth factor 1 pathway. A deletion mutation of another serotonin receptor gene, ser-4 , shortened early to mid lifespan. The results suggest that serotonin signal antagonistically modulates longevity through different serotonin receptors. This study may indicate serotonin receptors as a potential target for antigeric interventions.     

Vervet monkey (Cercopithecus aethiops sabaeus) whole blood serotonin level is determined by platelet uptake sites

Whole blood serotonin levels in adult male vervet monkeys living in social groups are sensitive to the animals' social environment. The mechanisms that translate different behavioral and environmental cues into altered whole blood serotonin levels are unknown. In this study, we have measured platelet number, size, serotonin content, and serotonin uptake, as well as the serum concentrations of tryptophan, Mg+2 and Ca+2. Results showed that whole blood serotonin levels, platelet serotonin content, and the serotonin uptake parameter Vmax were stable within animals on repeated sampling. The whole blood serotonin level was highly positively associated with platelet serotonin content, and the platelet serotonin content was highly positively associated with Vmax. These findings suggested that whole blood serotonin levels were a function of the number of platelet uptake sites.     

Effect of serotonin on small intestinal contractility in healthy volunteers

The physiological significance of serotonin released into the intestinal lumen for the regulation of motility is unknown in humans. The aim of this study was to evaluate the effect of serotonin infused into the lumen of the gastric antrum, duodenum or the jejunum, on antro-duodeno-jejunal contractility in healthy human volunteers. Manometric recordings were obtained and the effects of either a standard meal, continuous intravenous infusion of serotonin (20 nmol/kg/min) or intraluminal bolus infusions of graded doses of serotonin (2.5, 25 or 250 nmol) were compared. In addition, platelet-depleted plasma levels of serotonin, blood pressure, heart rate and electrocardiogram were evaluated. All subjects showed similar results. Intravenous serotonin increased migrating motor complex phase III frequency 3-fold and migrating velocity 2-fold. Intraluminal infusion of serotonin did not change contractile activity. Platelet-depleted-plasma levels of serotonin increased 2-fold following both intravenous and high doses of intraluminal infusions of serotonin. All subjects reported minor short-lived adverse effects following intravenous serotonin stimulation, while only half of the subjects reported minor short-lived adverse effects following intraluminal serotonin stimulations. We conclude that exogenous serotonin in the lumen of the upper part of the small intestine does not seem to change antro-duodeno-jejunal contractility significantly in healthy adult volunteers.     

Synthesis and characterization of photolabile derivatives of serotonin for chemical kinetic investigations of the serotonin 5-HT(3) receptor

A series of photolabile o-nitrobenzyl derivatives of serotonin (caged serotonin) were synthesized: the amine-linked serotonin derivatives N-(2-nitrobenzyl) serotonin (Bz-5HT) and N-(alpha-carboxy-2-nitrobenzyl) serotonin (N-CNB-5HT), and O-alpha-carboxy-2-nitrobenzyl) serotonin (O-CNB-5HT), which has the caging group attached to the phenolic OH group. All the derivatives released free serotonin when excited by 308-nm or 337-nm laser pulses. The time constant of serotonin release from N-CNB-5HT was 1. 2 ms, with a quantum yield of 0.08. This is too slow for rapid chemical kinetic measurements. O-CNB-5HT is suitable for transient kinetic investigations of the serotonin 5-HT(3) receptor. It released serotonin with a time constant of 16 micros and a quantum yield of 0.03. The biological properties of O-CNB-5HT were evaluated, and the applicability of the compound for kinetic studies of the 5-HT(3) receptor was demonstrated. O-CNB-5HT does not activate the 5-HT(3) receptor by itself, nor does it modulate the response of a cell when co-applied with serotonin. When irradiated with a 337-nm laser pulse, O-CNB-5HT released free serotonin that evoked 5-HT(3) receptor-mediated whole-cell currents in NIE-115 mouse neuroblastoma cells.     

Inhibition of serotonin reuptake.

