Thyrotropin-releasing hormone: regional distribution in rat brain.

A sensitive and specific radioimmnunoassay has been used to measure the distribution of thyrotropin-releasing hormone (TRH) in rat brain. All areas of brain tested, except cerebellum, contained readily measurable amounts of TRH. The hypothalamus contained only 31.2 percent of the total brain content of TRH. These results support recent suggestions of central actions for TRH in addition to its hypophysiotropic functions.

Nuclear localization of histamine in neonatal rat brain.

The concentration of histamine in the brains of neonatal rats is considerably higher than that in adults. Subcellular fractionation studies revealed that about 90 percent of the histamine content of neonatal rat brain is confined to the crude nuclear fraction obtained by differential fractionation. Purified nuclei prepared from these fractions retained 90 percent of their histamine content. The nuclear localization of histamine in the brains of neonatal rats suggests a function for histamine in modulating the growth processes of the neonatal brain.

A major direct GABAergic pathway from zona incerta to neocortex.

Retrograde fluorescent tracers were used to demonstrate a previously unknown but sizable direct gamma-aminobutyric acid (GABA)-containing neuronal pathway from the zona incerta to the neocortex in rats. This incertocortical pathway was found to project bilaterally to the entire neocortex and exhibited a rough corticotopic organization. Many of the zona incerta neurons projecting to the parietal and occipital cortices could also be immunohistochemically stained with antibodies to glutamic acid decarboxylase and GABA. Few of these neurons were immunoreactive to tyrosine hydroxylase antibodies, which identify dopamine-containing neurons. Injections in the frontal and entorhinal cortices labeled many neurons near or within the dopaminergic A13 subdivision of the zona incerta. In addition, the incertocortical system was found to be significantly larger during early postnatal (2 to 3 weeks) development. The projection pattern of this newly discovered pathway resembles that of the monoaminergic and cholinergic systems, arising from the brainstem and forebrain, suggesting possible similarities of function.

Immunolocalization of the thyrotropin-releasing hormone prohormone in the rat central nervous system.

The distribution of immunoreactive TRH prohormone in the rat central nervous system was studied by immunocytochemistry using an antiserum raised against a synthetic decapeptide hypothesized to represent a portion of the mammalian TRH precursor protein. Reaction product was identified in several regions of the brain in a distribution typical of that previously described for the tripeptide. In contrast to TRH, however, immunoreactive pro-TRH was largely confined to neuronal perikarya and only rarely seen in axons or axon terminals. In addition, immunoreactive pro-TRH was present in portions of the telencephalon and brainstem where TRH has not previously been described in neurons by immunocytochemistry. These studies indicate that in most regions of the brain the TRH prohormone is rapidly processed within the cell soma and not during axonal transport, and raise the possibility that in certain regions of the brain processing of the prohormone may be to non-TRH peptides, which may be of biological importance.

The regional distribution of somatostatin in the rat brain.

A sensitive and specific radioimmunoassay for somatostatin (SRIF) has been used to determine the regional distribution of SRIF in rat brain. The hypothalamus contained the highest concentration of SRIF. Lower, but significant amounts of SRIF were present outside of the hypothalalmus. Within the hypothalamus, the concentration of SRIF was highest in the median eminence and arcuate nucleus although all of the hypothalmic nuclei contained some fo this material. The implications of this distribution are discussed.

Immunohistochemical localization of adrenocorticotropin in the rat brain.

ACTH and some of its fragments have been shown to play a role in a variety of adaptative mechanisms. To clearly identify the nervous structures containing ACTH in the rat brain, an immunohistochemical localization of this peptide was conducted at both light and electron microscopic levels. Nervous fibers staining for ACTH were found to be largely distributed throughout regions of hypothalamus, thalamus, and midbrain. Positive fibers could also be occasionally observed in the spinal cord. Immunostained neuronal cell bodies were only detected in the arcuate nucleus. Essentially, the same results were obtained 2 and 8 weeks after hypophysectomy. In animals pretreated with colchicine, the intensity in the staining of cell bodies was markedly increased, making possible the detection of a larger number of cell bodies. At the electron microscopic level, it was demonstrated that ACTH is contained in dense core vesicles present in nervous fibers and endings. These results indicate that ACTH of nonpituitary origin is synthesized in the central nervous system and could probably be considered as a neurotransmitter of still undefined function.

