Tanemichi Chiba

Afferent and efferent connections of the medial preoptic area in the rat: A WGA-HRP study

Afferent and efferent connections of the medial preoptic area including medial preoptic nucleus (MP) and periventricular area at the MP level were examined using WGA-HRP as a marker. Injections were performed by insertion of micropipette containing (1) small amount of HRP powder or (2) dryed HRP solution for 24 to 48 hr until the fixation or for 5 min respectively. Dorsal and ventral approaches of injection micropipettes were performed and the results were compared. Previously reported reciprocal connections with lateral septum, bed nucleus of the stria terminalis, medial amygdaloid nucleus, lateral hypothalamic nucleus, paraventricular hypothalamic nucleus, ventromedial hypothalamic nucleus, arcuate nucleus, supramammillary nucleus, central gray at the mesencephalon, raphe dorsalis, raphe medianus, and lateral parabrachial nucleus have been confirmed. In addition, we found reciprocal connections with septo-hypothalamic nucleus, amygdalo-hipocampal nucleus, subiculum, parafascicular thalamic nucleus, posterior thalamic nucleus at the caudo-ventral subdivision, median preoptic nucleus, lateral preoptic nucleus, anterior hypothalamic nucleus, periventricular area at the caudal hypothalamic level, dorsomedial hypothalamic nucleus, posterior hypothalamic nucleus, dorsal and ventral premammillary nucleus, lateral mammillary nucleus, peripeduncular nucleus, periventricular gray, ventral tegmental area, interpeduncular nucleus, nucleus raphe pontis, nucleus raphe magnus, pedunculo-pontine tegmental nucleus, gigantocellular reticular nucleus and solitary tract nucleus. The areas which had only efferent connections from MP were accumbens, caudate putamen, ventral pallidum, substantia innominata, lateral habenular nucleus, paratenial thalamic nucleus, paraventricular thalamic nucleus, mediodorsal thalamic nucleus, reuniens thalamic nucleus, median eminence, medial mammillary nucleus, subthalamic nucleus, pars compacta of substantia nigra, oculomotor nucleus, red nucleus, laterodorsal tegmental nucleus, reticular tegmental nucleus, cuneiform nucleus, nucleus locus coeruleus, and dorsal motor nucleus of vagus among which substantia innominata and median eminence were previously reported. Efferent connections to the nucleus of Darkschewitsch, interstitial nucleus of Cajal, dorsal tegmental nucleus, ventral tegmental nucleus, vestibular nuclei, nucleus raphe obsculus were very weak or abscent in the ventral approach while they were observed in dorsal approach. Previously reported afferent connections from dorsal tegmental nucleus, cuneiform nucleus, and nucleus locus ceruleus were not detected in this study.(ABSTRACT TRUNCATED AT 400 WORDS)

A correlative quantitative study comparing the nerve fibers in the cervical sympathetic trunk and the locus of the somata from which they originate in the rat

Correlative quantitative analyses were performed on the rat comparing the number of fibers in the cervical sympathetic trunk (CST) and the horseradish peroxidase (HRP)-labeled neurons in the superior cervical sympathetic ganglion (SCG), stellate ganglion, as well as in the spinal cord. The total number of nerve fibers in the left CST was 4180 +/- 169 (mean +/- S.E.M.) among which 92 +/- 3 (mean +/- S.E.M.) were myelinated. The diameter of unmyelinated fibers was 0.68 +/- 0.22 (mean +/- S.D.) microns and showed single-peaked distribution. After the application of HRP to the proximal cut end of the CST, labeled neurons were found in the stellate ganglion as well as in the ipsilateral spinal cord from C7 to T4 segments. The total number of HRP-labeled neurons in the spinal cord was 1334 +/- 45 (mean +/- S.E.M.) with the range between 844 and 1808. Ninety-nine percent of labeled neurons were located in the intermediolateral column and in the lateral funiculus while 1% were in the intercalated region and central autonomic area. Labeled neurons were encountered only sporadically in the dorsal root ganglia (DRG) from C8 to T3 level. After the application of HRP to the distal cut end of the CST, about 200 labeled neurons were observed in the caudal part of the SCG. The present results were discussed with special reference to the organization of the CST of the rat.

Correlation of mitochondrial swelling after capsaicin treatment and substance P and somatostatin immunoreactivity in small neurons of dorsal root ganglion in the rat

Capsaicin injections into adult rats produced swelling of mitochondria in a population of small neurons of the dorsal root ganglia. Large neurons did not show the mitochondrial change. The mitochondrial swelling was confirmed to be a specific change produced by capsaicin treatment by the examination of different fixation conditions in both control and capsaicin-treated materials. Whether or not these mitochondrial changes occur in a subpopulation of small neurons which contain particular neurotransmitters was examined. The swelling of mitochondria was found in most substance P- or somatostatin-immunoreactive neurons and also in some non-immunoreactive small neurons. The present results indicate that the swelling of mitochondria is a specific change which may correlate with long-lasting desensitization of nociceptive neurons in the dorsal root ganglion after capsaicin treatment.

