Nitric oxide synthase and interferon-gamma receptor immunoreactivities in relation to ascending spinal pathways to thalamus, hypothalamus, and the periaqueductal grey in the rat

The retrograde tracer fluoro-gold was injected into the periaqueductal grey, thalamus or hypothalamus, and spinal cord sections were processed for neuronal nitric oxide synthase (nNOS) immunohistochemistry to investigate the relationships of nNOS immunoreactive, and spinomesencephalic, spinothalamic and spinohypothalamic projection neurones. In addition, in the lateral spinal nucleus the relationship between spinomesencephalic, -thalamic and -hypothalamic projection neurones, and nNOS and interferon-gamma receptor immunoreactive structures was investigated at the lumbar level. No single retrogradely labelled spinomesencephalic, -thalamic or -hypothalamic neurone showed nNOS immunoreactivity. In the lateral spinal nucleus, however, many fluoro-gold-labelled neurones were closely apposed by both nNOS and interferon-gamma receptor immunoreactive structures, especially prominent in the hypothalamic injection cases. This study gave no evidence for nNOS immunoreactivity in spinal neurones projecting to the periaqueductal grey, thalamus or hypothalamus, but suggests that in the lateral spinal nucleus such neurones are contacted by both nNOS- and interferon-gamma receptor-containing axon terminals.     

Organization of neuropeptide tyrosine-like immunoreactive system in the brain of the Antarctic fish, Trematomus bernacchii

Antarctic notothenioids have developed unique freezing-resistance adaptations, including brain diversification, to survive in the subzero waters of the Southern Ocean surrounding Antarctica. In this study we have investigated the anatomical distribution of neuropeptide tyrosine (NPY)-like immunoreactive elements in the brain of the Antarctic fish Trematomus bernacchii, by using an antiserum raised against porcine NPY. Perikarya exhibiting NPY-like immunoreactivity were observed in distinct regions of the brain. The most rostral group of immunoreactive perikarya was found in the telencephalon, within the entopeduncular nucleus. In the diencephalon, three groups of NPY-like immunoreactive perikarya were found in the hypothalamus. Two groups of positive cell bodies were found in distinct populations of the preoptic nucleus, whereas the other group was found in the nucleus of the lateral recess. More caudally, NPY immunoreactivity was detected in large neurons located in the subependymal layers of the dorsal tegmentum of the mesencephalon, medially to the torus semicircularis. NPY-like immunoreactive nerve fibres were more widely distributed throughout the telencephalon to the rhombencephalon. High densities of nerve fibres and terminals were observed in several regions of the telencephalon, olfactory bulbs, hypothalamus, tectum of the mesencephalon and in the ventral tegmentum of the rhombencephalon. The distribution of NPY-like immunoreactive structures suggests that, in Trematomus, this peptide may be involved in the control of several brain functions, including olfactory activity, feeding behaviour, and somatosensory and visual information. In comparison with other neuropeptides previously described in the brain of Antarctic fish, NPY is more widely distributed. Our data also indicate the existence of differences in the brain distribution of NPY between Trematomus and other teleosts. In contrast with previous results reported in other fish, Trematomus contains positive fibres in the olfactory bulbs and immunoreactive perikarya in the nucleus of the lateral recess, whereas NPY-immunopositive cell bodies are absent in the thalamus and rhombencephalon, and no NPY immunoreactivity is present in the pituitary. These differences could be related to the Antarctic ecological diversity of notothenioids living at subzero temperatures.     

Comparative immunocytochemical localization of putative opioid ligands in the central nervous system

We report a detailed comparative immunocytochemical mapping of enkephalin, CCK and ACTH/beta-endorphin immunoreactive nerves in the central nervous system of rat and guinea pig. Enkephalin immunoreactivity was detected in many groups of nerve cell bodies, fibers and terminals in the limbic system, basal ganglia, hypothalamus, thalamus, brain stem and spinal cord. beta-endorphin and ACTH immunoreactivity was limited to a single group of nerve cell bodies in and around the arcuate nucleus and in fibers and terminals in the midline areas of the hypothalamus, thalamus and mesencephalic periaqueductal gray with lateral extensions to the amygdaloid area. Cholecystokinin immunoreactive nerve fibers and terminals displayed a distribution similar to that of enkephalin in many regions; but striking differences were also found. An immunocytochemical doublestaining technique, which allowed simultaneous detection of two different peptides in the same tissue section, showed that enkephalin-, CCK- and ACTH/beta-endorphin-immunoreactive nerves although closely intermingled in many brain areas, occurred separately. The distributions of nerve terminals containing these neuropeptides showed striking overlaps and also paralleled the distribution of opiate receptors. This may suggest that enkephalin, CCK, ACTH and beta-endorphin may interact with each other and with opiate receptors.     