Localization of cells containing LHRH-like mRNA in rat forebrain using in situ hybridization.

Using in situ hybridization, we localized cells in the rat forebrain which contain mRNA that hybridizes with a radiolabeled, synthetic oligodeoxyribonucleotide (59-mer) complementary to human LHRH mRNA in the region which includes the coding sequence for the decapeptide. These brain areas have been shown previously to contain immunoreactive LHRH cell bodies.

Localization of thyrotropin-releasing hormone prohormone messenger ribonucleic acid in rat brain in situ hybridization.

We studied the distribution of pro- TRH mRNA in rat brain by in situ hybridization histochemistry using radiolabeled single stranded cRNA probes to confirm the hypothesis that the TRH precursor is distributed beyond regions that contain immunoreactive TRH. All regions of the central nervous system previously recognized to contain TRH showed hybridization. Hypophysiotropic neurons in the medial parvocellular division of the paraventricular nucleus showed more intense hybridization than anterior parvocellular division cells, suggesting regional differences in expression. In addition, regions not previously recognized to contain TRH in neuronal perikarya by immunocytochemistry showed specific hybridization for pro-TRH mRNA. These include cells in the olfactory bulbs, dorsal motor nucleus of the vagus, ventrolateral periaqueductal gray, reticular nucleus of the thalamus, and anterior commissural nucleus. Only a single hybridizing band was observed on Northern blots of RNA extracts of the periaqueductal gray and reticular nucleus, identical to that seen in extracts of the paraventricular nucleus. The appearance of pro-TRH mRNA in neurons not previously recognized to contain TRH but which contain the prohormone suggests that non-TRH peptides within the TRH precursor may be preferentially expressed in certain regions of the brain.

Angiotensinogen levels in the brain and cerebrospinal fluid of the genetically hypertensive rat.

The present experiments were designed to document changes in the regional distribution of angiotensinogen in the rat brain with the development of hypertension in spontaneously hypertensive rats (SHR) relative to age-matched normotensive Wistar-Kyoto rats (WKY). Levels of angiotensinogen were measured in discrete brain nuclei and cerebrospinal fluid from rats at 4, 7, and 16 weeks of age and in cerebrospinal fluid obtained by cisternal puncture at 7 and 16 weeks. Age-dependent changes in angiotensinogen were found, with levels higher in both strains at 4 weeks of age compared with 7 or 16 weeks. In contrast, plasma levels of angiotensinogen were essentially the inverse of the brain levels, low at 4 weeks and higher at 7 and 16 weeks. Levels in a number of regions adjacent to the rostral third ventricle from the 4-week-old SHR (prehypertensive phase) were significantly elevated relative to the WKY (p less than 0.05), while levels in the amygdala and posterior hypothalamus were significantly lower in the SHR (p less than 0.05). In 7-week-old rats (evolving phase), levels in nine brain regions were significantly elevated in the SHR relative to the WKY and included the nucleus tractus solitarii (p less than 0.01). Unlike the prehypertensive and evolving phases, in 16-week-old rats (maintenance phase) only two brain areas, the nucleus of the diagonal band and the lateral hypothalamus, had significantly elevated levels in the SHR (p less than 0.05). Cerebrospinal fluid levels of angiotensinogen did not correlate well with brain levels of angiotensinogen.(ABSTRACT TRUNCATED AT 250 WORDS)

Distribution of serotonin 5-HT1C receptor mRNA in adult rat brain.

Based on in situ hybridization histochemistry (ISHH), we describe the anatomical distribution of the serotonin 5-HT1C receptor mRNA. In addition to the very high levels in epithelial cells of the choroid plexus, 5-HT1C receptor mRNA is found throughout the limbic system, in catecholaminergic cells and in serotonergic neurons. Receptor transcripts are also present in the hypothalamus, numerous motor nuclei and the subthalamus. Our results correlate well with serotonin (5-HT) innervation and receptor binding. Receptor mRNA is present in many brain structures in addition to regions previously shown to have 5-HT1C receptor binding. The distribution of this receptor mRNA suggests that the 5-HT1C receptor may mediate a number of the central effects of 5-HT.

The distribution of substance P immunoreactive fibers in the rat central nervous system.