An uptake system on the serotonin neuronal membrane apparently functions to inactivate serotonin that has been released into the synaptic cleft. Various inhibitors of this active transport system on serotonin neurons are known, and some are specific in the sense that they do not inhibit the active uptake system on norepinephrine neurons. The most widely studied specific inhibitor of the serotonin neuron pump is fluoxetine, 3-(p-trifluoromethylphenoxy-N-methyl-3-phenyl propylamine (Lilly 110140). When fluoxetine or other effective but less specific serotonin uptake inhibitors are given, a rapid decrease in serotonin turnover occurs and the rate of firing of single neural units in the serotonin rich raphe area of brain is reduced. This decrease in serotonin turnover and release may be a compensatroy mechanism in response to an enhanced action of serotonin on synaptic receptors. Through the use of fluoxetine and other serotonin uptake inhibitors, the role of serotonin neurons in various brain functions--behavior, sleep, regulation of pituitary hormone release, thermoregulation, pain responsiveness, and so on--can be studied.     

Alteration of the acetylcholine response by intra- and extracellular serotonin application in intracellularly perfused neurons ofLymnaea stagnalis

1. The effect of serotonin on the acetylcholine (ACh) response has been studied by means of voltage clamp and intracellular perfusion in unidentified isolated neurons from parietal and visceral ganglia of Lymnaea stagnalis. 2. In most cells studied serotonin added to the internal or external solution decreases the response to ACh. 3. In other neurons serotonin added to the intracellular solution increases the response to ACh; when it is added extracellularly it produces the opposite effect on the same cells. 4. The decreasing effect of serotonin on ACh currents is mimicked by cyproheptadine, an antagonist of serotonin receptors, and by the intracellular application of cyclic AMP (cAMP) forskolin. 5. The enhancing effect of intracellularly applied serotonin on ACh currents is blocked by cyproheptadine and is not obtained by the intracellular administration of cAMP and forskolin. In some cells the enhancing effect of serotonin appears after forskolin. 6. The results suggest a modulating effect of serotonin on cholinergic synaptic transmission in the nervous system of mollusks. The possible existence of intracellular serotonin receptors is discussed.     

What do we know about serotonin?

The present review focuses on what is known of basic serotonin physiology in the human body. Here, we describe serotonin biochemistry and metabolism and summarize the results of studies that have contributed significantly to our understanding of serotonin physiology. We report the well-established role of serotonin in cardiovascular, gastrointestinal, and circulatory physiology. Emphasis is placed on the role of serotonin in peripheral physiological systems rather than in the central nervous system. A brief overview is provided on the emerging role of serotonin in novel areas such as bone pathways and glucose uptake. We also report a select few animal studies and animal models that have provided worthwhile contributions to the understanding of serotonin in human physiology. In addition, we summarize the results of large-scale genetic studies on serotonin and serotonin transporter genes, performed in relation to behavioral and mood disorders.     

Biochemistry and ultrastructure of serotonergic nerve endings in the lobster: serotonin and octopamine are contained in different nerve endings

In this article we report that the distribution of serotonin in the lobster nervous system parallels the distribution of octopamine and that the same tissues that contain endogenous serotonin can synthesize it from tryptophan. Octopamine and serotonin are highly concentrated in a neurosecretory region of the second thoracic roots in association with a group of neurosecretory cells. The roots possess separate high-affinity uptake systems for both serotonin and tryptophan. Radioactive serotonin, accumulated in tissues during incubations with either tritiated serotonin or tritiated tryptophan, can be released, in a calcium-dependent manner, by depolarization with potassium. A detailed morphological examination of the second thoracic roots shows four distinct categories of nerve endings in the vicinity of the neurosecretory cells. Octopamine is synthesized in one of these types of endings and serotonin in another. The high-affinity uptake systems for serotonin and tryptophan are found only in association with the endings that make serotonin. These endings and all the biochemical parameters of serotonin metabolism in the roots are selectively destroyed by previous injection of animals with the neurotoxin 5,7-dihydroxytryptamine.     

3,4-methylenedioxymethamphetamine (MDMA)-induced release of endogenous serotonin from the rat dorsal raphe nucleus in vitro: Effects of fluoxetine and tryptophan

The serotonin releasing action of 3,4-methylenedioxymethamphetamine on slices of dorsal raphe nucleus from rat was investigated. The slices were maintained in a gas-liquid interface perfusion chamber used for electrophysiological recording. Microdialysis probes designed for use on the slice surface were employed to measure the release of endogenous serotonin which was determined using liquid chromatography with electrochemical detection. Three minute duration exposure of the slices to 100 micromolar 3,4-methylenedioxymethamphetamine caused a long lasting release of endogenous serotonin. Fluoxetine, a serotonin transport inhibitor, reduced the amount of serotonin release. Tryptophan added to the perfusion solution increased both the duration and amount of serotonin released. These results further support earlier work on the mechanism of 3,4-methylenedioxymethamphetamine induced inhibition of serotonin neuronal firing.     