Thiamine-like fibers in the monkey brain: an immunocytochemical study

The distribution of thiamine-immunoreactive structures was studied in the brain of the monkey using an indirect immunoperoxidase technique. Fibers containing thiamine, but no thiamine-immunoreactive cell bodies, were found. The highest density of fibers containing thiamine was observed in the pulvinar nucleus and in the region extending from the pulvinar nucleus to the caudate nucleus. In the mesencephalon, immunoreactive fibers containing thiamine were only found at rostral level close to the medial lemniscus (at the mesencephalic-diencephalic junction). In the thalamus, the distribution of thiamine-immunoreactive structures was more widespread. Thus, immunoreactive fibers were found in nuclei close to the midline (centrum medianum/parafascicular complex), in the ventrolateral thalamus (medial geniculate nucleus, inferior pulvinar nucleus), and in the dorsolateral thalamus (lateral posterior nucleus, pulvinar nucleus). Finally, in the anterior commissure and in the cerebral cortex a low density immunoreactive fibers was visualized. Thus, in the brainstem, no immunoreactive structures were visualized in the medulla oblongata, pons, or in the medial-caudal mesencephalon, and no immunoreactive fibers were observed in the cerebellum, hypothalamus and in the basal ganglia. The present report describes the first visualization and the morphological characteristics (thick, smooth and short, medium or long in length) of the thiamine-immunoreactive fibers in the primate central nervous system using an antiserum directed against this vitamin. The distribution of thiamine-immunoreactive structures in the monkey brain suggests that this vitamin could be involved in several physiological mechanisms.     

The vasotocinergic system in the hypothalamus and limbic region of the budgerigar (Melopsittacus undulatus)

We report a morphological and biochemical analysis on the presence, distribution and quantification of vasotocin in the hypothalamus and limbic region of the budgerigar Melopsittacus undulatus, using immunohistochemistry on serial sections and competitive enzyme linked immunoadsorbent assay measurements on tissue extracts. Analysis of the sections showed large vasotocin-immunoreactive neurons in three main regions of the diencephalon, of both male and female specimens. Vasotocinergic cell bodies were located in the ventral and lateral areas of the hypothalamus, dorsal to the lateral thalamus and medial to the nucleus geniculatus lateralis. Immunoreactive neurons were placed also periventricularly, close to the walls of the third ventricle, at the level of the magnocellular paraventricular nucleus. Well evident bundles of immunoreactive fibers were placed ventral to the anterior commissure in the same regions of the hypothalamus and thalamus where vasotocinergic perikarya are localized. Fibers were identified close to the third ventricle, and in the lateral hypothalamic area along the lateral forebrain bundle. In contrast to what reported for other oscine and non-oscine avian species, we were not able to identify immunopositive neurons in any region above the anterior commissure, or detect relevant differences on the distribution of the vasotocin immmunoreactivity between sexes. Competitive enzyme linked immunoadsorption assay and image analysis of the extension of immunoreactivity in the tissue sections were consistent with the qualitative observations and indicated that there is no statistically significant dimorphism in the content of vasotocin or in the location and distribution of vasotocinergic elements in the investigated areas of male and female parrot brains.     

