Effects of 5-HT and 8-OH-DPAT on forebrain monoamine synthesis after local application into the median and dorsal raphe nuclei of the rat

Summary 5-HT (10 and 40 g) and 8-OH-DPAT (1 and 5 g) were locally applied into the dorsal or median raphe nuclei in awake, unrestrained, rats. All animals were also treated with the 5-HTP and DOPA decarboxylase inhibitor NSD-1015, 100mg kg^–1 SC, 30 min before decapitation. 5-HT or 8-OH-DPAT were administered 5 min before NSD-1015. The regional brain in vivo rate of tyrosine and tryptophan hydroxylase activity was estimated by measuring the accumulation of DOPA and 5-HTP. The following brain regions were sampled: neocortex, hippocampus, dorso-lateral neostriatum, ventro-medial neostriatum, nucleus accumbens, olfactory tubercle, globus pallidus, septum and the amygdala.Compared to normal controls, there were small and inconsistent effects on forebrain 5-HTP accumulation by saline injections into the dorsal or the median raphe (an increase in 3 out of 36 experiments), whereas strong effects by the injection procedure were noted on forebrain DOPA accumulation (an increase in 15 out of 36 experiments).Injections of 5-HT (same effect by 10 or 40 g) into the dorsal raphe, produced a decrease in 5-HTP accumulation in all forebrain areas except for the hippocampus and the septum, whereas no effects were seen in any area after median raphe injections. In contrast, 8-OH-DPAT preferentially produced a decrease in forebrain 5-HTP accumulation after median raphe injections and less, but statistically significant effects by dorsal raphe injections. The 8-OH-DPAT injection into the median raphe primarily affected limbic forebrain areas (hippocampus, nucleus accumbens, ventro-medial neostriatum, amygdala and the septum).This dissociation of the effects of 5-HT and 8-OH-DPAT on forebrain 5-HT synthesis after local application into the dorsal or the median raphe strongly supports the contention of heterogeniety in the brain 5-HT receptor population in terms of receptor subtypes and/or receptor regulation.     

Anesthetics block morphine-induced increases in serotonin release in rat CNS

The effect of morphine on serotonin (5-HT) was examined by microdialysis in unanesthetized and anesthetized rats. In unanesthetized rats, morphine (10 mg/kg, s. c.) produced increases in extracellular 5-HT in nucleus accumbens (n. accumbens) and dorsal raphe nucleus (DRN), but not in the dorsal hippocampus. Similarly, extracellular 5-HT in the n. accumbens, but not the dorsal hippocampus, was increased after morphine (1 μM) was infused for 60 min by reverse dialysis into the DRN. Chloral hydrate, pentobarbital, and ketamine anesthesia had different effects on 5-HT in the n. accumbens. Chloral hydrate induced a transient increase and ketamine a sustained increase in extracellular 5-HT. Pentobarbital caused a sustained decrease. The effects of systemic and intraraphe administration of morphine were abolished by all three anesthetics. Infusion of muscimol, a GABA_A receptor agonist, into the DRN also induced a decrease in 5-HT and abolished the effects of morphine on 5-HT in the DRN and n. accumbens. These results are consistent with other evidence suggesting that morphineinduced increases in monoamine neurotransmission are a disinhibitory effect resulting from opioid-mediated inhibition of GABA release. More conclusively, it is apparent that anesthetized animals are inappropriate for testing the effect of morphine on 5-HT neurotransmission. © 1994 Wiley-Liss, Inc.     

