Significant non-serotonergic raphe projection to the visual cortex of the rat. An immunohistochemical study combined with retrograde tracing.

The present study investigated the distribution of serotonergic and non-serotonergic raphe neurons with direct projections to the visual cortex. The study employed the WGA-apoHRP-Au retrograde transport technique combined with 5-HT immunohistochemical staining. Retrogradely labeled cells were observed in the dorsal raphe nucleus, the median raphe nucleus, and in the B9 and B6 cell groups. One notable finding was the great number of retrogradely labeled, non-5-HT immunoreactive cells. The average percentages of such cells in the various raphe regions were as follows: DR: 52% (n = 401); MR: 35% (n = 311); B9: 24% (n = 129); B6: 95% (n = 200). The present study demonstrated the presence of a significant proportion of non-serotonergic raphe region neurons projecting to the primary visual cortex in the rat. It is suggested that these neurons may complement the aminergic neurons as part of the ascending system which controls the functions of the visual cortex.     

Organization of raphe-cortical projections in rat: A quantitative retrograde study

Retrograde transport of a fluorescent dye was employed to study the projections from raphe nuclei to neocortex in the rat. The spatial distributions of labeled raphe cells were analyzed quantitatively to determine whether the nuclei are topographically organized with respect to different cortical targets. The dorsal raphe nucleus (DRN), exclusive of the lateral wing regions, has a predominantly (3:1) ipsilateral projection with decreasing numbers of cells projecting to frontal, parietal, and occipital cortex. Overlapping cell groups within the DRN project differentially to these three cortical areas: DRN cells innervating frontal cortex extend more rostrally and laterally than those to either parietal or occipital cortex. The medium raphe and B9 projections are bilaterally symmetric, with equal cell numbers projecting to frontal, parietal, and occipital cortex. The rostro-caudal distributions of cells that project to disparate cortical areas differ in B9 but not in MR. The percentage of cortically projecting cells that are serotonergic is 80% for the DRN, 60% in the MR and 33% in the B9 cell group. The dorsal raphe nucleus and the B9 cell group are organized heterogeneously, and overlapping sets of neurons project differentially upon particular areas of neocortex. In contrast, the median raphe nucleus projects uniformly upon the neocortex and does not exhibit topographic organization. The three rostral raphe nuclei (DR, MR and B9) are each organized according to different rules with regard to their efferent projections to cortex. The differential organization of the raphe nuclei suggests that groups of cells within these three raphe nuclei are likely to innervate different combinations of cortical targets and thus to have different functional effects.     

Effects of parachloroamphetamine upon the serotonergic innervation of the rat hippocampus

The ability of hippocampal serotonergic (5-HT) axons to proliferate in response to damage by para-chloroamphetamine (PCA) was examined in this study. Synaptosomal uptake of 5-HT in the hippocampal formation was decreased to 40% of control 3 days after systemic administration of PCA. Six weeks after PCA, uptake values were 44% of control. Retrograde tracing combined with 5-HT immunocytochemistry showed a significant reduction (18% of control) in the number of 5-HT raphe neurons projecting to the hippocampus 3 days after PCA. The number of 5-HT neurons projecting to the hippocampal formation increased to 69% of control by 6 weeks. The dorsal raphe nucleus was not retrogradely labeled after PCA; the increase in labeled neurons was observed in the median raphe nucleus. PHA-L, injections of the median raphe nucleus demonstrated a reduction of raphe axons in the hippocampal formation after PCA. In rats treated with PCA, raphe axons labeled with PHA-L also appeared to have fewer boutons than raphe axons labeled in control cases. The density of PHA-L containing axons in the hippocampal formation of rats injected 3 days and 6 weeks after PCA was less than control but there was no difference between the experimental groups. Based upon the results from synaptosomal uptake and anterograde tracing experiments, we feel that compensatory proliferation of 5-HT axons does not occur within 6 weeks of PCA-induced damage to the 5-HT plexus of the hippocampal formation. The data derived from the retrograde tracing experiment are thought to reflect reduced uptake and transport of WGA-HRP as an acute effect of PCA.     