A detailed account of the distribution of immunoreactive substance P-containing structures in the rat central nervous system is presented, from results obtained by applying an indirect immunofluorescent technique. High densities of substance P-containing nerve terminals were present in sensory nuclei and other non-sensory structures such as thalamus, hypothalamus and extrapyramidal system. Substance P-reactive neuron cell bodies were present in spinal root ganglia, nucleus habenulae medialis, nucleus interpeduncularis, caudoputamen and globus pallidus. Most of the neocortex and the cerebellar cortices had no substance P-positive elements. The results support the hypothesis that substance P may be a widespread neurotransmitter in the central nervous system.

Distribution of serotonin in the brain of the mormyrid teleost Gnathonemus petersii.

The distribution of serotonin-immunoreactive neurons and fibers was studied in the highly developed brain of the weakly electric fish Gnathonemus petersii with the aid of specific antibodies against serotonin. Serotoninergic cell bodies occur in three regions: the raphe region of the brainstem, the hypothalamus, and the transition zone between the dorsal thalamus and the pretectum. Serotoninergic raphe neurons are clustered in three groups: nucleus raphes superior, intermedius, and inferior. The latter has not been described in other teleosts and thus might be the source of the serotoninergic innervation of specific mormyrid electrosensory brain regions. Most hypothalamic serotoninergic neurons have cerebrospinal-fluid (CSF)-contacting processes and thus belong to the paraventricular organ (PVO), which in Gnathonemus is located around a number of small infundibular recesses. The distribution of serotonin in the PVO precisely matches the distribution of dopamine, as described previously. Serotoninergic cells in the thalamopretectal transition zone also have been described in other teleosts, but not in other vertebrate groups, and thus seem to represent a teleostean specialization. Serotoninergic fiber density is especially high in the medial forebrain bundle and surrounding preoptic and hypothalamic regions as well as in several telencephalic and preoptic subependymal plexus. Serotoninergic fibers appear to be almost completely absent in the large and differentiated corpus and valvula cerebelli. Comparison with the literature on teleostean serotoninergic innervation patterns reveals several mormyrid specializations, including the absence of serotonin in large parts of the mormyrid telencephalic lobes, a differentiated innervation pattern of distinct electrosensory and mechanosensory subnuclei of the torus semicircularis, a refined serotoninergic lamination pattern in the midbrain tectum, and a prominent innervation of the electrosensory lateral line lobe, the associated caudal cerebellar lobe, and the electromotor medullary relay nucleus. A distinct innervation of several types of (pre)motor neurons, such as the Mauthner cells and facial motor neurons, has not been reported previously for other teleosts. Consequently, the distribution of serotoninergic fibers as well as neurons in the mormyrid brain is substantially adapted to the high degree of differentiation of its electrosensory and telencephalic brain regions, but serotoninergic innervation is not involved in the circuitry of the most impressive part of the mormyrid brain; i.e., its large corpus and valvula cerebelli.

The distribution of dopamine immunoreactivity in the forebrain and midbrain of the lizard Gekko gecko: an immunohistochemical study with antibodies against dopamine.

The distribution of dopamine (DA) immunoreactivity in the forebrain and the midbrain of the lizard Gekko gecko was studied by using recently developed antibodies against DA. Dopamine-containing cells were found around the glomeruli of the olfactory bulb, in several parts of the periventricular hypothalamic nucleus, in the periventricular organ, the ependymal wall of the infundibular recess, the lateral hypothalamic area and the pretectal posterodorsal nucleus of the diencephalon, and in the ventral tegmental area, the substantia nigra, and the presumed reptilian equivalent of the mammalian A8 cell group of the mesencephalon. Dopaminergic fibers and terminals were observed throughout the whole brain, but particularly in the diencephalon and the telencephalon. The nucleus accumbens appears to have the most dense innervation, but also the striatum, amygdaloid complex, olfactory tubercle, septum, and dorsal ventricular ridge (especially its superficial zone) show numerous DA-containing fibers and terminals. Except for the lateral cortex, cortical areas are not densely innervated by DA fibers. In several respects DA distribution in the gekkonid brain differs from that in other reptiles studied. For instance, in the Gekko the dorsal ventricular ridge is densely innervated by DA fibers, whereas in turtles and crocodiles the same structure shows only weak catecholaminergic histofluorescence. When compared to the distribution of DA immunoreactivity in mammals, it appears that the DA system in the gekkonid telencephalon resembles the distribution of DA in the limbic forebrain and striatum of mammals. Whether these similarities in distribution of DA also imply similarities in function will be discussed.