Functional dissociation between serotonergic pathways in dorsal and ventral hippocampus in psychotomimetic drug-induced locomotor hyperactivity and prepulse inhibition in rats

Altered hippocampal function and brain serotonin activity are implicated in the development and symptoms of schizophrenia. We have previously shown that lesions of the median raphe nucleus, but not the dorsal raphe nucleus, produced a marked enhancement of locomotor hyperactivity induced by phencyclidine and disruption of prepulse inhibition. The dorsal and ventral hippocampus receive serotonin projections predominantly from the median raphe nucleus and dorsal raphe nucleus, respectively. Therefore, we investigated the effect of local lesions of serotonin projections into the dorsal and ventral hippocampus on psychotomimetic drug-induced locomotor hyperactivity and prepulse inhibition. Male Sprague-Dawley rats were anaesthetized with pentobarbitone and stereotaxically microinjected with 5 microg of the serotonergic neurotoxin 5,7-dihydroxytryptamine into either the dorsal or the ventral hippocampus. Two weeks after surgery, dorsal hippocampus-lesioned rats showed a 100% enhancement of the locomotor hyperactivity caused by phencyclidine treatment and a slight but significant reduction of the effect of amphetamine. Prepulse inhibition was significantly disrupted in lesioned rats and serotonin levels in the dorsal hippocampus were reduced by 80%. Rats with lesions of the ventral hippocampus showed 85% depletion of serotonin and partial disruption of prepulse inhibition, but no significant changes in the effect of phencyclidine or amphetamine. These results suggest that serotonin projections from the median raphe nucleus to the dorsal hippocampus play an important role in locomotor hyperactivity and prepulse inhibition in rats, animal models of aspects of schizophrenia. This suggests that these serotonin projections may be involved in the pathophysiology of schizophrenia symptomology.     

Stress induces rapid changes in serotonergic activity: restraint and exertion

Rapid activation of central serotonergic systems occurs in response to the social stress of aggression in dominant lizards. The most rapid expression of serotonergic activity occurs in nucleus accumbens, hippocampus and brainstem. To compare previously measured responses induced by social stressors with those provoked by physical stress, serotonergic activity was examined following restraint stress (handling) and forced physical exertion. After handling, some male Anolis carolinensis were placed on a race track and either run until there was no movement following 1 min of prodding, or half that time. Controls were killed without treatment. Lizards stressed by handling showed rapid (25 s) increases in serotonergic activity (5-HIAA/5-HT) in striatum, dorsal cortex, locus ceruleus, and nucleus accumbens. Other changes in serotonergic systems caused by stress occurred in raphe and hippocampus. Serotonergic changes induced by handling stress were reversed by exercise (to 50% maximal exertion time) in subiculum, striatum and nucleus accumbens. The serotonergic profile of lizards run until they would no longer respond to prodding (maximal exertion time) was significantly different from that for more acute exertion in hippocampus, subiculum, striatum, medial amygdala, locus ceruleus, area postrema, and raphe. Physical stress (handling) mimicked social stress by producing rapid serotonergic changes in hippocampus, subiculum, nucleus accumbens and locus ceruleus. In contrast, the medial amygdala, which has previously been demonstrated to respond serotonergically to social stress only after a temporal delay, did not show a rapid response to restraint stress.     

Effects of tactile startle on serotonin content of midbrain raphe neurons in rats

Rats selected to exhibit comparable levels of behavioral reactivity to tactile startle stimuli were used to examine the effects of the startle testing itself on the levels of serotonin (5-HT) in the dorsal and median raphe nuclei and in the hippocampus. A series of 240 air puff stimuli given in 1 h immediately before sacrifice decreased the intracellular 5-HT content, as measured by microspectro-fluorimetry, in the median raphe nucleus, without affecting 5-HT levels in the dorsal raphe nucleus. Animals which were housed in the startle chambers but were not given any stimuli were not different from controls. A comparable decrease in 5-HT in the startled animals was found in the hippocampus, which receives its serotonergic innervation from the median raphe nucleus. The results are discussed in the context of the general inverse relationship between startle reactivity and the functional status of the mesolimbic serotonergic pathway, and in the context of the effects of physical stressors on the utilization of brain 5-HT.     