Immunohistochemical localization of neurokinin-l receptor in the lumbar spinal cord of young rats: morphology and distribution

The type and distribution of neurokinin-1 (NK-1) receptor-expressing neurones were studied in young (14-day-old) rats' lumbar spinal cord using pre-embedding immunohistochemistry. The heaviest immunoreactivity was observed in the middle part and lateral fourth of lamina I where the great majority of immunoreactive perikarya represented fusiform and multipolar cells. In lamina II the middle and medial part showed moderate immunoreactivity, most of the cells resembled stalked cells. In lamina III the labelled perikarya were evenly distributed, while those in lamina IV accumulated mainly in the lateral part. In both laminae most of the labelled neurones represented central cells, the rest of them belonged to the antenna-type cells with long dorsally directed dendrites penetrating the superficial laminae. The immunoreactivity in laminae V-VII was uniform and relatively weak. In lamina VIII the immunopositive perikarya were encountered only rarely while in lamina IX virtually all motoneurones showed weak immunoreactivity. Lamina X contained small, multipolar and fusiform labelled perikarya. In conclusion, we found that the general appearance of the NK-1 receptor immunostaining and the major type of NK-I receptor-expressing neurones were similar to that found previously in adult spinal cord. Using the same method as Brown and colleagues the number of labelled NK- 1 receptor immunoreactive cells was similar in young and adult animals except lamina I where the number of immunoreactive neurones was twice that in adults.     

The electrophysiological study of axonal branching of the C6-C7 neurones of the cat's spinal cord projecting to the lateral reticular nucleus and the cerebellum.

Ascending projections of the cervical spinal cord neurones to the lateral reticular nucleus (LRN) and the restiform body (RB) were electrophysiologically investigated in alpha-chloralose anaesthetized cats. The main purpose of the study was to demonstrate whether the some of C6/C7 segments neurones gave off collateral branches that reached both the LRN and RB. Antidromic action potentials were recorded from 50 neurones located in laminae IVVIII of the grey matter of C6/C7 segments following stimulation of the LRN and RB with tungsten electrodes. Analysis of the antidromic responses allowed to distinguish three populations of neurones according to their projection patterns, 41% cells ascended exclusively to either one of the two structures investigated (26% to LRN and 15% to RB), while in the remaining cases (59%) collateral projections ascended to both LRN and RB. The axonal conduction velocities were comprised in the range of 14-90 m/s and no differences were found when compared mean values for particular groups of neurones. In conclusion, the results obtained indicate that in a great number of C6/C7 neurones both spinoreticular and spinocerebellar projections are present. It is likely that afferent input from forelimbs may be directly of indirectly transmitted to the cerebellum through the same neurones.     

Immunohistochemical mapping of perineuronal nets containing chondroitin unsulfate proteoglycan in the rat central nervous system

Subsets of neurons ensheathed by perineuronal nets containing chondroitin unsulfate proteoglycan have been immunohistochemically mapped throughout the rat central nervous system from the olfactory bulb to the spinal cord. A variable proportion of neurons were outlined by immunoreactivity for the monoclonal antibody (Mab 1B5), but only after chondroitinase ABC digestion. In forebrain cortical structures the only immunoreactive nets were around interneurons; in contrast, throughout the brainstem and spinal cord a large proportion of projection neurons were surrounded by intense immunoreactivity. Immunoreactivity was ordinarily found in the neuropil between neurons surrounded by an immunopositive net. By contrast, within the pyriform cortex the neuropil of the plexiform layer was intensely immunoreactive even though no perineuronal net could be found. The presence of perineuronal nets could not be correlated with any single class of neurons; however a few functionally related groups (e.g., motor and motor-related structures: motor neurons both in the spinal cord and in the efferent somatic nuclei of the brainstem, deep cerebellar nuclei, vestibular nuclei; red nucleus, reticular formation; central auditory pathway: ventral cochlear nucleus, trapezoid body, superior olive, nucleus of the lateral lemniscus, inferior colliculus, medial geniculate body) were the main components of the neuronal subpopulation displaying chondroitin unsulfate proteoglycans in the surrounding extracellular matrix. The immunodecorated neurons found in the present study and those shown by different monoclonal antibodies or by lectin cytochemisty, revealed consistent overlapping of their distribution patterns.     