Central somatostatin systems revealed with monoclonal antibodies.

The distribution of central neurons displaying somatostatin immunoreactivity was studied using three monoclonal antibodies to cyclic somatostatin. The sensitive ABC immunoperoxidase technique was employed. A large number of positive cell groups including many previously undescribed populations were detected throughout the brain and spinal cord. Telencephalic somatostatin neurons included periglomerular cells in the olfactory bulb, mitral cells in the accessory olfactory bulb, and multipolar cells in the anterior olfactory nuclei, neocortex, amygdala, hippocampus, lateral septum, striatum, and nucleus accumbens. Within the hypothalamus, positive neurons were found in the periventricular, suprachiasmatic, and arcuate nuclei, and throughout the anterior and lateral hypothalamus. The entopeduncular nucleus and zona incerta contained many positive neurons, and the lateral habenula had a dense terminal field suggesting a pallidohabenula somatostatin pathway. Somatostatin neurons were also found in association with many sensory systems. Positive cells were present in the superior and inferior colliculi, the ventral cochlear nuclei, the ventral nucleus of the lateral lemniscus, nucleus cuneatus, nucleus gracilus, and the substantia gelatinosa. Various cerebellar circuits also displayed somatostatin immunoreactivity. Golgi cells throughout the cerebellar cortex were intensely stained, and some Purkinje cells in the paraflocculus also showed a positive reaction. Positive fibers were present in the granular layer and large varicose fibers were present in the inferior cerebellar peduncle. Many nuclei known to project to the cerebellum, including the nucleus reticularis tegmenti pontis, the medial accessory inferior olive, the nucleus prepositus hypoglossi, and many areas of the reticular formation contained positive neurons. These studies demonstrate that these new monoclonal antibodies are of great value for the study of central somatostatin systems. Previously described somatostatin systems are readily detected with these antibodies, and in addition, many otherwise unrecognized somatostatin cell groups have been discovered.

Distribution and morphology of vasopressin-, neurophysin II-, and oxytocin-immunoreactive cell bodies in the forebrain of the cat.

Experiments were done to provide a detailed map of the location and a description of morphological characteristics of vasopressin (AVP-IR)-, neurophysin II (NII-IR)- and oxytocin (OXY-IR)-immunoreactive neuronal perikarya in the forebrain of the cat. In addition, the location of cells in the forebrain retrogradely labeled following injections of tracers into the neurohypophysis was determined. The distribution of AVP-IR and NII-IR was similar in all cases studied. Most of the cells containing AVP-IR and OXY-IR were observed in the hypothalamic paraventricular (PVH) and supraoptic (SON) nuclei. In addition, AVP-IR and OXY-IR cell bodies were found in the regions of the nucleus of the diagonal band of Broca, the dorsal chiasmatic nucleus, the anterior hypothalamic-preoptic area, the periventricular area, the nucleus circularis, the perifornical area of the lateral hypothalamus, the accessory SON, the area of the tuber cinereum (Tca), and the medial nucleus of the amygdala. The density of AVP-IR cells was greater than that of OXY-IR cells in these regions. Several forebrain areas were also observed to contain only AVP-IR perikarya: the suprachiasmatic nucleus (Sc), the bed nucleus of the stria terminalis, and the region of the substantia innominata and ventral globus pallidus (SI/GP). In addition, the dorsomedial nucleus of the hypothalamus only contained OXY-IR perikarya. Most of the cells immunoreactive to AVP were multipolar and had spinelike processes over their somata and proximal dendrites. In addition, the majority of cells in the PVH and SON were round or oval, whereas those outside these nuclei were fusiform or triangular. The mean somal area of AVP-IR cells in the region of the SI/GP was significantly (P less than 0.05) larger than that of AVP-IR cells in all other regions examined, whereas the mean somal area of Sc AVP-IR cells was significantly (P less than 0.05) smaller than that of all other groups of AVP-IR cells examined. Most OXY-IR cells were similar morphologically to those immunoreactive to AVP, except that OXY-IR cell bodies and their appendages did not have spinelike processes. In addition, OXY-IR perikarya were generally of uniform size. OXY-IR cells in the PVH and accessory SON were significantly (P less than 0.05) larger than AVP-IR cells in the same regions, but were not different from AVP-IR cells in the lateral hypothalamus and SON.(ABSTRACT TRUNCATED AT 400 WORDS)

Organization of atriopeptin-like immunoreactive neurons in the central nervous system of the rat.