Differential role of serotonin projections from the dorsal and median raphe nuclei in phencyclidine-induced hyperlocomotion and fos-like immunoreactivity in rats

Altered brain serotonin activity is implicated in schizophrenia. We have previously shown differential involvement of serotonergic projections from the dorsal or median raphe nucleus in phencyclidine‐induced hyperlocomotion in rats, a behavioral model of aspects of schizophrenia. Here we further investigated the effects of serotonergic lesions of the raphe nuclei on phencyclidine‐induced hyperlocomotion by parallel assessment of Fos‐like immunoreactivity (FLI), a marker of neuronal activation in the brain. Male Sprague‐Dawley rats were anesthetized with pentobarbitone and stereotaxically microinjected with 5 μg of the serotonergic neurotoxin, 5,7‐dihydroxytryptamine (5,7‐DHT), into either the dorsal raphe (DRN) or median raphe nucleus (MRN). Two weeks after the surgery, rats with lesions of the MRN, but not those with lesions of the DRN, showed significant enhancement of the hyperlocomotion induced by injection of 2.5 mg/kg of phencyclidine. Rats with MRN lesions also showed significantly higher levels of FLI in the polymorphic layer of the dentate gyrus in the dorsal hippocampus (PoDG) when compared with sham‐operated controls. Rats with lesions of the DRN showed significantly higher levels of FLI in the nucleus accumbens (NAcc). These results indicate that FLI in the PoDG, but not the NAcc, correlates with enhanced phencyclidine‐induced locomotor hyperactivity in MRN‐lesioned rats. These results support our previous studies suggesting a role of serotonergic projections from the MRN to the dorsal hippocampus in some of the symptoms of schizophrenia. Synapse, 2012. © 2012 Wiley Periodicals, Inc.     

Corticotropin-releasing factor in the dorsal raphe nucleus increases medial prefrontal cortical serotonin via type 2 receptors and median raphe nucleus activity

Interactions between central corticotropin-releasing factor (CRF) and serotonergic systems are believed to be important for mediating fear and anxiety behaviors. Recently we demonstrated that infusions of CRF into the rat dorsal raphe nucleus result in a delayed increase in serotonin release within the medial prefrontal cortex that coincided with a reduction in fear behavior. The current studies were designed to study the CRF receptor mechanisms and pathways involved in this serotonergic response. Infusions of CRF (0.5 μg/0.5 μL) were made into the dorsal raphe nucleus of urethane-anesthetized rats following either inactivation of the median raphe nucleus by muscimol (25 ng/0.25 μL) or antagonism of CRF receptor type 1 or CRF receptor type 2 in the dorsal raphe nucleus with antalarmin (25–50 ng/0.5 μL) or antisauvagine-30 (2 μg/0.5 μL), respectively. Medial prefrontal cortex serotonin levels were measured using in-vivo microdialysis and high-performance liquid chromatography with electrochemical detection. Increased medial prefrontal cortex serotonin release elicited by CRF infusion into the dorsal raphe nucleus was abolished by inactivation of the median raphe nucleus. Furthermore, antagonism of CRF receptor type 2 but not CRF receptor type 1 in the dorsal raphe nucleus abolished CRF-induced increases in medial prefrontal cortex serotonin. Follow-up studies involved electrical stimulation of the central nucleus of the amygdala, a source of CRF afferents to the dorsal raphe nucleus. Activation of the central nucleus increased medial prefrontal cortex serotonin release. This response was blocked by CRF receptor type 2 antagonism in the dorsal raphe. Overall, these results highlight complex CRF modulation of medial prefrontal cortex serotonergic activity at the level of the raphe nuclei.     

Differential effect of immobilization stress on in vivo synthesis rate of monoamines in medial prefrontal cortex and nucleus accumbens of conscious rats.