Differential distribution of orexin-A and orexin-B immunoreactivity in the rat brain and spinal cord

The orexins are recently identified appetite-stimulating hypothalamic peptides. We used immunohistochemistry to map orexin-A and orexin-B immunoreactivity in rat brain, spinal cord, and some peripheral tissues. Orexin-A- and orexin-B-immunoreactive cell bodies were confined to the lateral hypothalamic area and perifornical nuclei. Orexin-A-immunoreactive fibers were densely distributed in the hypothalamus, septum, thalamus, locus coeruleus, spinal cord, and near the ventricles, but absent from peripheral sites investigated. In contrast, orexin-B-immunoreactive fibers were distributed sparsely in the hypothalamus. Orexin cells are strategically sited to contribute to feeding regulation, but their widespread projections suggest that orexins have other physiological roles.     

The distribution of GABA-like-immunoreactive neurons in the brain of the newt,Triturus cristatus carnifex,and the green frog,Rana esculenta

Summary The distribution of -aminobutyric acid (GABA) immunoreactivity was studied in the brain of two amphibian species (Triturus cristatus carnifex, Urodela; Rana esculenta, Anura) by employing a specific GABA antiserum. A noteworthy immunoreactive neuronal system was found in the telencephalic dorsal and medial pallium (primordium pallii dorsalis and primordium hippocampi) and in the olfactory bulbs. In the diencephalic habenular nuclei there was a rich GABAergic innervation, and immunoreactive neurons were observed in the dorsal thalamus. In the hypothalamus the GABA immunoreactivity was found in the preoptic area, the paraventricular organ and in the hypothalamo-hypophysial complex. In the preoptic area of the frog some GABA-immunoreactive CSF-contacting cells were shown. In the optic tectum immunolabeled neurons were present in all the cellular layers. A rich GABAergic innervation characterized both the fibrous layers of the tectum and the neuropil of the tegmentum and interpeduncular nucleus. In the cerebellum, in addition to the Purkinje cells showing a variable immunopositivity, some immunoreactive cell bodies appeared in the central grey. Abundant immunolabeled nerve fibers in the acoustico-lateral area and some immunopositive neurons in the region of the raphe nucleus were observed. In conclusion, the GABAergic central systems, well-developed in the amphibian species studied, were generally characterized by close similarities to the pattern described in mammals.Dedicated to Professor Valdo Mazzi (Dipartimento di Biologia Animale, Università di Torino), in honor of his 70th birthday     

Topographical localisation of endothelin mRNA and peptide immunoreactivity in neurones of the human brain

The distribution of endothelin mRNA and immunoreactivity in the human brain was investigated using the technique of in situ hybridization and immunocytochemistry. Cryostat sections from 22 cases of neurologically normal adult human brain, collected 3-7 h post-mortem were hybridized with 35S-labelled complementary (c)RNA probes prepared from the 3' non-coding region of endothelin-1 cDNA, and the chromosomal genes encoding endothelin-2 and -3. In situ hybridization with all three cRNA probes revealed labelled neuronal cell bodies in laminae III-VI of the parietal, temporal and frontal cortices. Labelled cells were also seen, scattered throughout the para- and periventricular, supraoptic and lateral hypothalamic nuclei, the caudate nucleus, amygdala, hippocampus, basal nucleus of Meynert, substantia nigra, raphe nuclei, Purkinje cell layer of the cerebellum and in the dorsal motor nuclei of the vagus of the medulla oblongata. The distribution of neurones immunoreactive to endothelin was similar to that of endothelin mRNA, although fewer immunoreactive cells throughout the brain, were noted. Immunoreactive fibres were present mainly in the cortex and hypothalamus, and to a lesser extent in the brain stem. Combined in situ hybridization and immunocytochemistry on the same section revealed the presence of endothelin-1 mRNA and immunoreactivity in the same cortical neuronal cell. Colocalisation studies in the cortex revealed endothelin-1 mRNA and immunoreactivity in a number of cells which also expressed neuropeptide Y mRNA and immunoreactivity. In the hypothalamus and basal nucleus of Meynert endothelin immunoreactivity was colocalised to a subset of neurophysin- and galanin-immunoreactive cell bodies respectively. Endothelin mRNA and immunoreactivity was also seen in some blood vessel endothelial cells. The findings of endothelin mRNAs and immunoreactivity in heterogenous neuronal populations further emphasises the potential role of endothelin as a neuropeptide, probably having diverse actions in the nervous system of man.     