The atrial natriuretic peptide, atriopeptin, is a circulating hormone that plays an important role in the regulation of fluid and electrolyte homeostasis. Several recent studies have shown that atriopeptin-like immunoreactivity is present within the central nervous system as well as peripheral tissues. In the present report, we describe in detail the organization of atriopeptin-like immunoreactive (APir) perikarya and fibers in the central nervous system of the rat. The most prominent collection of APir perikarya was found in the hypothalamus, adjacent to the anteroventral tip of the third ventricle. Additional groups of APir perikarya were observed along the wall of the third ventricle and in the paraventricular and arcuate nuclei. Separate, smaller groups with distinctive morphology were seen in the lateral hypothalamic area, in the supra-mammillary, medial, and lateral mammillary nuclei, medial habenular nucleus, bed nucleus of the stria terminalis, and the central nucleus of the amygdala. In the pons and brain-stem, APir neurons were observed in the pedunculopontine and laterodorsal tegmental nuclei, as well as in the ventral tegmental area, Barrington's nucleus, the parabrachial nucleus, and the nucleus of the solitary tract. The densest terminal fields of APir fibers were found in the paraventricular nucleus of the hypothalamus, the bed nucleus of the stria terminalis, the median eminence, and the interpeduncular nucleus. The presence of atriopeptin immunoreactivity within the central nervous system suggests that atriopeptin may function as a central neuromediator. Potential functions of this candidate neuromediator deduced from its anatomical distribution are discussed, including the possibility that atriopeptin may function as both a central neuromediator and a systemic hormone in the regulation of the cardiovascular system.

Immunocytochemical localization of serotonin-containing neurons in the myelencephalon, brainstem and diencephalon of the sheep.

Using immunocytochemistry, morphological characteristics and distribution of serotonin-containing neurons and fibers of the sheep myelencephalon, brainstem and diencephalon were studied, employing highly specific antibodies to serotonin. The immunocytochemical procedure described here allowed the visualization of endogenous, and thus presumably physiological, pools of serotonin, because no pharmacological treatments (colchicine, inhibitors of monoamine oxidase or 5-hydroxytryptophan) were used to increase the endogenous amount of antigen. The distribution of serotonin cell bodies observed in the study is in agreement with that described by other authors in the rat using a similar method. The present work also shows more numerous groups than the formaldehyde-induced fluorescence method, because five additional groups were revealed, designated S1 to S5. Compared with those in the rat, sheep serotonergic structures exhibit striking specific characteristics: (1) greater scattering of cell bodies within the different groups visualized, (2) absence of group B4 and hypothalamic groups, (3) only a weak serotonergic innervation of the suprachiasmatic nuclei area.

Ultrastructural localization and afferent sources of substance P in the rat parabrachial region.