Extent of colocalization of serotonin and GABA in the neurons of the rat raphe nuclei

Previous investigations of the distribution of neurons containing both serotonin and GABA in the brainstem raphe nuclei have yielded discrepant results amongst different authors. This study attempted to clarify the distribution as well as the proportions of raphe and other brainstem neurons that contain both neurotransmitters. All the nine serotonergic cell groups known to be present in the brainstem were examined with an indirect immunofluorescence method using antibodies against serotonin and glutamic acid decarboxylase in colchicine-treated rats. Sections were incubated either simultaneously or sequentially for the two immunolabels. Brainstem neurons that were labelled for both markers were generally infrequent. Of all the serotonin cell groups in the brainstem, the nucleus raphe magnus contained the most double-labelled cells (a mean of 3.6% of a total of 625–1155 serotonin-immunoreactive cells counted in this nucleus), followed by the nucleus raphe obscurus (1.5% of a total of 220–550 serotonin-immunoreactive neurons counted). The dorsal, median and pontine raphe nuclei as well as the supralemniscal nucleus (the B9 group) contained very few double-labelled cells, which comprised a mean of 0.1–0.7% of all serotonin-immunoreactive cells in each of these nuclei. No double labelled cells were present in the caudal linear raphe nucleus or the nucleus raphe pallidus, nor in the B4 group. These results suggest that only a very small percentage of serotonergic neurons in the medullary raphe nuclei (raphe magnus and raphe obscurus) also contain GABA, whereas such cells are virtually absent in the midbrain raphe nuclei or in the non-raphe serotonergic cell groups in the brainstem.     

Chronic degeneration of dorsal raphe serotonergic neurons modulates cortical spreading depression: A possible pathophysiology of migraine

The vascular serotonergic system in the brain has been implicated in the pathophysiology of migraine, however, involvement of the serotonergic nervous system of the brain parenchyma in the pathophysiology remains unclear. To investigate whether the brain parenchymal serotonergic nervous system is involved in the etiology of migraine, we prepared an experimental model of migraine by generation of cortical spreading depression (SD), characterized by spreading of neuronal/glial membrane depolarization accompanied by temporal elevation of the cerebral blood flow (CBF) throughout the cerebral cortical hemisphere in rats, which underwent pharmacological treatment for degeneration of serotonergic neurons in the dorsal raphe nucleus. We show here that 1) significant degeneration of serotonergic neurons in the dorsal raphe nucleus and serotonergic fibers in the cerebral cortex was observed in treated rats, 2) spreading velocity of the CBF changes was significantly increased in these rats, and 3) calculated width of the depolarization wave was significantly extended in these rats. These results indicate that the dorsal raphe serotonergic neurons modulate cortical spreading depression and might be involved in migraine pathology via a similar mechanism. © 2013 Wiley Periodicals, Inc.     

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.     

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.     

Proportion of glutamate- and aspartate-immunoreactive neurons in the efferent pathways of the rat visual cortex varies according to the target.

