5,7-dihydroxytryptamine lesions of dorsal and median raphé nuclei and performance in the social interaction test of anxiety and in a home-cage aggression test.

Micro-injections of the neurotoxin 5,7-dihydroxytryptamine into the dorsal raphe nucleus produced a behavioural profile in the social interaction test of anxiety similar to that seen in rats treated chronically with benzodiazepines. Neurotoxin injections into the median raphé nucleus did not produce a profile significantly different from that of the controls. In the control rats and in the rats with lesions of the median raphé nucleus, ACTH1-24 (corticotrophin) significantly reduced active social interactions, whereas it was without effect on the rats with lesions of the dorsal raphé nucleus. In the home-cage intruder test, the median raphé-lesioned rats submitted less to the intruder and stood and jumped on him more often than did the controls. The dorsal raphé-lesioned rats showed significantly fewer interactions of all kinds, compared with control rats when an intruder was placed in their home cages.     

Selective accumulation of hydroxytryptamines by frog tectal neurones

By means of histofluorescence microscopy, 5,7-dihydroxytryptamine was shown to be taken up by selective populations of brain neurones of the frog,Rana pipiens, following both intracranial administration andin vitro incubation with isolated brain preparations. Presumptive non-aminergic cell bodies of the superficial aspect of tectal lamina 6 exhibited more avid uptake than did putative serotonin perikarya of the raphe complex. Within the tectum, 5,7-dihydroxytryptamine uptake appeared to be restricted to large piriform neurons; in the torus semicircularis, it occurred in a morphologically dissimilar group of scattered cells. The same tectal cell system accumulated 5-hydroxytryptamine and 6-hydroxytryptamine, but notN-acetylserotonin, melatonin, or noradrenaline. 5,7-Dihydroxytryptamine uptake was insensitive to cold or imipramine; however, it was blocked by ouabain at high but not low temperature. At concentrations ≥ 100 μM, 5,7-dihydroxytryptamine-induced fluorescence was sufficiently intense to permit tracing of intratectal dendrites and tectofugal axonal processes projecting to a lateral diencephalic neuropil and an ipsilateral isthmic neuropil.While previous monoamine histofluorescence and immunohistologic studies have not revealed serotonin-containing perikarya in the ranid tectum, our findings demonstrate that lamina 6 piriform projection neurones, presumably lacking indolamine-synthesizing enzymes, possess a striking capability for accumulating hydroxylated tryptamines.     

The effects of monoamine neurotoxins on peptides in the rat spinal cord

The coexistence of two neuronally-localised peptides, substance P and thyrotropin-releasing hormone (TRH), in descending serotoninergic nerve fibres to the spinal cord was investigated using immunocytochemical and biochemical methods. Substance P-like material in the spinal cord was shown to be identical to the undecapeptide substance P by the criteria of gel filtration, high performance liquid chromatography and behaviour in substance P specific radioimmunoassays. Immunocytochemical staining for 5-hydroxytryptamine, substance P, and TRH showed that all three substances had a similar distribution in nerve fibres and terminals in the ventral and lateral grey matter of the spinal cord. After treatment with the serotonin neurotoxin 5,7-dihydroxytryptamine, neuronal elements containing 5-hydroxytryptamine, substance P and TRH degenerated and disappeared from these parts of the spinal cord in parallel with one another.Biochemical measurements of 5-hydroxytryptamine, substance P and TRH in the spinal cord after treatment with 5,7-dihydroxytryptamine confirmed that these three substances were all depleted from the ventral horn and, in addition, showed that there was a small depletion of substance P from the dorsal horn. Two other neuropeptides, somatostatin and methionine-enkephalin were not depleted from the spinal cord by treatment with 5,7-dihydroxytryptamine nor was substance P in other parts of the brain. Substance P in the spinal cord was unaffected by 6-hydroxydopamine, a drug known to destroy catecholamine-containing neurones.These results are consistent with coexistence of substance P and TRH together with 5-hydroxytryptamine in the descending axons and terminals of bulbospinal neurones.     