The ultrastructural morphology and afferent sources of terminals containing substance P-like immunoreactivity were examined in the rat parabrachial region. In the first portion of the study, a polyclonal antiserum to substance P was localized in the ventrolateral parabrachial region using the peroxidase-antiperoxidase labeling technique combined with electron microscopy. The antiserum was tested for cross-reaction with substance P, physalaemin, substance K and neuromedins B, C and K. Cross-reactivity was most intense with substance P. However, substance K, neuromedin K and physalaemin also exhibited limited cross-reactions with the antiserum. In the ventrolateral parabrachial region of untreated adult animals, substance P-like immunoreactivity was localized in axon terminals containing numerous small (40-60 nm) clear vesicle and 1-3 large (90-120 nm) dense-core vesicles. At least 54% of the labeled terminals formed asymmetric synapses with unlabeled dendrites; and at least 30% of the recipient dendrites received more than one labeled axon terminal. In addition, the labeled terminals were associated less frequently with other unlabeled soma, axon terminals and blood vessels. In the second part of the study, we examined whether or not perikarya in various extrinsic regions contributed to the substance P-like immunoreactivity in axon terminals in the parabrachial region. Wheat-germ agglutinin conjugated horseradish peroxidase was injected unilaterally into the parabrachial region of adult rats two days prior to being killed and one day prior to intraventricular injection of colchicine (100 micrograms in 7.5 microliter saline) which enhanced the detection of immunoreactivity in perikarya. Sections were first processed by a tetramethylbenzidine reaction stabilized with cobalt-diaminobenzidine for demonstration of the transported peroxidase then were immunocytochemically labeled for substance P. Perikarya containing both the black granular retrograde labeling and brown peroxidase-immunoreactivity were found in the nuclei of the solitary tracts, the caudal ventrolateral reticular formation, the lateral dorsal tegmental nucleus and the paraventricular, dorsomedial and lateral hypothalamic nuclei. The projections were largely, but not exclusively, from perikarya located on the same side as the parabrachial injection. We conclude that substance P, or a closely related tachykinin, is a putative transmitter or modulator within a number of pathways to the parabrachial region and that these afferents act primarily through axodendritic synapses with intrinsic neurons.

Substance P in the human brain.

The distribution of substance P immunoreactive sites was investigated by immunoenzymatic methods in a large series of paraffin embedded human brain sections from the collection assembled by Oscar and Cécile Vogt several decades ago, as well as from more recent post-mortem material. These studies demonstrated that substance P immunoreactivity was preserved in archival material permitting a detailed account of the localization of immunoreactive cell bodies, fibre networks and tracts in the human brain. Previous observations made on experimental animals and man were confirmed and extended. Additionally, substance P immunoreactive cell bodies were seen in most cortical areas and novel features were noted in the distribution of substance P-containing elements in the tuberal region, corpus striatum, substantia nigra (particularly in relationship to blood vessels) and in association with melanin-containing cells. Reconstruction of some substance P pathways was attempted by the analysis of semi-serial sections in more than one plane. Immunocytochemistry, in combination with image analysis, enabled some measurements of the differential concentrations of substance P immunoreactive material to be made and allowed a close correlation of this with defined anatomical landmarks or enkephalin immunoreactive sites.

Distribution of neuropeptide Y-like immunoreactivity in the rat central nervous system--II. Immunohistochemical analysis.

The distribution of neuropeptide Y-like immunoreactivity in the rat brain and spinal cord was investigated by means of the peroxidase-antiperoxidase procedure of Sternberger using a rabbit anti-neuropeptide Y serum. A widespread distribution of immunostained cells and fibres was detected with moderate to large numbers of cells in the following regions: olfactory bulb, anterior olfactory nucleus, olfactory tubercle, striatum, nucleus accumbens, all parts of the neocortex and the corpus callosum, septum including the anterior hippocampal rudiment, ventral pallidum, horizontal limb of the diagonal band, amygdaloid complex. Ammon's horn, dentate gyrus, subiculum, pre- and parasubiculum, lateral thalamic nucleus (intergeniculate leaflet), bed nucleus of the stria terminalis, medial preoptic area, lateral hypothalamus, mediobasal hypothalamus, supramammillary nucleus, pericentral and external nuclei of the inferior colliculus, interpeduncular nucleus, periaqueductal central gray, locus coeruleus, dorsal tegmental nucleus of Gudden, lateral superior olive, lateral reticular nucleus, medial longitudinal fasciculus, prepositus hypoglossal nucleus, nucleus of the solitary tract and spinal nucleus of the trigeminal nerve. In the spinal cord cells were found in the substantia gelatinosa at all levels, the dorsolateral funiculus and dorsal gray commissure in lumbosacral cord. The pattern of staining was found to be similar to that observed with antisera to avian and bovine pancreatic polypeptide, but to differ in some respects from that observed with antisera to molluscan cardioexcitatory peptide. The presence of neuropeptide Y immunoreactive fibres in tracts such as the corpus callosum, anterior commissure, lateral olfactory tract, fimbria, medial corticohypothalamic tract, medial forebrain bundle, stria terminalis, dorsal periventricular bundle and other periventricular areas, indicated that in addition to the localisation of neuropeptide Y-like peptide(s) in interneurons in the forebrain, neuropeptide Y may be found in long neuronal pathways throughout the brain.