Immunohistochemistry, with antisera directed against glutamate (Glu) or aspartate (Asp), was combined with wheat germ agglutinin-horseradish peroxidase (WGA-HRP) histochemistry to examine the distribution, morphology, and proportions of Glu- and Asp-containing neurons that give rise to corticofugal and callosal projections of the rat visual cortex. WGA-HRP injections in the dorsal lateral geniculate nucleus resulted in retrograde labelling of small and medium-sized cells throughout layer VI of the visual cortex. Of these cells, 60% were also Glu-immunoreactive and 61% Asp-positive. WGA-HRP injections in the superior colliculus labelled large and medium-sized neurons in the upper portion of layer V of the visual cortex. Of these cells, 46% were also stained for Glu and 66% for Asp. Injections in the pontine nuclei resulted in retrograde labelling of cells in the deeper part of cortical layer V. Retrogradely labelled cells, which were also immunoreactive for Glu or Asp, were large pyramidal cells. Corticopontine neurons, which were also Glu-positive, accounted for 42% of the total number of WGA-HRP labelled cells, whilst for Asp-positive neurons this percentage was 51%. Finally, after injections in the visual cortex, retrogradely labelled small and medium-sized cells were found throughout layers II-VI in the contralateral visual cortex. Of these neurons, 38% were also labelled for Glu while 49% were also Asp-immunoreactive. The present results demonstrate that substantial proportions of projection neurons in the rat visual cortex are immunoreactive for Glu or Asp, suggesting that these excitatory amino acids are the major transmitters used by the cortical efferent systems examined. Furthermore, the proportions of these immunoreactive neurons in the efferent pathways vary according to the target.     

Further studies on the activation of rat median raphe serotonergic neurons by inescapable sound stress

Previous studies, using a biochemical measure of serotonergic neuronal function, show that inescapable, randomly presented sound pulses activate serotonergic neurons in the rat median raphe but not dorsal raphe nucleus. The present study reveals that this activation also occurs in serotonin projection areas, in hippocampus, nucleus accumbens and cortex but not in caudate nucleus. The selectivity of this response is examined by comparing the response to sound stress with that produced by morphine, a treatment known to selectively activate dorsal raphe but not median raphe serotonergic neurons. Two approaches are used in Sprague-Dawley rat to measure the activation of serotonergic neurons: (1) determination ex vivo of accumulation of 5-hydroxytryptophan (5-HTP) in tissue from the dorsal and median raphe nuclei, hippocampus, cortex, caudate nucleus, and nucleus accumbens following in vivo inhibition of aromatic amino acid decarboxylase; and (2) measurement of extracellular serotonin levels in hippocampus, caudate nucleus, and nucleus accumbens. Sound stress increases 5-HTP accumulation in median raphe nucleus, hippocampus, cortex, and nucleus accumbens, but not dorsal raphe nucleus or caudate nucleus. Sound stress also enhances extracellular serotonin levels in hippocampus and nucleus accumbens, but not caudate nucleus. In contrast, the morphine treatment enhances 5-HTP accumulation in dorsal raphe nucleus, cortex and caudate nucleus, but not in median raphe nucleus, hippocampus or nucleus accumbens. Furthermore, it increases extracellular serotonin levels in only the caudate nucleus. The combined effects of sound stress and morphine on 5-HTP accumulation are identical to those obtained by each treatment individually. These findings provide further support for the presence of serotonergic neurons within the median raphe nucleus that have a unique response profile. These neurons may have an important role in responses or adaptations to stress.     

Serotonergic and non-serotonergic projections from the raphe nuclei to the piriform cortex in the rat: a cholera toxin B subunit (CTb) and 5-HT immunohistochemical study

Retrograde axonal transport of the cholera toxin B subunit (CTb) was combined with 5-HT immunohistochemistry to determine the origin of the serotonergic innervation of the piriform cortex (PC) in the rat. After iontophoretic CTb injections in the PC, a substantial number of retrogradely labeled cells were found in the middle and medio-ventral part of the dorsal raphe nucleus (RD). A few retrogradely labeled cells were also observed in the median raphe nucleus (MnR) and the B9 serotonergic cell groups. Following CTb and 5-HT immunohistochemistry on the same sections, double-labeled cells were observed in the RD, MnR and B9 groups. In the RD, 30% of CTb stained cells were immunoreactive to 5-HT. After colchicine or nialamide (a monoamine oxidase inhibitor) pretreatment the percentage of these double-labeled cells reached 70%. These results indicate that both 5-HT and non-5-HT neurons in the RD innervate the PC and that the percentage of double-labeled cells is influenced by drug pretreatment. To determine the terminal fields of the RD efferent fibers in the PC, injections of the anterograde tracer PHA-L were also performed. Analysis of the fiber distribution in the PC further revealed some medio-lateral and antero-posterior differences.     