We have used microdialysis to measure the in vivo hydroxylation level of tyrosine and tryptophan in the medial prefrontal cortex and nucleus accumbens of conscious rats that were subjected to immobilization. The brain was perfused with an inhibitor of aromatic L-amino acid decarboxylase, 3-hydroxybenzylhydrazine, and the amount of 3,4-dihydroxyphenylalanine (DOPA) and 5-hydroxytryptophan (5-HTP) accumulating in the dialysate was measured as an index of the in vivo hydroxylation rate of tyrosine and tryptophan. One hour of immobilization caused a significant increase in extracellular DOPAin the medial prefrontal cortex but not nucleus accumbens. The same manipulation produced a significant and more prolonged elevation in extracellular 5-HTP in the nucleus accumbens as well as medial prefrontal cortex. The observed profile of stress-induced 5-HTP response was comparable in two brain regions. The results suggest that in vivo catecholamine synthesis is heterogenous, whereas in vivo serotonin synthesis is homogenous, with respect to responsiveness to stress in the medial prefrontal cortex and nucleus accumbens.     

Mirtazapine enhances frontocortical dopaminergic and corticolimbic adrenergic, but not serotonergic, transmission by blockade of alpha2-adrenergic and serotonin2C receptors: a comparison with citalopram

Mirtazapine displayed marked affinity for cloned, human alpha2A-adrenergic (AR) receptors at which it blocked noradrenaline (NA)-induced stimulation of guanosine-5'-O-(3-[35S]thio)-triphosphate ([35S]-GTPgammaS) binding. Similarly, mirtazapine showed high affinity for cloned, human serotonin (5-HT)2C receptors at which it abolished 5-HT-induced phosphoinositide generation. Alpha2-AR antagonist properties were revealed in vivo by blockade of UK-14,304-induced antinociception, while antagonist actions at 5-HT2C receptors were demonstrated by blockade of Ro 60 0175-induced penile erections and discriminative stimulus properties. Mirtazapine showed negligible affinity for 5-HT reuptake sites, in contrast to the selective 5-HT reuptake inhibitor, citalopram. In freely moving rats, in the dorsal hippocampus, frontal cortex (FCX), nucleus accumbens and striatum, citalopram increased dialysate levels of 5-HT, but not dopamine (DA) and NA. On the contrary, mirtazapine markedly elevated dialysate levels of NA and, in FCX, DA, whereas 5-HT was not affected. Citalopram inhibited the firing rate of serotonergic neurons in dorsal raphe nucleus, but not of dopaminergic neurons in the ventral tegmental area, nor adrenergic neurons in the locus coeruleus. Mirtazapine, in contrast, enhanced the firing rate of dopaminergic and adrenergic, but not serotonergic, neurons. Following 2 weeks administration, the facilitatory influence of mirtazapine upon dialysate levels of DA and NA versus 5-HT in FCX was maintained, and the influence of citalopram upon FCX levels of 5-HT versus DA and NA was also unchanged. Moreover, citalopram still inhibited, and mirtazapine still failed to influence, dorsal raphe serotonergic neurons. In conclusion, in contrast to citalopram, mirtazapine reinforces frontocortical dopaminergic and corticolimbic adrenergic, but not serotonergic, transmission. These actions reflect antagonist properties at alpha2A-AR and 5-HT2C receptors.     

Histologic and enzymatic studies of the mesolimbic and mesostriatal serotonergic pathways.