Visualization of serotonin neurones in the nodose ganglia of the cat. An autoradiographic study

After in vitro incubation of the nodose ganglia (right and left) in tritiated serotonin mixed with norepinephrine, the autoradiographic detection of the amine at the light-microscopic level showed an intense labeling over cell bodies; they have been characterized as gray type neurones at the ultrastructural level. Moreover, a labeling was observed over unmyelinated intraganglionic fibers.The uptake of exogenous serotonin by these neurones was demonstrated to be specific by using different experimental conditions. The same distribution of labeling was observed either after the incubation in ‘high affinity’ concentration (10^?7M), or in saturated concentrations (10^?5M or 10^?4M) of [³H]serotonin. The number of labeled cell bodies decreased after adding fluoxetine (an inhibitor of serotonin uptake) to the tracer.The specificity of the neuronal accumulation of serotonin was confirmed by interganglionic comparisons between the nodose and the superior cervical ganglia incubated under the same conditions. Several hundred cell bodies were labeled in the nodose ganglion incubated in [³H]serotonin (10^?6M) whereas there was no labeling in the superior cervical ganglion. On the other hand, most of the perikarya were labeled in the superior cervical ganglion incubated in [³H] norepinephrine (10^?6M), whereas no significant labeling was found in the nodose ganglion.These results provide some evidence that peripheral sensory neurones are able to take up exogenous serotonin.     

Increased expression of preproneuropeptide Y and preprosomatostatin mRNA in striatum after selective serotoninergic lesions in rats

The levels of neuropeptide Y and somatostatin may change when serotoninergic neurotransmission is altered in different brain regions. To assess whether serotonin regulates the synthesis of these peptides, we measured the levels of preproneuropeptide Y (ppNPY) and preprosomatostatin (ppSOM) mRNA in different brain regions after intracerebroventricular injection of 5,7-dihydroxytryptamine (5,7-DHT), a selective serotonin neurotoxin. The mRNA of these peptides significantly increased in the striatum but not in hippocampus and frontal cortex. It thus appears that serotonin has an inhibitory effect on the biosynthesis of neuropeptide Y and somatostatin in striatum whereas it probably acts by stimulating the release of these peptides in hippocampus and frontal cortex.     

Interaction between Serotonin and Nitric Oxide (NO) in the Activation of the Serotoninergic System in the Common Snail

The effects of serotonin and NO donors on serotoninergic neurons (more than 60) in the brain of the common snail Helix lucorum were studied. Serotonin and NO donors induced depolarization in all neurons, and increased spike activity and activated the synchronous synaptic input, including train-like input, resulting in the onset of synchronous train activity in all these neurons. The excitatory effect of serotonin was significantly decreased by 5,7-dihydroxytryptamine (5,7-DOT) and monomethylarginine – a blocker of endogenous NO synthesis. Both these substances blocked the serotonin activation of the synchronous train input. 5,7-DOT also blocked the activation of this input by NO donors, but had no effect on their excitatory actions. The effects of 5,7-DOT developed quickly, were reversible, and were comparable to the effects of serotonin receptor antagonists. The data obtained here provide evidence that serotonin and NO have similar regulatory effects on the serotoninergic system in the snail brain. Not only do they excite serotoninergic neurons, but they also coordinate their functioning by activating common synaptic inputs, which are apparently also serotoninergic. It is suggested that NO has the role of a second messenger during serotoninergic excitation and functions as a co-transmitter for the presynaptic input.     

Denervation supersensitivity to serotonin in the facial nucleus.