Transport and storage of serotonin by thrombin-treated platelets

Repeated thrombin treatment of washed platelets prepared from rabbits can decrease the serotonin content of the platelets by about 80%. When these platelets are deaggregated they reaccumulate serotonin but their storage capacity for serotonin is reduced by about 60%. If thrombin-pretreated platelets are allowed to equilibrate with a high concentration of serotonin (123 mu M), they release a smaller percentage of their total serotonin upon further thrombin treatment, in comparison with the percentage of serotonin released from control platelets equilibrated with the same concentration of serotonin calculations indicate that in thrombin-treated platelets reequilibrated with serotonin, two-thirds of the serotonin is in the granule compartment and one-third is in the extragranular compartment, presumably the cytoplasm. Analysis of the exchange of serotonin between the suspending fluid and the platelets showed that thrombin treatment does not alter the transport rate of serotonin across the platelet membrane and does not cause increased diffusion of serotonin from the platelets into the suspending fluid. The primary reason for the reduced serotonin accumulation by the thrombin-treated platelets appears to be loss of amine storage granules or of the storage capacity within the granules.     

Die Bildung von Serotonin in Juglans regia L.

Summary The serotonin content of growing fruits and of germinating seeds of Juglans regia has been studied. In the embryo 0.4–0.6 mg serotonin/g FW were found; in contrast no serotonin was detectable in the fleshy pericarp and in the seed coat. Serotonin was also not detectable in leaves, stems and roots of the adult plant. Most of the serotonin found in the embryo is formed after abscission of the seeds. During the synthesis of serotonin there are no dramatic changes in the chemical composition of the seeds (Tables 3–5).The formation of serotonin could be followed in isolated cotyledons and under sterile conditions. This serotonin formation is stimulated by exogenous tryptophan (Fig. 2). That tryptophan acts as a precursor of serotonin could be demonstrated with labelled DL-tryptophan (benzene ring ¹⁴C) (U). The possibility of stimulating serotonin formation in isolated cotyledons by the addition of tryptophan is limited to a certain stage of development and cannot be observed with material from fully matured seeds (Fig. 3).No serotonin was found in callus tissue and adventitious roots formed by isolated cotyledons; all the serotonin remained in the cotyledons. This was also the case in young seedlings, in which only the cotyledons showed the characteristic high serotonin content, whereas leaves, stems and roots contained no serotonin (Table 6).From these data we conclude that serotonin formation in the embryo of Juglans regia is not a special type of nitrogen storage but a way of ammonia detoxification in which ammonia from protein amino acid degradation is incorporated into serotonin via tryptophan.     

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.     

Serotonin: an inducer of collagenase in myometrial smooth muscle cells.

Rat myometrial smooth muscle cells in culture actively produce collagenase in medium containing fetal bovine serum, but not in medium containing newborn bovine serum or containing fetal serum adsorbed with dextran-coated charcoal. A dialyzable molecule has been isolated from fetal bovine serum, which restores the ability of the smooth muscle cells to produce collagenase. The molecule has been purified and identified as serotonin (5-hydroxytryptamine). Cells cultured in medium depleted of serotonin for 3 days fail to produce collagenase, as assessed both enzymatically and immunologically. Addition of serotonin promptly restores the ability of the cells to produce the enzyme. The EC50 for serotonin is approximately 2 microM; maximum stimulation of collagenase production is observed at 5 microM. The response is specific for serotonin: a wide variety of compounds tested, either related to serotonin or of potential reproductive significance, were without effect in the induction of collagenase production by the cells. No changes in DNA content, general protein synthesis, or cellular collagen production were observed as a consequence of serotonin depletion or restoration, suggesting a selective effect of the compound on collagenase production. The effect of serotonin was also selective to myometrial smooth muscle cells; collagenase-producing fibroblasts from skin and cervix displayed no serotonin requirement for enzyme production. Studies using specific agonists or antagonists for a variety of serotonin receptor subtypes suggest that the 5-HT-2 receptor mediates the serotonin induction of collagenase in these cells. Preliminary evidence indicates that cultured human myometrial smooth muscle cells are also dependent upon serotonin for collagenase production. The evidence in this study suggests the possibility that serotonin serves as a signal to begin the massive collagen degradation that occurs in the postpartum uterus.     