Neuronal location of the bombesin-like immunoreactivity in the central nervous system of the rat

The immunohistochemical distribution of bombesin-like immunoreactivity in the central nervous system of the rat was revealed using a rabbit antibody against [Glu⁷]bombesin(6–14). In radioimmunoassay, the antibody had minimal cross reactivity with substance P thereby enhancing the significance of histochemical controls proving that the immunoreactivity detected was related to bombesin but not to substance P. Bombesin-immunoreactive neurons were detected in several brain structures including the hypothalamus, interpeduncular nucleus, central grey, dorsolateral tegmental nucleus, dorsal parabrachial nucleus, nucleus of the solitary tract and trigeminal complex. In the spinal cord, intense immunoreactivity was found in the superficial layers of the posterior horn. Since in this area the reaction diminished after rhizotomy the location of the peptide in afferent neurons was considered. In the anterior horn the bombesin-like immunoreactivity located in nerve terminal-like structures was unchanged after rhizotomy suggesting that the cell bodies were located in CNS.     

New data on the immunocytochemical localization of thyrotropin-releasing hormone in the rat central nervous system

An antiserum raised against the synthetic tripeptide pyroglutamyl-histidyl-proline (free acid) was used to localize thyrotropin-releasing hormone (TRH) in the rat central nervous system (CNS) by immunocytochemistry. The distribution of TRH-immunoreactive structures was similar to that reported earlier; i.e., most of the TRH-containing perikarya were located in the parvicellular part of the hypothalamic paraventricular nucleus, the suprachiasmatic portion of the preoptic nucleus, the dorsomedial nucleus, the lateral basal hypothalamus, and the raphe nuclei. Several new locations for TRH-immunoreactive neurons were also observed, including the glomerular layer of the olfactory bulb, the anterior olfactory nuclei, the diagonal band of Broca, the septal nuclei, the sexually dimorphic nucleus of the preoptic area, the reticular thalamic nucleus, the lateral reticular nucleus of the medulla oblongata, and the central gray matter of the mesencephalon. Immunoreactive fibers were seen in the median eminence, the organum vasculosum of the lamina terminalis, the lateral septal nucleus, the medial habenula, the dorsal and ventral parabrachial nuclei, the nucleus of the solitary tract, around the motor nuclei of the cranial nerves, the dorsal vagal complex, and in the reticular formation of the brainstem. In the spinal cord, no immunoreactive perikarya were observed. Immunoreactive processes were present in the lateral funiculus of the white matter and in laminae V-X in the gray matter. Dense terminal-like structures were seen around spinal motor neurons. The distribution of TRH-immunoreactive structures in the CNS suggests that TRH functions both as a neuroendocrine regulator in the hypothalamus and as a neurotransmitter or neuromodulator throughout the CNS.     

Alpha-melanocyte-stimulating hormone (alpha-MSH) in the brain of the cartilagenous fish. Immunohistochemical localization and biochemical characterization

The distribution of immunoreactive alpha-melanocyte-stimulating hormone (alpha-MSH) in the central nervous system and pituitary of the elasmobranch fish Scyliorhinus canicula was determined by the indirect immunofluorescence and the peroxidase-antiperoxidase methods using a highly specific antiserum. Perikarya containing alpha-MSH-like immunoreactivity were localized in the dorsal portion of the posterior hypothalamus, mainly in the tuberculus posterioris and sacci vasculosus nuclei. Immunoreactive alpha-MSH cell bodies were found in the dorsal wall and ventral region of the caudal part of the tuberculum posterioris. These structures were densely innervated by fine beaded immunoreactive fibers. Some alpha-MSH immunoreactive cells were occasionally detected in the ventral part of the nucleus periventricularis. Scattered cell bodies and fibers were also observed in the dorsal wall of the posterior recess. Outside the hypothalamus very few fibers were detected in the dorsal thalamus and mesencephalon. No immunoreactivity was found in any other parts of the brain. The alpha-MSH immunoreactive material localized in the brain was characterized by combining high-performance liquid chromatography (HPLC) analysis and radioimmunological detection. Brain and pituitary extracts exhibited displacement curves which were parallel to that obtained with synthetic alpha-MSH. The concentrations of alpha-MSH immunoreactive material were determined in 5 different regions of the brain. The highest concentration was found in the hypothalamus. HPLC analysis resolved two major forms of immunoreactive alpha-MSH in the hypothalamus, which had been same retention times as des-N alpha-acetyl-alpha-MSH and its sulfoxide derivative. These results provide the first evidence for the presence of alpha-MSH-like peptides in the fish brain.(ABSTRACT TRUNCATED AT 250 WORDS)     

Distribution of FMRFamide-like immunoreactivity in the forebrain of the catfish, Clarias batrachus (Linn.).