The present investigation was designed to determine whether denervation supersensitivity to serotonin could be produced in the facial nucleus of the rat. The mechanism for the development of this phenomenon was also explored. Serotonergic denervation was accomplished chemically by injection of 200μg of 5,7-dihydroxytryptamine into the lateral ventricle. The response of facial motoneurons to microiontophoretic application of 5-HT, 5-methoxydimethyltryptamine and norepinephrine was determined by extracellular unit recording. There was a marked increase in the sensitivity to serotonin 1 day following lesions. Histochemical fluorescence indicated that serotonin-containing terminals were destroyed at this time. In contrast, supersensitivity to 5-methoxydimethyltryptamine took 7 days to develop following lesions. The serotonin uptake inhibitor, fluoxetine, potentiated the response of motoneurons to serotonin in control animals but not in lesioned rats. The supersensitivity to serotonin and 5-methoxydimethyltryptamine was attenuated in animals 14 days after the lesions. The diminution of the effect of serotonin and 5-methoxydimethyltryptamine was temporally correlated with regeneration of serotonin-containing fibers and terminals in the facial nucleus.The functional importance of the serotonin input to the facial nucleus and possible pre- and postsynaptic mechanisms involved in the development of supersensitivity to serotonin are discussed.     

Differential effects of 5,7-dihydroxytryptamine-induced serotoninergic degeneration on 5-HT1A receptors and 5-HT uptake sites in the rat brain

The time-course of 5,7-dihydroxytryptamine-induced lesions (2, 5 and 14 days after i.c.v. injection of 150 μg) and the effects of acute reserpine treatment (10 mg/kg, i.p., one or 5 days before scheduled death), were evaluated by autoradiography of [³H]paroxetine binding sites in the rat brain. Reserpine had no significant effect on [³H]paroxetine binding, indicating that the depletion of serotonin is not sufficient per se to alter the serotonin uptake sites in any region. Two days after the 5,7-dihydroxytryptamine lesion, [³H]paroxetine binding was already decreased in the majority of brain regions. In the caudate putamen these binding sites were significantly decreased only 14 days after the lesion, whereas the ventral tegmental area (or the enclosed median forebrain bundle), the dorsal raphe (mainly the ventral portion) and the median raphe maintained their high density of serotonin uptake sites even after 14 days. Results were similar using [³H]citalopram as ligand for the serotonin uptake sites, in the brains of rats lesioned 5 days before death; an exception was the ventral portion of the dorsal raphe, where there was a significant increase with [³H]paroxetine and a decrease with [³H]citalopram binding. In adjacent sections of the same brains we also measured [³H]8-OH-DPAT binding, confirming that it completely disappears in the dorsal raphe after the lesion. Thus, considering the extent of serotonin cell body degeneration, there appears to be a paradoxical mismatch between the excessive loss of [³H]8-OH-DPAT binding and the resistance of [³H]citalopram or [³H]paroxetine binding in the dorsal raphe, suggesting that the two binding sites may undergo adaptive regulation in surviving neurons.     

Brain serotonin depletion impairs short-term memory, but not long-term memory in rats

Intracerebroventricular injection of 5,7-dihydroxytryptamine (5,7-DHT) (150 microg; 4.5 microl/ventricle), a serotonergic neurotoxin, significantly decreased spontaneous alternation in Y-maze task and working memory in radial 8 arm-maze task, suggesting effects on short-term memory, without affecting long-term memory, explored by reference memory in radial 8 arm-maze task and step-through latency in multi-trial passive avoidance task. Parachlorophenylalanine (PCPA) (3 days treatment 200 microg, i.c.v.), a serotonin synthesis inhibitor, did not impair step-through-latency in multi-trial passive avoidance task, suggesting no effects on long-term memory. These results suggest that serotonin, among other neurotransmitters, play an important role in cognitive functions, especially short-term memory.     