Developmental alteration of serotonin neurons in the raphe nucleus of rats with methylazoxymethanol-induced microcephaly

Summary Prenatal exposure of pregnant rats to methylazoxymethanol acetate (MAM), an anti-mitotic agent, on day 15 of gestation induces severe microcephaly in the offspring. The present study first investigated a developmental alteration of serotonin (5HT) neurons immunohistochemically in the dorsal and median raphe nuclei in serial sections in both control and microcephalic rats (MAM-rats) at 35 days of age. 5HT-immunoreactive neurons in the MAM-rats were reduced in number and irregularly distributed in the dorsal and median raphe nuclei compared with those in the control. The dendrites of neurons in these nuclei in the MAM-rats were very short and twisted. A follow-up observation on the development of the cerebral cortex at 5, 9 and 28 days of age was performed using Nissl-stained preparations, which revealed a disorganized cell arrangement in the cerebral cortex of the MAM-rats at the very early postnatal period. Furthermore, the distribution of 5HT-immunoreactive fibers into the cerebral cortex was also examined using brains of 28 days of age. In MAM-rats of this age, abnormally tortuous 5HT-immunoreactive fibers were observed in the cerebral cortex. 5HT neurons in the raphe nuclei are known to project their ascending axons widely into the entire cortical area during the 1st postnatal week. Thus, the association of disorganized cortical cell arrangement and the hyperdense and tortuous distribution of 5HT-immunoreactive fibers in the cerebral cortex support the idea of target-dependent secondary degeneration of 5HT neurons in the dorsal and median raphe nuclei of the MAM-rats.     

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.     

Origin of the serotonergic innervation to the rat dorsolateral hypothalamus: retrograde transport of cholera toxin and upregulation of tryptophan hydroxylase mRNA expression following selective nerve terminals lesion.

The regulation of serotonin synthesis was investigated in the serotonergic neurons, which provide afferents to the dorsolateral hypothalamus (DLH). The origin of the DLH projection neurons within the raphe nucleus was identified by retrograde transport of Cholera toxin (CTb) and their serotonergic nature confirmed by tryptophan hydroxylase (TPH) immunocytochemistry. Disruption of serotonin synthesis steady-state was induced unilaterally by a selective and local destruction of serotonergic nerve terminals with 5,7-dihydroxytryptamine (5,7-DHT), stereotaxically injected in the right DLH. The results show that most of the serotonergic dorsal raphe neurons projecting to the DLH have an ipsilateral localization within the lateral aspects of the nucleus. In rats with unilateral DLH lesion, a population of serotonergic cells within the raphe nucleus exhibited a clear increase in TPH mRNA. These cells were about five times more numerous in the ipsilateral as compared to the contralateral dorsal raphe nucleus and they had, for the most part, a lateral localization within the raphe nucleus. Sham-operated rats did not exhibit any upregulation of TPH mRNA. Together, the present results provide the first demonstration that a discreet and selective destruction of serotonergic terminals induces a circumscribed and striking increase in TPH mRNA expression in a subset of brainstem serotonergic neurons projecting to and/or passing through the DLH. On the basis of these results and previous in vivo measurements of TPH activity (e.g., 5-HT synthesis), we suggest that this upregulation in TPH mRNA expression results from the loss of pre-synaptic and/or post-synaptic regulation of serotonin synthesis. These new findings raise important issues related to the repercussions of a local disruption in serotonergic neurotransmission on brain areas remote from the site of injury.     

Development of the projections from the dorsal lateral geniculate nucleus to the lateral suprasylvian visual area of cortex in the cat.