The anatomical distribution of FMRFamide-like immunoreactivity in the forebrain and pituitary of the catfish, Clarias batrachus, was investigated. Immunoreactive cells were observed in the ganglion cells of the nervus terminalis (NT) and in the medial olfactory tracts. In the preoptic area, FMRFamide-containing perikarya were restricted to the lateral preoptic area, paraventricular subdivision of the nucleus preopticus, nucleus suprachiasmaticus and nucleus preopticus periventricularis posterior. In the postoptic area, some cells of the nucleus postopticus lateralis and nucleus of the horizontal commissure showed moderate immunoreactivity. In the tuberal area, immunoreactivity was observed in few cells of the nucleus hypothalamicus ventralis and nucleus arcuatus hypothalamicus (NAH). Nucleus ventromedialis thalami was the only thalamic nucleus with FMRFamide immunoreactivity. Immunoreactive processes were traceable from the NT through the medial as well as lateral olfactory tracts into the telencephalon and the area ventralis telencephali pars supracommissuralis (Vs). Further caudally, the immunoreactive fibers could be traced into discrete areas, including habenular and posterior commissures, neurohypophysis and pituitary; isolated fibers were also observed in the pineal stalk. A loose network of immunoreactive processes was observed in the olfactory bulbs and the entire telencephalon, with higher densities in some areas, including Vs. A dense plexus of immunoreactive fibers was seen in the pre- and postoptic areas and around the paraventricular organ, while relatively few were observed in the thalamus. A high concentration of fiber terminals was found in the caudal tuberal area.     

Distribution and characterization of endozepine-like immunoreactivity in the central nervous system of the frog Rana ridibunda.

The localization of endozepine-like immunoreactivity in the brain of the frog Rana ridibunda was investigated by indirect immunofluorescence, using an antiserum against synthetic rat octadecaneuropeptide (ODN). A specific immunoreaction was detected in ependymal cells lining the ventricular system of the brain and in circumventricular organs. Numerous immunoreactive cells were found covering the walls of the lateral ventricles in the telencephalon, as well as in the diencephalic and mesencephalic ventricles. In the hypothalamus, both the preoptic nucleus and the infundibular region showed numerous immunopositive cells. Ependymal cells lining the rhomboencephalic fourth ventricle and the central canal of the spinal cord were also immunoreactive. The concentration of endozepine-like immunoreactivity was measured in various regions of the brain using a sensitive and specific radioimmunoassay for rat ODN. The highest levels of ODN-like immunoreactivity were found in the infundibulum, cerebellum and preoptic area. Reverse phase high performance liquid chromatography and radioimmunoassay quantification were used to characterize endozepines in the frog brain. The elution profiles of the different brain regions revealed four major immunoreactive peaks. The present results demonstrate the presence of peptides immunologically related to the endozepine family in the central nervous system of the frog. The localization of immunoreactive endozepines in ependymal cells suggests that these peptides play important neuromodulatory functions in the amphibian brain.     

Glucagon-related peptides in the rat hypothalamus

Summary Immunohistochemically, nerve fibers and terminals reacting with anti-N-terminal-specific but not with anti-C-terminal-specific glucagon antiserum were observed in the following rat hypothalamic regions: paraventricular nucleus, supraoptic nucleus, anterior hypothalamus, arcuate nucleus, ventromedial hypothalamic nucleus and median eminence. Few fibers and terminals were demonstrated in the lateral hypothalamic area and dorsomedial hypothalamic nucleus. Radioimmunoassay data indicated that the concentration of gut glucagon-like immunoreactivity was higher in the ventromedial nucleus than in the lateral hypothalamic area. In food-deprived conditions, this concentration increased in both these parts. This was also verified in immunostained preparations in which a marked enhancement of gut glucagon-like immunoreactivity-containing fibers and terminals was observed in many hypothalamic regions. Several immunoreactive cell bodies were found in the ventromedial and arcuate nuclei of starved rats. Both biochemical and morphological data suggest that glucagon-related peptides may act as neurotransmitters or neuromodulators in the hypothalamus and may be involved in the central regulatory mechanism related to feeding behavior and energy metabolism.     