NG108-15 hybrid cells take up but do not synthesize serotonin

The mouse neuroblastoma × rat glioma hybrid cell line, NG108-15, does not synthesize serotonin from tryptophan, although the cells take up tryptophan and serotonin in a saturable manner from serum-supplemented incubation medium. Since serum commonly used to supplement the growth medium contains serotonin, it is concluded that appreciable levels of serotonin found in NG108-15 cells are attributable to uptake of serotonin from the serum-supplemented medium.     

Effects of 5,6-Dihydroxytryptamine on the Release, Synthesis, and Storage of Serotonin: Studies Using Rat Brain Synaptosomes

5,6-Dihydroxytryptamine is a neurotoxic analogue of serotonin which can have profound cardiovascular effects within minutes of administration in vivo (Korner and Head, 1981). These effects have been attributed to 5,6-dihydroxytryptamine-induced serotonin release, although there has been no biochemical assessment of the extent to which this occurs. The present study utilized an in vitro synaptosomal assay to determine the short-term effects of 5,6-dihydroxytryptamine on endogenous serotonin release, synthesis, storage, and metabolism. 5,6-Dihydroxytryptamine produced a rapid depletion of serotonin. At lower concentrations of 5,6-dihydroxytryptamine (0.1-1 microM), this depletion was associated primarily with an increase in the levels of 5-hydroxyindoleacetic acid, the deaminated metabolite of serotonin, with small increases in the amount of serotonin release. At higher concentrations (10-100 microM), a greater proportion of the depleted serotonin was released with less metabolism occurring. When metabolism was prevented by inhibiting monoamine oxidase, the amount of serotonin which was released equalled the amount of serotonin depletion. Thus monoamine oxidase activity was important in controlling the amount of serotonin which could be released by 5,6-dihydroxytryptamine. Further studies demonstrated that an impairment in serotonin synthesis and vesicular storage could account for the rapid depletion produced by 5,6-dihydroxytryptamine. Taken together, the results indicate that 5,6-dihydroxytryptamine acts to displace serotonin from vesicular stores into the cytoplasm where it can either be deaminated by monoamine oxidase or be released. Moreover, it is hypothesized that the intraneuronal concentration of 5,6-dihydroxytryptamine is important in determining the extent of serotonin release, because it can inhibit the deamination of serotonin by monoamine oxidase.     

Serotonin localization and its functional significance during mouse preimplantation embryo development

Serotonin is a neurotransmitter functioning also as a hormone and growth factor. To further investigate the biological role of serotonin during embryo development, we analysed serotonin localization as well as the expression of specific serotonin 5-HT1D receptor mRNA in mouse oocytes and preimplantation embryos. The functional significance of serotonin during the preimplantation period was examined by studying the effects of serotonin on mouse embryo development. Embryo exposure to serotonin (1 microM) highly significantly reduced the mean cell number, whereas lower concentrations of serotonin (0.1 microM and 0.01 microM) had no significant effects on embryo cell numbers. In all serotonin-treated groups a significant increase in the number of embryos with apoptotic and secondary necrotic nuclei was observed. Expression of serotonin 5-HT1D receptor mRNA in mouse oocytes and preimplantation embryos was confirmed by in situ hybridization showing a clearly distinct punctate signal. Immunocytochemistry results revealed the localization of serotonin in oocytes and embryos to the blastocyst stage as diffuse punctate cytoplasmic labelling. It appears that endogenous and/or exogenous serotonin in preimplantation embryos could be involved in complex autocrine/paracrine regulations of embryo development and embryo-maternal interactions.     