In the study reported in the preceding paper, we used retrograde labeling methods to show that the enhanced projection from the thalamus to the posteromedial lateral suprasylvian (PMLS) visual area of cortex that is present in adult cats following neonatal visual cortex damage arises at least partly from surviving neurons in the dorsal lateral geniculate nucleus (LGN). In the C layers of the LGN, many more cells than normal are retrogradely labeled by horseradish peroxidase (HRP) injected into PMLS cortex ipsilateral to a visual cortex lesion. In addition, retrogradely labeled cells are found in the A layers, which normally have no projection to PMLS cortex in adult cats. The purpose of the present study was to investigate the mechanisms of this enhanced projection by examining the normal development of projections from the thalamus, especially the LGN, to PMLS cortex. Injections of HRP were made into PMLS cortex on the day of birth or at 1, 2, 4, or 8 weeks of age. Retrogradely labeled neurons were present in the lateral posterior nucleus, posterior nucleus of Rioch, pulvinar, and medial interlaminar nucleus, as well as in the LGN, at all ages studied. Within the LGN of the youngest kittens, a small number of retrogradely labeled cells was present in the interlaminar zones and among the cells in the A layers that border these zones. Such labeled cells were virtually absent by 8 weeks of age, and they are not found in normal adult cats. Sparse retrograde labeling of C-layer neurons also was present in newborn kittens.(ABSTRACT TRUNCATED AT 250 WORDS)     

Bihemispheric Axonal Bifurcation of the Afferents to the Visual Cortical Areas during Postnatal Development in the Rat

Numerous cortical neurons in the juvenile and adult rat project to visual areas of both hemispheres whereas the vast majority of subcortical structures projecting to the visual cortex send strictly ipsilateral projections (Dreher et al., 1990). In the present study, the authors have sought to determine whether this pattern of axonal bifurcation in the connectivity of the visual areas undergoes a change during postnatal development. Two retrograde fluorescent dyes were used, fast blue (FB) and diamidino yellow (DY). Large multiple injections of one of the dyes were placed in all visual areas of one hemisphere and a small injection of the other dye was placed in area 17 of the opposite hemisphere. Labelled neurons were observed in subcortical and cortical structures on the side of the small injection. The experiments were performed on ten neonatal albino rat pups aged between 3 and 12 postnatal days (p.n.d.) at the time of injection and the results were compared with those obtained in the juvenile and adult animals, as reported in the preceding paper. In the thalamus of newborn animals, neurons belonging to nuclei located away from the midline send strictly ipsilateral cortical projections. However, in the midline nuclei of the intralaminar thalamic complex, a small region of overlap was observed between neurons projecting ipsilaterally and neurons projecting contralaterally in animals aged less than 9 postnatal days. In addition, in these neonatal animals a small number of bilaterally projecting neurons was detected in this region of overlap. In all other subcortical structures examined (ventral tegmental area, diagonal band of Broca, claustrum), the laterality of the projection was the same in the newborn and the adult animals. In particular, in the claustrum of neonatal animals, as in adult animals, there was a large contingent of contralaterally projecting neurons and only a very small number of bilaterally projecting neurons. The results in the cortex contrast with those observed in subcortical structures. Whereas ipsilaterally projecting neurons were distributed in a broadly similar way in newborn and adult animals, the laminar and areal distribution of contralaterally projecting neurons in newborn animals clearly differed from those observed in the adult animals. Furthermore, double labelled neurons were more numerous in animals aged less than 12 days than in adults. The proportions of such bilaterally projecting neurons were computed with respect to the numbers of neurons sending ipsilateral projections to area 17. These proportions are constant at all ages in the claustrum and cortical area 8. In areas 18a, 29 and 35 on the other hand, the proportions of bilaterally projecting neurons increase after 5 days and reach a peak in the period extending from 9 to 11 days of age when more than half of the neurons projecting ipsilaterally also send an axonal branch to the contralateral cortex. In cortical areas 29 and 35, this peak is followed by a sudden drop to the adult level at 12 postnatal days, whereas the return to the adult level is gradual in area 18a. These results demonstrate that, in subcortical structures and in cortical area 8, the laterality of the afferent connections to the visual cortex does not change during postnatal development. By contrast, cortical areas 18a, 29 and 35 go through a stage when numerous cells send bifurcating connections to both hemispheres. The timing of the decrease in proportions of bilaterally projecting neurons in these areas suggests that numerous neurons retract their callosal axonal branch when the adult pattern of callosal connectivity is established at 9 - 11 days of age.     