Effects of amyloid-β peptides on the serotoninergic 5-HT1A receptors in the rat hippocampus

A recent [¹⁸F]MPPF-positron emission tomography study has highlighted an overexpression of 5-HT_1A receptors in the hippocampus of patients with mild cognitive impairment compared to a decrease in those with Alzheimer's disease (AD) [Truchot, L., Costes, S.N., Zimmer, L., Laurent, B., Le Bars, D., Thomas-Antérion, C., Croisile, B., Mercier, B., Hermier, M., Vighetto, A., Krolak-Salmon, P., 2007. Up-regulation of hippocampal serotonin metabolism in mild cognitive impairment. Neurology 69 (10), 1012-1017]. We used in vivo and in vitro neuroimaging to evaluate the longitudinal effects of injecting amyloid-β (Aβ) peptides (1-40) into the dorsal hippocampus of rats. In vivo microPET imaging showed no significant change in [¹⁸F]MPPF binding in the dorsal hippocampus over time, perhaps due to spatial resolution. However, in vitro autoradiography with [¹⁸F]MPPF (which is antagonist) displayed a transient increase in 5-HT_1A receptor density 7 days after Aβ injection, whereas [¹⁸F]F15599 (a radiolabelled 5-HT_1A agonist) binding was unchanged suggesting that the overexpressed 5-HT_1A receptors were in a non-functional state. Complementary histology revealed a loss of glutamatergic neurons and an intense astroglial reaction at the injection site. Although a neurogenesis process cannot be excluded, we propose that Aβ injection leads to a transient astroglial overexpression of 5-HT_1A receptors in compensation for the local neuronal loss. Exploration of the functional consequences of these serotoninergic modifications during the neurodegenerative process may have an impact on therapeutics targeting 5-HT_1A receptors in AD.     

Loss of vision-guided adaptation of the vestibulo-ocular reflex after depletion of brain serotonin in the rabbit

Brain serotonin in pigmented rabbits was depleted by intraventricular injection of 5,7-dihydroxytryptamine (5,7-DHT). Before depletion, an adaptive increase in the gain of the horizontal vestibulo-ocular reflex (HVOR) could be induced regularly by continuously rotating the animals in combination with optokinetic stimulus. After the depletion, such an increase in HVOR gain did not occur, even though dynamic characteristics of the HVOR and of the optokinetic eye movement were not altered. 5,7-DHT treatment also reduced brain norepinephrine, but depletion of norepinephrine to a similar extent by intraventricular injection of 6-hydroxydopamine in other rabbits did not affect the HVOR adaptation. These results suggest that brain serotonin plays an important role in maintaining adaptive modifiability of the HVOR.     

Interaction of serotonin- and dopaminergic systems of the brain in mechanisms of latent inhibition in rats

The activity of serotoninergic neurons of the medial raphé nucleus was turned off by local injection of the neurotoxin, 5,7-dihydroxytryptamine. Seven days later, a conditioned passive avoidance reaction was developed following 20 preexposures to the conditional stimulus (the presentation of the experimental chamber). As compared with the sham-operated control, in which 20 preexposures to the stimulus elicited latent inhibition, i. e., attenuation of the reproduction of the conditioned passive avoidance reaction, prolonged maintenance at the formed level, and lack of susceptibility to amnesia, all of the enumerated features were disturbed in animals administered the neurotoxin. The administration of haloperidol (0.5 mg/kg) 1 h prior to training reestablished the state of latent inhibition which had been disrupted by the switching off of the mesolimbic serotonin system. It is hypothesized that the process of the suppression of attention to the nonreinforced signals during the preexposure to the stimulus is achieved by the interaction of the serotoninergic and dopaminergic systems.Translated from Zhurnal Vysshei Nervnoi Deyatel'nosti imeni I. P. Pavlova, Vol. 39, No. 4, pp. 714–720, July–August, 1990.     