Choline acetyltransferase immunocytochemical staining of the rat supraoptic nucleus and its surroundings

Summary Two different monoclonal antibodies raised against choline acetyltransferase were used, together with preembedding immunocytochemical techniques, to visualize the possible cholinergic innervation of the supraoptic and paraventricular nuclei of the rat hypothalamus. Light microscopy confirmed the presence of a group of bipolar and multipolar immunoreactive neurones in the hypothalamus dorsolateral to the supraoptic nucleus as well as numerous immunopositive fibers. Electron microscopy showed that the immunopositive cell bodies contained the usual perikaryal organelles while most immunoreactive fibers appeared dendritic; immunonegative terminals made synaptic contact onto these profiles. Immunopositive terminals making synaptic contact onto dendritic profiles were also noted in this area. In contrast, light microscopy showed no immunoreactivity to choline acetyltransferase in the magnocellular nuclei themselves. Electron microscopy revealed some immunopositive profiles along the boundaries of both nuclei, along the optic chiasm adjacent to the supraoptic nucleus and in the ventral glial lamina but not within the nuclei proper. Surprisingly, these immunopositive profiles appeared dendritic and were often contacted by one or more immunonegative synapses. Our observations thus indicate that cell bodies and dendrites in the supraoptic and paraventricular nuclei are not directly innervated by cholinergic synapses. The functional significance of the putative cholinergic dendrites in close proximity to magnocellular neurones remains to be determined.     

Demonstration of a different localization of perikarya immunoreactive to oxytocin and vasopressin in the cat hypothalamus

Using immunohistochemical techniques, we demonstrated oxytocin (OT) and vasopressin (AVP) neurons in the cat hypothalamus. The OT immunoreactive neurons were found mainly in the paraventricular nucleus, supraoptic nucleus and dorsal accessory group located lateral to the fornix. In addition to these hypothalamic structures, the AVP immunoreactive neurons were observed in the suprachiasmatic nucleus, ventral accessory group located in the retrochiasmatic area and lateral accessory group, dorsal to the supraoptic nucleus caudally, and ventral to the medial part of the internal capsule rostrally. We further demonstrated a different localization of the OT and AVP immunoreactive neurons in the paraventricular and supraoptic nuclei.     

Molecular characterization of the human VIP prohormone

Two distinct cell groups contain α-melanocyte-stimulating hormone-like immunoreactivity in rat hypothalamus. Only one group, located in the arcuate nucleus, contains other opiocortin peptide immunoreactivity. Combined immunocytochemistry, radioimmunoassay and high performance liquid chromatography, using two different antisera, were used in an attempt to characterise the immunoreactive material present in each cell group. The results thus obtained from normal rats, using an antiserum against α-melanocyte-stimulating hormone and one against COOH-terminal adrenocorticotropin, were compared with those obtained from rats treated neonatally with monosodium glutamate, which destroys the arcuate nucleus.In animals treated with monosodium glutamate, cells of the arcuate nucleus, staining with both antisera, were reduced in number. Cells containing only α-melanocyte-stimulating hormone-like immunoreactivity in the lateral hypothalamus were unaffected. Peptide levels detected by radioimmunoassay with both antisera were reduced in parallel. Chromatographed extracts showed parallel reductions in α-melanocyte-stimulating hormone-like and COOH-terminal adrenocorticotropin-like immunoreactivities.These results suggest that if the immunostained cells of the lateral hypothalamus contain conventional α-melanocyte-stimulating hormone, it constitutes only a very small proportion of the total hypothalamic concentration. However, the possibilities that the antiserum is crossreacting with a different molecular species, or with a similar compound synthesised by a different pathway cannot be excluded.