Activity of serotonin-containing neurons in nucleus raphe magnus in freely moving cats

Serotonergic neurons were recorded in the nucleus raphe magnus in freely moving cats and were initially identified on-line by their characteristic slow and regular spontaneous activity during quiet waking (3.42 +/- 0.33 spikes/s; mean +/- SE). Discharge rates of these serotonergic neurons were highest during active waking (4.49 +/- 0.40 spikes/s), intermediate during slow-wave sleep (middle: 2.14 +/- 0.23 spikes/s), and lowest during REM sleep (0.20 +/- 0.03 spikes/s). Although these cells fired at a rate 31.3% higher during active waking than during quiet waking, their activity displayed no correlation with phasic elevations of the nuchal EMG or overt body movements. In addition, no relationship was observed between the activity of these neurons during slow-wave sleep and the occurrence of sleep spindles in the cortical EEG or pontogeniculooccipital waves recorded from the lateral geniculate nucleus. Serotonergic neurons of nucleus raphe magnus were also relatively unresponsive to phasic auditory and visual stimuli, with about half of the cells examined showing weak excitatory responses. These neurons did respond, however, to the administration of a small dose of the serotonin specific agonist, 5-methoxy-N,N-dimethyltryptamine (250 micrograms/kg, i.m.) with a mean decrease in unit activity of 73.6 +/- 4.5%. The results of this study are compared with those previously reported for serotonergic neurons in the dorsal raphe nucleus, nucleus centralis superior, and nucleus raphe pallidus of freely moving cats.     

鱼藤酮对大鼠脑内5-羟色胺能神经元损伤的机制研究

目的研究鱼藤酮对大鼠脑内5-羟色胺能神经元的毒性作用及其机制。方法健康成年雄性Wistar大鼠背部皮下注射鱼藤酮制备帕金森病动物模型。采用免疫组化、蛋白印迹及分光吸光度法检测大鼠中脑中缝背核5-羟色胺能神经元的损伤及中缝背核氧化应激参数(丙二醛和还原型谷胱甘肽)的改变。结果和对照组相比,鱼藤酮组大鼠脑内中缝背核5-羟色胺(5-HT)免疫反应阳性神经元数明显少于对照组(P<0.01),Western blot结果显示中缝背核5-HT表达在鱼藤酮组明显降低(P<0.01);鱼藤酮组大鼠中缝背核区域丙二醛含量明显增高(P<0.01)、还原型谷胱甘肽(GSH)含量明显减少(P<0.01)。结论鱼藤酮在损伤大鼠脑内多巴胺能神经元的同时对5-HT能神经元也具有明显的毒性作用,氧化应激可能是导致5-HT神经元损伤的主要原因。     

Intraocular co-grafts of fetal dorsal raphe nucleus and suprachiasmatic nucleus

Serotonergic neurons in the fetal dorsal raphe nucleus were grafted together with fetal anterior hypothalamic tissue including the suprachiasmatic nucleus (SCN) to the anterior eye chamber of adult rats. After 6 weeks transplantation, the double grafts were immunocytochemically examined using antisera against serotonin, arginine vasopressin (AVP) and vasoactive intestinal polypeptide (VIP). The raphe grafts contained a large number of serotonin-immunoreactive neurons and fibers, but only a few AVP-immunoreactive fibers and VIP-immunoreactive neurons and fibers. On the other hand, numerous AVP- and VIP-immunoreactive neurons and fibers were found in the SCN of the anterior hypothalamic graft. Outgrowing serotonin-immunoreactive fibers from the raphe tissue were densely distributed in the anterior hypothalamic graft. In the SCN, however, only a few fibers were detected. The results demonstrate that the isolated anterior hypothalamic grafts can be innervated by the serotonergic neurons from the raphe grafts, but the innervation pattern of these fibers was quite different from the normal rat. The present results indicate that the isolated SCN has an inhibitory influence on the growth of serotonergic fibers.     