Ultrastructural organization of regenerated serotonin axons in the dorsomedial hypothalamus of the adult rat

Summary The ultrastructural organization of regenerated serotonin (5-HT) axons was examined in the dorsomedial hypothalamus (DMH) of the adult rat using high-resolution radioautography after intraventricular infusion of [³H]5-HT. An analysis of the microenvironment of the [³H]5-HT-labelled terminals in the DMH was made 30 and 50 days after unilateral injection of 5,7-dihydroxytryptamine (5,7-DHT) or vehicle solution into the dorsolateral hypothalamus. In sham-treated animals [³H]5-HT-labelled axons were small, contained many small clear vesicles, one or more large granular vesicles, and showed only rare synaptic specializations. In 5,7-DHT-treated animals the internal organization of [³H]5-HT-labelled profiles resembled that of sham-treated animals. A tendency toward increased synaptic frequency was found for [³H]5-HT-labelled terminals in the 5,7-DHT-treated group 50 days post-lesion, and an increase in the number of [³H]5-HT-labelled terminals abutting unlabelled perikarya was found in both 30- and 50-day post-lesion groups as compared to sham-treated groups. No other differences in ultrastructural environment were found between sham- and 5,7-DHT-treated animals at either 30 or 50 days post-lesion. These results suggest that 5-HT fibres in the hypothalamus regenerate with a great deal of cellular specificity.     

Role of enkephalinergic brain structures in regulating motivated behavior in rats with lesioned serotoninergic neurons

The behavioral effects of injections of enkephalin into the substantia nigra or dorsal raphe nucleus were studied in rats with lesioned serotoninergic structures. Lesions were produced by intracerebral administration of the neurotoxin 5,7-dihydroxytryptamine. Treated rats showed normalization of conditioned drinking reflex extinction. It is proposed that there is a tight connection between the normalizing effects of enkephalin on rat behavior in conditions of deficient brain serotoninergic system function and increases in the efficiency of presynaptic inhibition of dopaminergic neurons. Laboratory for Conditioned Reflex Morphophysiology, Science Research Institute of the Brain, Russian Academy of Medical Sciences, 5 Obukh Lane, 103064 Moscow, Russia. Translated from Rossiiskii Fiziologicheskii Zhurnal imeni I. M. Sechenova, Vol. 83, No. 1-2, pp. 71–75, January–February, 1997.     

Indirect effects of apomorphine on serotoninergic neurons in rats

We have studied the effects of apomorphine on central serotoninergic system by using fluorescence histochemistry and high performance liquid chromatography coupled with electrochemical detection. Apomorphine has been shown selectively to elevate intracellular serotonin fluorescence in dorsal raphe neurons and serotonin and/or 5-hydroxyindoleacetic acid concentrations in the major terminal area of the dorsal raphe, the striatum. Apomorphine has no effect on serotonin neurons in the median raphe or its corresponding projection site, the hippocampus. In the present study, intraventricular 6-hydroxydopamine, when administered together with desimipramine and pargyline, antagonized the apomorphine-induced elevations of serotonin fluorescence in the dorsal raphe and serotonin and 5-hydroxyindoleacetic acid levels in the striatum. This antagonism was found when 6-hydroxydopamine was given either 3 or 10 days before 3 mg/kg apomorphine. Apomorphine also elevated extraperikaryal serotonin fluorescence and catecholamine fluorescence in the dorsal raphe selectively and these effects were also blocked by 6-hydroxydopamine. Additionally, 6-hydroxydopamine accelerated striatal serotonin turnover when it was given 10 days prior to death. This phenomenon probably reflects some compensatory change of mesostriatal serotonin neurons in response to the prolonged depletion of brain dopamine. The above results suggest that the observed effects of apomorphine on the serotoninergic system are mediated indirectly through dopaminergic neurons and that postsynaptic dopamine receptors are probably not responsible for apomorphine's effects.     

5,7-dihydroxytryptamine lesions of the dorsal and median raphe nuclei interfere with lithium-induced conditioned gaping, but not conditioned taste avoidance, in rats

Both the dorsal and median raphe nuclei of the midline brainstem region in rats were lesioned with the neurotoxin 5,7-dihydroxytryptamine. Rats were then surgically implanted with intraoral cannulas for fluid delivery and received a single conditioning trial in which 2-min saccharin infusion was followed by either lithium or saline administration. The conditioned gaping seen in the lithium-conditioned rats was significantly attenuated by raphe lesions, indicating that reduction of forebrain serotonin levels interferes with conditioned gaping. However, lesioned rats still expressed comparable conditioned taste avoidance as measured by both the 1- and 2-bottle consumption tests. These results parallel previous pharmacological findings indicating that reduction of serotonin activity interferes with conditioned gaping, but not conditioned taste avoidance.     