Distribution of substance P immunoreactive nerve terminals within the nucleus tractus solitarius of the rat

The relative density of the substance P (SP) immunoreactive nerve terminals in the subnuclei of the nucleus tractus solitarius (NTS) of the rat was examined by immunohistochemical light and electron microscopy. A higher density of SP immunoreactive terminals was observed in the medial and dorsolateral subnuclei of the rostral levels to the obex, while SP immunoreactive terminals were only sparsely found in the medial, ventrolateral and interstitial subnuclei from levels 1 mm rostral to 0.5 mm caudal to the obex. Most other parts of the NTS were moderately innervated by SP immunoreactive nerve terminals.

Projection pathways, co-existence of peptides and synaptic organization of nerve fibers in the inferior mesenteric ganglion of the guinea-pig.

SummaryThe presence of immunoreactive enkephalin, dynorphin, vasoactive intestinal polypeptide, cholecystokinin, substance P and neuropeptide Y in nerve fibers that project to the guinea-pig inferior mesenteric ganglion was analysed, after different denervation and ligation procedures. A quantitative analysis demonstrates that enkephalin- and substance P fibers reach the ganglion mainly via lumbar splanchnic and partly via intermesenteric nerves. Dynorphin-, vasoactive intestinal polypeptide- and cholecystokinin fibers reach the ganglion mainly via colonic and partly via hypogastric or intermesenteric nerves. Neuropeptide Y fibers enter via intermesenteric, lumbar splanchnic and hypogastric nerves and pass through the ganglion. Analysis of serial 0.5 μm sections tends to confirm co-existence: of dynorphin, vasoactive intestinal polypeptide and cholecystokinin in fibers projecting from the colon; of dynorphin with substance P in the lumbar splanchnic nerves; and of neuropeptide Y with substance P in the hypogastric and colonic fibers. Synaptic contacts, predominantly axodendritic, onto the ganglion cells from enkephalin-, vasoactive intestinal polypeptide-, and substance P-containing terminals were revealed by electron microscopy. Enkephalin-immunoreactive axon varicosities are filled with small, clear vesicles with a few large, cored vesicles and form asymmetric synapses; dynorphin-, vasoactive intestinal polypeptide- and cholecystokinin-immunoreactive axon varicosities are rich in large, dense-cored vesicles and form symmetric synapses.

Direct synaptic contacts of catecholamine axons on the preganglionic sympathetic neurons in the rat thoracic spinal cord

Preganglionic sympathetic neurons were labelled by retrograde transport of horseradish peroxidase, while catecholamine axon varicosities were marked by the uptake of 5-hydroxydopamine in the intermediolateral nucleus of the rat. The direct synaptic contacts from the catecholamine axons to the preganglionic sympathetic neurons were demonstrated. Catecholamine axons formed symmetric synapses.

Architecture and synaptic relationships in the intermediolateral nucleus of the thoracic spinal cord of the rat: HRP labelling, catecholamine histochemistry and electron microscopic studies

SummaryThe organization of the intermediolateral nucleus (IML) of the thoracic spinal cord was examined using glyoxylic acid-induced fluorescence histochemistry, retrograde horseradish peroxidase (HRP) labelling and electron microscopy. In serial sections of T2, it was found that the distribution of catecholamine nerve terminals was intimately related to the neuronal perikarya of IML. Potassium permanganate fixation and 5-hydroxydopamine treatment revealed small dense-cored vesicles in axon varicosities with or without synaptic specializations. A gelatinous region, composed of small diameter dendrites and unmyelinated axons, formed a narrow longitudinal bundle in the centre of the nucleus. The population of the axon varicosities in the IML was 0.17 ± 0.02/μm2 in 75 nm sections. The average size of the axon varicosities with flat synaptic vesicles was 1.44 ± 0.05 μm2 and that of varicosities with spherical vesicles was 0.97 ± 0.02 μm2. After HRP injection into the superior cervical ganglion, ipsilateral IML neurons were labelled in T1–T3 segments of the spinal cord. Axon varicosities with flat and others with spherical synaptic vesicles synapsed on the dendrites labelled by HRP. Among axon varicosities synapsing on the preganglionic sympathetic neurons, 74.8 ± 7.1% at axo-somatic synapses and 46.0 ± 6.7% at synapses on proximal dendrites contained flat synaptic vesicles.