Transient cortical pathways in the pyramidal tract of the neonatal ferret

Anterograde transport of wheat germ agglutinin conjugated to horseradish peroxidase (WGA-HRP) was used to study transient axons from the visual cortex in the pyramidal tract. Injections at birth restricted to the visual cortex labeled axons in the vicinity of the pontine nuclei. Two to eight days after birth, axons from the occipital cortex were found posterior to the pontine nucleus, their caudalmost stable target. Transient corticospinal axons from the presumptive primary visual cortex did not grow caudal to the pyramidal decussation. Innervation of more distal targets preceded innervation of proximal targets. Innervation of the pontine nucleus is initiated around 68 hours after birth, when the transient extension in the medullary pyramidal tract has attained its maximum caudal extent. Innervation of the superior colliculus begins 9 days after birth. Retrograde tracers were used to follow the developmental changes in the cortical distribution of the parent neurons giving rise to axons in the pyramidal tract. In the adult, labeled neurons following injection of retrograde tracer in the pyramidal tract occupied less than a third of the neocortex and were centred on the anterior part of the coronal and spleniocruciate gyri. In the immature brain, labeled neurons covered more than two-thirds of the neocortex. Areal density measurements in the neonate showed that peak labeling was centred in the anterior coronal and spleniocruciate gyri, where corticospinal cells in the adult are located. There was a marked rostral-caudal gradient so that labeled neurons were very scarce towards the occipital pole. These results, showing transient neocortical axons in the pyramidal tract in a carnivore, suggest that this may be a common feature of mammalian development. The finding that the adult pattern of corticospinal projections does not emerge from a uniform distribution is discussed with respect to the areal specification of cortical connectivity. © 1993 Wiley-Liss, Inc.     

Distribution,quantification, and morphological characteristics of serotonin-immunoreactive cells of the supralemniscal nucleus (B9) and pontomesencephalic reticular formation in the rat

In their initial report on the rat, Dahlstrom and Fuxe ([1964] Acta Physiol. Scand. 62:1–55) identified nine brainstem serotonin-containing cell groups, which they termed B1–B9. B9 has received considerably less attention than other serotonergic nuclei (B1–B8) due in part to the fact that its precise location and extent have not been well documented in subprimates. B9 (supralemniscal nucleus; SLN) has been viewed as a minor serotonergic cell group. In addition, 5-hydroxytryptamine (5-HT)-containing cells have been shown to be only sparsely distributed throughout the pontomesencephalic reticular formation (PMRF). By using 5-HT immunohistochemical techniques, we examined the distribution and morphological characteristics of SLN and PMRF 5-HT neurons of the pontomesencephalic tegmentum. We showed that 5-HT cells of both the SLN and the PMRF extend rostrocaudally from the rostral midbrain to the midpons. 5-HT SLN cells are located within or dorsal to the medial lemniscus (ML); those of the PMRF are widely distributed throughout the PMRF. The mean numbers of 5-HT containing cells in the SLN, PMRF, dorsal raphe, and median raphe nuclei were 4,571, 1,948, 15,191, and 4,114, respectively. The SLN (B9) contains more 5-HT neurons than any serotonergic group other than the dorsal raphe nucleus. The dendrites of both SLN and PMRF 5-HT cells are primarily oriented mediolaterally and generally extend for long distances (75–300 μm), running perpendicular to the fibers of the ML (SLN) or, to those coursing through the brainstem (PMRF). The present anatomical delineation of SLN and PMRF shows that they are major 5-HT-containing cell groups in the rat and provides the foundation for the further examination of their properties and functions. J. Comp. Neurol. 378:411–424, 1997. © 1997 Wiley-Liss, Inc.