Effect of serotonin on the formation of neurons producing gonadotropin-releasing hormone in Wistar rats

The effect of serotonin on the formation of neurons producing gonadotropin-releasing hormone (GnRH) during embryogenesis of Wistar rats was studied. The neurons producing GnRH were detected immunocytochemically on days 18 and 21 of embryonic development and on day 15 of postnatal development of rats with normal serotonin metabolism and rats in which the synthesis of serotonin was inhibited by p-chlorophenylalanine. The total number of GnRH neurons in serotonin deficiency was larger than in the case of its normal metabolism at all developmental stages studied. This is an indirect evidence for the inhibitory effect of serotonin on the formation of GnRH neurons. To confirm the morphogenetic effect of serotonin, we studied the rate of formation of GnRH neurons by injecting bromodeoxyuridine in the formation period of these neurons. It was found that serotonin deficiency had no effect on the time of formation of GnRH neurons: over 97% of neurons formed on days 11 to 15 of embryonic development both in the experimental and control groups. Note that, in serotonin deficiency, the maximum number of GnRH neurons formed one day later than in the normal state. Thus, serotonin inhibits the proliferation of GnRH neuron progenitor cells and thereby has a morphogenetic effect on the development of these neurons.     

Regional distribution of 5-hydroxytryptamine in cat brain.

The regional distribution of serotonin (5-HT) in the brain of the cat is poorly understood. In this work, serotonin was analyzed fluorometrically along the brain stem and prosencephalon of the cat. The hypothalamus had the highest concentration of serotonin. Serotonin decreased gradually at the mesencephalon, preoptic area, medulla oblongata, hippocampus, pons, visual cortex, spinal cord and frontal cortex. Significant differences were found between the raphe (3 mm thick) and the lateral blocks of the brain stem. The concentration of serotonin is higher in the raphe blocks, though it decreases caudally. There is no significant difference between the raphe (4 mm thick) and the lateral block of the brain stem. The results demonstrate the regional concentration of serotonin in the CNS of a normal cat, the relationships between serotoninergic neuron groups and serotonin concentration,and the probable significance of nerve terminals and varicosities in storing serotonin.     

Ultrastructural immunocytochemical co-localization of serotonin and PNMT in adrenal medullary vesicles

Summary Previous immunocytochemical studies at the light microscopic level have demonstrated serotonin immunoreactivity in rat adrenal epinephrine-containing cells. In this study we have used electron microscopic immunocytochemical methods to study the subcellular distribution of serotonin and the enzyme responsible for epinephrine biosynthesis, phenylethanolamine-N-methyltransferase (PNMT). The distribution of the immunostaining was compared in adjacent serial thin sections using a post-embedding method in conjunction with peroxidase-antiperoxidase (PAP) immunocytochemistry. Serotonin immunoreactivity was associated with the limiting membrane as well as with the core of the chromaffin vesicles. In adjacent sections PNMT immunoreactivity was also seen in the serotonin-containing vesicles. However, its intravesicular distribution was different from that of serotonin; PNMT occupied the eccentric zone of the vesicles between the serotonin immunoreactive sites.These results are interpreted to be in support of biochemical studies claiming a serotonin uptake and storage capacity of adrenal chromaffin vesicle fractions as well as those which suggest serotonin is synthesized by chromaffin cells. The relative contribution of uptake and synthesis to the pool of serotonin that is stored in the vesicles is an open question. The co-localization of serotonin and PNMT in the same vesicle is suggestive of a capacity for co-release of serotonin and epinephrine by the adrenal medulla.     

Single- and multi-photon excited fluorescence from serotonin complexed with beta-cyclodextrin.

The fluorescence of serotonin on binding with beta-cyclodextrin has been studied using both steady state and time-resolved methods. Steady state fluorescence intensity of serotonin at 340 nm showed approximately 30% increase in intensity on binding with K(A) approximately 60 dm(3) mol(-1) and the fluorescence lifetimes showed a corresponding increase. In contrast, the characteristic green fluorescence ('hyperluminescence') of serotonin observed upon multiphoton near-infrared excitation with sub-picosecond pulses was resolved into two lifetime components assigned to free and bound serotonin. The results are of interest in relation to selective imaging and detection of serotonin using the unusual hyperluminescence emission and in respect to recent determinations of serotonin by capillary electrophoresis in the presence of cyclodextrin. The results also suggest that hyperluminescence occurs from multiphoton excitation of a single isolated serotonin molecule.