Selective lesions of the dorsal (B7), median (B8), or lateral (B9) raphe nuclei were made stereotaxically in male rats 4 weeks before sacrifice. The extent of damage to each raphe nucleus was quantified histologically by means of a simplified formaldehyde histochemical method for visualization of serotonin in cryostat sections. A detailed mapping of the distribution of the yellow-fluorescent raphe perikarya provided the basis for quantification. Tryptophan hydroxylase activity was measured in 6 forebrain regions from each animal, and the results were correlated with the per cent damage to each raphe nucleus. Tyrosine hydroxylase was also assayed in 5 of these regions; it was not significantly affected by any of the raphe lesions. Dorsal raphe lesions reduced tryptophan hydroxylase activity in the striatum, thalamus, cortex, and hypothalamus, but not in the septal nuclei or hippocampus. Damage to B8 resulted in decrements in this serotonergic enzyme in the septal nuclei, hippocampus, cortex, and hypothalamus, but not in the striatum or thalamus. Lesions of the scattered B9 cells had no significant effect on enzyme activity in any region examined. These data suggest that the dorsal and median raphe nuclei provide two distinct though perhaps overlapping serotonergic systems innervating different parts of the forebrain: a mesostriatal pathway originating in B7 and a mesolimbic system derived from B8. Behavioral studies on the animals, which are presented in a companion paper, indicated that damage to the median nucleus is responsible for many of the behavioral effects previously reported after combined lesions of both major raphe nuclei.     

Involvement of lateral habenula-dorsal raphe neurons in the differential regulation of striatal and nigral serotonergic transmission cats

The importance of the lateral habenula-dorsal raphe pathway in the control of in vivo [3H]serotonin release in the cat basal ganglia was examined using the push-pull cannula technique and an isotopic method for the estimation of [3H]serotonin continuously formed from [3H]tryptophan. [3H]Serotonin was measured in both caudate nuclei and substantiae nigra and, in some cases, in the dorsal raphe. Electrical stimulation of the lateral habenula decreased [3H]serotonin release in all structures studied. Blockade of the GABA inhibitory pathway to the lateral habenula by the local application of picrotoxin reduced [3H]serotonin release in both substantiae nigra and increased release of the 3H-amine in the dorsal raphe but was without effect on [3H]serotonin release in either caudate nucleus. This inhibition of nigral [3H]serotonin release was antagonized by simultaneous application of picrotoxin to the dorsal raphe. Substance P delivery to the dorsal raphe produced the same effects on [3H]serotonin release as described for picrotoxin application to the lateral habenula except that inhibition of nigral [3H]serotonin release was not prevented by local co-administration of picrotoxin. These results suggest that the lateral habenula can control serotonergic transmission in the basal ganglia and that this regulation may be different for those serotonergic neurons innervating the caudate nucleus versus those projecting to the substantia nigra.     

Changes of serotonin and dopamine metabolism in various forebrain areas of rats injected with morphine either systemically or in the raphe nuclei dorsalis and medianus

The regional brain metabolism of serotonin (5-HT) and dopamine (DA) was studied in rats injected with morphine either systemically or in the nuclei raphe medianus (MR) or dorsalis (DR). A subcutaneous injection of 10 mg/kg morphine significantly raised the levels of 5-hydroxyindoleacetic acid (5-HIAA) in the diencephalon, striatum, nucleus accumbens and cortex with no effect in the hippocampus. Similar changes in 5-HT metabolism were found in animals injected with 5 micrograms/0.5 microliter in the DR whereas morphine injected in the MR raised 5-HIAA levels only in the nucleus accumbens. A subcutaneous or direct injection of morphine in the DR significantly raised the levels of homovanillic acid (HVA) and dihydroxyphenylacetic acid (DOPAC) in the striatum and nucleus accumbens, but injection in the MR was ineffective. All the effects of morphine were blocked by naloxone, injected either intraperitoneally (1 mg/kg) or directly in the raphe nuclei (2 micrograms/0.5 microliter). Pretreatment with parachlorophenylalanine, an inhibitor of serotonin synthesis, significantly reduced the effect of morphine injected in the DR on dopamine metabolism in the striatum and nucleus accumbens. The data suggest that a major mechanism by which morphine increases 5-HT metabolism in the rat forebrain is activation of 5-HT cells in the nucleus raphe dorsalis, and this action may contribute to the increased DA metabolism found in the animal injected with morphine in this brain area.     