Regeneration of the recurrent laryngeal nerve in the guinea pig: reorganization of motoneurons after freezing injury.

The purpose of this study was to clarify the morphologic changes resulting from reinnervation after a freezing injury. We chose the freezing injury as the most promising nerve regeneration model in order to examine the mechanism behind the production of misdirected reinnervation. The left recurrent laryngeal nerve of the adult guinea pig was injured by freezing (-80 degrees C) at the level of the 10th tracheal ring. At intervals ranging from 2 weeks to 6 months after the injury, horseradish peroxidase was injected into the left posterior cricoarytenoid muscle to ascertain the presence of retrograde-labeled perikarya in the medulla oblongata. Projections to the individual laryngeal muscles and to the entire recurrent laryngeal nerve served as normal controls. In addition, we observed by electron microscopy the degeneration and regeneration processes of the recurrent laryngeal nerve following injury. From 2 to 6 months after the freezing injury, the number of labeled neurons in the nucleus ambiguus increased gradually from 20 to 90. In addition, the area occupied by neurons which project to the posterior cricoarytenoid muscle was expanded, but was confined within the region of perikarya projecting to the normal recurrent laryngeal nerve. Most axons degenerated within 3 days and showed regenerative sprouting with growth cones by 7 days postinjury. Despite the fact that freezing injury preserved the basal lamina tunnel with minimal disturbance of the recurrent laryngeal nerve fiber structure, target-specific reinnervation was incomplete.

An immunohistochemical observation of polypeptides and monoamines in the nucleus preopticus medianus of the rat

A detailed regional distribution of nerve cells and terminals immunoreactive to polypeptides or monoamines was examined in the 5 subdivisions (rostral, mid-dorsal, mid-ventral, caudo-dorsal and caudo-ventral parts) of the nucleus preopticus medianus (POMe) of the rat. In general, immunoreactive nerve cells and terminals are more numerous in the ventral parts of the middle and caudal POMe. Nerve cells immunoreactive to neurotensin (NT), Met-enkephalin-Arg6-Gly7-Leu8 (mENK8) or cholecystokinin-octapeptide (CCK8) are distributed throughout the POMe, while those immunoreactive to luteinizing hormone-releasing hormone (LHRH) are found in the rostral and middle POMe. Nerve cells immunoreactive to substance P (SP) are seen in the middle and caudal POMe and those immunoreactive to somatostatin (SRIF) are scattered in the middle part of the nucleus. The densities of nerve terminals immunoreactive to neuropeptide tyrosine, mENK8, SP or noradrenaline are high throughout the POMe, while nerve terminals immunoreactive to CCK8, LHRH, NT, SRIF or vasoactive intestinal polypeptide are moderate and those immunoreactive to calcitonin gene-related peptide, serotonin or dopamine are sparse. This varied distributional pattern of immunoreactive nerve cells and terminals suggests regional differences in function within the POMe.

Formation of synaptic graft-host connections by noradrenergic locus coeruleus neurons transplanted into the adult rat hippocampus

Transplants of cell suspension obtained from the locus coeruleus region of 13- to 14-day-old rat fetuses were implanted into the hippocampal formation of intact adult rats or rats from which the noradrenergic afferents to the hippocampus had been removed by bilateral 6-hydroxydopamine (6-OHDA) injections into the dorsal tegmental noradrenergic bundle. The growth noradrenergic axons into the host hippocampus from the implant was studied at 4-8 months after surgery by immunohistochemistry using antisera raised against tyrosine hydroxylase or noradrenaline. In the animals with an intact noradrenergic system the host noradrenergic afferents were removed by bilateral dorsal bundle lesions 2 weeks before sacrifice. Fine axon-like fibers (diameter about 0.3 micron) and thick dendrite-like fibers (diameter about 1.3 micron), labeled immunohistochemically, were abundant and spread far from the graft. By electron microscopy, immunolabeled axon-like fibers formed mostly symmetrical synaptic contacts with nonlabeled spines and shafts of dendrites in the host. Labeled dendrite-like fibers of presumed graft origin penetrated deep into the host neuropil and received abundant afferents from nonlabeled axon terminals. The extent of graft-derived noradrenergic axons and the synapses established with the host hippocampal neurons were similar in the chronically denervated animals and in the animals where the intrinsic noradrenergic afferents had been left intact until 2 weeks before sacrifice. The results show that implanted embryonic noradrenergic neurons are able to innervate the hippocampus in both the presence and the absence of an intact intrinsic noradrenergic innervation and that the ingrowing axons form abundant synaptic connections with the host hippocampal neurons under both conditions. Dendritic processes from the grafted noradrenergic neurons that extend deep into the host tissue may receive a reciprocal synaptic host afferent input.