Co-localization of serotonin and GABA in neurons of the Xenopus laevis retina

Serotonin-synthesizing neurons in the retina of Xenopus laevis have been identified using anti-phenylalanine hydroxylase (PH) antibody which recognizes tryptophan 5-hydroxylase, the rate-limiting enzyme for serotonin synthesis. Double-labelling experiments, using anti-PH antibody and anti-serotonin antibody/5,7-dihydroxytryptamine (5,7-DHT) uptake, have shown that some serotonin-like immunoreactive/5,7-DHT-labelled neurons exhibit PH-like immunoreactivity (PH-LI) (serotonin-synthesizing neurons), but the others do not (serotonin-accumulating neurons). In the present study, triple-labelling experiments were performed using 5,7-DHT uptake and antibodies raised against GABA and PH, to determine the possible co-localization of γ-aminobutyric acid (GABA) in serotonin-synthesizing and/or -accumulating neurons in the Xenopus retina. All 5,7-DHT-labelled bipolar cells lacked PH-LI; all of them were immunoreactive to GABA. In contrast, all 5,7-DHT-labelled large amacrine cells exhibited PH-LI, but none of them expressed GABA-LI. Small amacrine cells labelled with 5,7-DHT but not PH-LI exhibited GABA-LI, whilst the small amacrine cells with PH-LI lacked GABA-LI. These observations indicate that GABA is co-localized in serotonin-accumulating amacrine and bipolar cells, whereas serotonin-synthesizing large and small amacrine cells do not contain GABA-LI.     

Differential recovery of adrenocortical responses to neural stimuli following administration of 5,7-dihydroxytryptamine into the hypothalamus

Summary In view of the role of serotonin in adrenocortical regulation, the effects of depletion of hypothalamic serotonin, using localized injections of the neurotoxin 5,7-dihydroxytryptamine into the hypothalamic paraventricular nucleus, on the rise in plasma corticosterone following afferent neural stimulation, were studied. The neurotoxin caused a significant reduction (p<0.001) in hypothalamic serotonin content of about 50% during the first month and about 30% up to two months later. Basal and ether stress-induced rises in plasma corticosterone levels were unaffected at all times after this treatment, but responses to stimulation of the sciatic nerve were reduced for up to four weeks (p<0.01), recovering at later times. Responses to photic and acoustic stimuli were almost entirely prevented up to four weeks following the treatment (p<0.001) but showed a gradual recovery to full, or almost full, adrenocortical responses at eight weeks, following acoustic and photic stimulation respectively. These results demonstrate a differential recovery of the adrenocortical responses, following the neurotoxin injection and indicate that different neural modalities require different 5-HT concentrations in the PVN for the expression of a full adrenocortical response.     

Serotonin uptake by human platelets as a method of screening antidepressants

Antidepressants of the imipramine group considerably inhibit serotonin uptake by human platelets in concentrations of 10^–7 to 10^–5 M. Of the neuroleptics, only haloperidol (10^–5 M) significantly inhibits serotonin uptake by platelets. Amphetamine and cholinolytics do not affect serotonin uptake by platelets even in a concentration of 10^–4 M. The specificity of the effect of the antidepressants confirms the view that their thymoanaleptic action is an indirect effect brought about through activation of serotoninergic processes.Laboratory of Psychopharmacology, V. M. Bekhterev Leningrad Psychoneurological Research Institute. (Presented by Academician of the Academy of Medical Sciences of the USSR V. V. Zakusov.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 81, No. 1, pp. 51–53, January, 1976.