Effects of castration and adrenalectomy on in vitro rates of tryptophan hydroxylation and levels of serotonin in microdissected brain nuclei of adult male rats

Rates of 5-hydroxytryptophan (5-HTP) synthesis and levels of serotonin (5-HT) were measured in microdissected brain nuclei following castration or adrenalectomy of adult male rats. Fourteen days following gonadectomy, 5-HTP synthesis decreased in the nucleus raphe dorsalis (DR) and nucleus centralis superior (NCS), while levels of 5-HT were unchanged in the 7 brain nuclei examined. Administration of testosterone to castrated rats not only did not reverse the castration-induced decrease in 5-HTP synthesis in the DR and NCS, but also decreased 5-HT synthesis in the nucleus amygdaloideus centralis (AGC) and the nucleus septalis lateralis (LS). Following administration of testosterone, 5-HT levels were unchanged. 10 days following bilateral adrenalectomy, 5-HTP synthesis increased in the NCS and the median eminence. Levels of 5-HT increased only in the median eminence. The increased 5-HTP synthesis and 5-HT levels following adrenalectomy were not reversed by corticosterone administration. In addition, these selective changes in 5-HT metabolism did not result from hormonal effects on the availability of tryptophan to the brain.We conclude that there are subsets of serotonergic neurons in rat central nervous system which respond uniquely to removal of the gonads and adrenals. Furthermore, the dissociation between serum and brain tryptophan concentrations and changes in rates of 5-HTP synthesis argue against tryptophan availability as being a primary determinant of 5-HT biosynthesis and for a direct endocrine central nervous system interaction with serotonergic neurons.     

Site-specific activation of dopamine and serotonin transmission by aniracetam in the mesocorticolimbic pathway of rats

The effects of aniracetam on extracellular levels of dopamine (DA), serotonin (5-HT) and their metabolites were examined in five brain regions in freely moving stroke-prone spontaneously hypertensive rats (SHRSP) using in vivo microdialysis. Basal DA release in SHRSP was uniformly lower in all regions tested than that in age-matched control Wistar Kyoto rats. 3,4-Dihydroxyphenylacetic acid and homovanillic acid levels were altered in the basolateral amygdala, dorsal hippocampus and prefrontal cortex of SHRSP. While basal 5-HT release decreased in the striatum and increased in the basolateral amygdala, there was no associated change in 5-hydroxyindoleacetic acid levels. Systemic administration of aniracetam to SHRSP enhanced both DA and 5-HT release with partly associated change in their metabolite levels in the prefrontal cortex, basolateral amygdala and dorsal hippocampus, but not in the striatum and nucleus accumbens shell, in a dose-dependent manner (30 and/or 100 mg/kg p.o.). Microinjection (1 and 10 ng) of aniracetam or its metabolites (N-anisoyl-GABA and 2-pyrrolidinone) into the nucleus accumbens shell produced no turning behavior. These findings indicate that SHRSP have a dopaminergic hypofunction throughout the brain and that aniracetam elicits a site-specific activation in mesocorticolimbic dopaminergic and serotonergic pathways in SHRSP, possibly via nicotinic acetylcholine receptors in the ventral tegmental area and raphe nuclei. The physiological roles in the aniracetam-sensitive brain regions may closely link with their clinical efficacy towards emotional disturbances appearing after cerebral infarction.     

Serotonergic neurons in the brainstem of the wallaby, Macropus eugenii.

The organisation and cytoarchitecture of the serotonergic neurons in a diprotodont marsupial were examined by using serial sections of the brainstem processed for serotonin immunohistochemistry and routine histology. The topographic distribution of serotonergic neurons in the brainstem of the adult wallaby (Macropus eugenii) was similar to that of eutherian mammals. Serotonergic neurons were divided into rostral and caudal groups, separated by an oblique boundary through the pontomedullary junction. Approximately 52% of the serotonergic neurons in the wallaby brainstem were located in the rostral midline nuclei (caudal linear nucleus, dorsal, median, and pontine raphe nuclei and the interpeduncular nucleus), whereas 21% were found in the caudal midline region (nuclei raphe magnus, obscurus, and pallidus). The remaining serotonergic neurons (27%) were located in more lateral regions such as the pedunculopontine tegmental nuclei, the supralemniscal nuclei (B9 group), and the ventrolateral medulla. The largest serotonergic group, the dorsal raphe, contained one-third of the brainstem serotonergic neurons and showed five subdivisions, similar to that described in other species. In contrast, the median raphe did not show clear subdivisions. The internal complexity of the raphe nuclei and the degree of lateralisation of serotonergic neurons suggest that the wallaby serotonergic system is similar in organisation to that described for the cat and rabbit. This study supports the suggestion that the serotonergic system is evolutionally well conserved and provides baseline data for a quantitative study of serotonergic innervation of the developing cortex in the wallaby.     

Voluntary wheel running produces resistance to inescapable stress-induced potentiation of morphine conditioned place preference

In rodents, exposure to acute inescapable, but not escapable, stress potentiates morphine conditioned place preference (CPP), an effect that is dependent upon hyperactivation of serotonin (5-HT) neurons in the dorsal raphe nucleus (DRN). Six weeks of voluntary wheel running constrains activation of DRN 5-HT neurons during exposure to inescapable stress. Six weeks of voluntary wheel running before inescapable stress blocked stress-induced potentiation of morphine CPP.     

Evidence that serotonergic neurons in the dorsal raphe nucleus exert a stimulatory effect on the secretion of renin but not of corticosterone

Data from previous experiments in rats indicate that release of serotonin in the central nervous system increases renin and corticosterone secretion. To determine which serotonergic neurons are involved, lesions of the dorsal or median raphe nuclei were produced by local injections of 5,7-dihydroxytryptamine (5,7-DHT) in rats, and 2 weeks later, the renin responses to parachloroamphetamine (PCA) were determined. Plasma corticosterone was also measured. PCA produced significant increases in plasma renin activity and plasma corticosterone in sham-lesioned animals and animals with median raphe lesions. The plasma corticosterone response to PCA was also normal in rats with dorsal raphe lesions but the renin response was significantly reduced. The data support the hypothesis that serotonergic neurons in the dorsal raphe nucleus are part of a neural pathway mediating increased renin secretion, and that the stimulatory effect of serotonin on corticosterone secretion is mediated by a different pathway.     

Distribution and neurochemical characterization of neurons expressing GIRK channels in the rat brain

G-protein inwardly rectifying potassium (GIRK) channels mediate the synaptic actions of numerous neurotransmitters in the mammalian brain and play an important role in the regulation of neuronal excitability in most brain regions through activation of various G-protein-coupled receptors such as the serotonin 5-HT(1A) receptor. In this report we describe the localization of GIRK1, GIRK2, and GIRK3 subunits and 5-HT(1A) receptor in the rat brain, as assessed by immunohistochemistry and in situ hybridization. We also analyze the co-expression of GIRK subunits with the 5-HT(1A) receptor and cell markers of glutamatergic, gamma-aminobutyric acid (GABA)ergic, cholinergic, and serotonergic neurons in different brain areas by double-label in situ hybridization. The three GIRK subunits are widely distributed throughout the brain, with an overlapping expression in cerebral cortex, hippocampus, paraventricular nucleus, supraoptic nucleus, thalamic nuclei, pontine nuclei, and granular layer of the cerebellum. Double-labeling experiments show that GIRK subunits are present in most of the 5-HT(1A) receptor-expressing cells in hippocampus, cerebral cortex, septum, and dorsal raphe nucleus. Similarly, GIRK mRNA subunits are found in glutamatergic and GABAergic neurons in hippocampus, cerebral cortex, and thalamus, in cholinergic cells in the nucleus of vertical limb of the diagonal band, and in serotonergic cells in the dorsal raphe nucleus. These results provide a deeper knowledge of the distribution of GIRK channels in different cell subtypes in the rat brain and might help to elucidate their physiological roles and to evaluate their potential involvement in human diseases.     

Brain serotonin depletion by lesions of the median raphe nucleus enhances the psychotomimetic action of phencyclidine, but not dizocilpine (MK-801), in rats

We have previously shown that brain serotonin depletion by lesions of the median raphe nucleus (MRN) causes enhancement of phencyclidine-induced locomotor hyperactivity [S. Kusljic, D.L. Copolov, M. van den Buuse, Differential role of serotonergic projections arising from the dorsal and median raphe nuclei in locomotor hyperactivity and prepulse inhibition, Neuropsychopharmacology 28 (2003) 2138-2147]. In this study, we extend our previous work by (1) comparing the effect of phencyclidine with that of another NMDA receptor antagonist, dizocilpine (MK-801); (2) investigate behavioral changes in more detail; (3) assess in detail the effect of raphe lesions on regional serotonin levels in the brain. Male Sprague-Dawley rats received microinjection of the serotonergic neurotoxin 5,7-dihydroxytryptamine into the MRN or dorsal raphe nucleus (DRN). The effects of treatment with saline, phencyclidine and MK-801 on locomotor activity were determined 2 weeks after the surgery. MRN lesions caused serotonin depletion in the dorsal hippocampus, whereas DRN lesions caused serotonin depletion in the frontal cortex, striatum and ventral hippocampus. There was a significant increase in phencyclidine-induced locomotor hyperactivity in the MRN-lesioned group compared to sham-operated controls. Further analysis of behavior showed that phencyclidine-induced hyperambulation, but not stereotypy or rearing, was significantly higher in MRN-lesioned rats compared to controls. In contrast, there was no significant effect of the lesions on the psychotomimetic effect of MK-801. These results indicate that a hyposerotonergic state induced by destruction of projections from the MRN leads to altered brain circuitry that is responsible for the regulation of phencyclidine-but not MK-801-induced locomotor hyperactivity. Thus, MRN projections may play an inhibitory role in mechanisms involved in symptoms of schizophrenia.     

Serotonergic lesion of median raphe nucleus alters nerve growth factor content and vulnerability of cholinergic septohippocampal neurons in rat.

About 45% of the serotonergic raphe neurons are reported to express nerve growth factor (NGF) receptors. We therefore investigated whether selective serotonergic lesions of the median or dorsal raphe nuclei are associated with changes in NGF protein levels of the brain and whether the loss of serotonergic function alters the vulnerability of cholinergic septohippocampal neurons. In adult rats the hippocampal NGF content changed in a biphasic way after lesion of the median raphe nucleus by 5,7-dihydroxytryptamine (5,7-DHT), with a significant increase after 2-3 weeks of up to 35%, followed by a significant reduction of 22% below control levels after 7 weeks, and a return to control levels within the following 4 weeks. By contrast, the decrease in hippocampal serotonin and 5-hydroxyindoleacetic acid remained throughout the observation period of 11 weeks, being still reduced to 15 and 30% of the control levels, respectively. In the frontal cortex the partial loss of the serotonergic innervation projecting from the median raphe was associated 5 weeks after 5,7-DHT injection with an increase in NGF protein of 39.7+/-9.6% (P<0.05), which remained elevated up to 11 weeks. At 9 weeks after 5,7-DHT, the lesion of the septohippocampal cholinergic neurons induced by the cholinotoxin ethylcholine aziridinium (AF64A) was exaggerated (P<0.05) as compared to AF64A-treated rats with intact serotonergic innervation. The present data indicate that a serotonergic lesion of the median raphe nucleus results in biphasic changes of NGF protein content and in a delayed increase in the vulnerability of septohippocampal cholinergic neurons.