Immunocytochemical localization of somatostatin-containing neurons in the rat hypothalamus

Summary The rat hypothalamus was studied at the light microscopic level with the use of single and double immunocytochemical staining methods. It was shown that the rat supraoptic and paraventricular hypothalamic nuclei, and their accessory neurosecretory nuclei, do not contain magnocellular somatostatin neurons. The distribution of the hypothalamic parvocellular somatostatin cells is described. The parvocellular component of the rat hypothalamic paraventricular nucleus is, at least partly, composed of somatostatin cells: they form a fairly well circumscribed periventricular cell mass. The rat suprachiasmatic nuclei contain separate somatostatin neurons and vasopressin neurons. Scattered somatostatin cells are present in the entire arcuate nucleus. In addition to the periventricular somatostatin cells located in the preopticanterior hypothalamic area and in the arcuate nucleus, the rat hypothalamus also contains numerous scattered somatostatin cells located distant from the third ventricle.This investigation was supported by a grant from the Belgian Nationaal Fonds voor Geneeskundig Wetenschappelijk Onderzoek     

Nuclei and fields of the rabbit hypothalamus]

Common features and distinctions in the structure of certain fields of the rabbit hypothalamus were established on the basis of cytoarchitectonical and cytological analysis. It has been shown that there are three types of nerve cells in the nuclei and fields of the hypothalamus which can be referred to somato-, cyto- and karyochromic elements of the nervous system in accordance with the Nissl classification. All the cellular structures of the hypothalamus can be divided into heteromorphic and isomorphic types. The medial and lateral hypothalamic fields of all the rostro-caudal length of the hypothalamus are referred to the first type. The hypothalamic nuclei occupying its basal part are referred to the second type. On the basis of the obtained data concerning the neuronal composition of the fields and nuclei of the hypothalamus, it can be divided into three zones: the medial zone, including 3 medial hypothalamic fields disposed along the 3d ventricle; the lateral zone comprizing two a lateral hypothalamic fields occupying its lateral parts along its all length and the basal zone including hypothalamic nuclei disposed mainly in the ventral part of the hypothalamus.     

Lateral septal projections onto tubero-infundibular neurons in the hypothalamus of the guinea pig

Efferent projections of the lateral septal nucleus (LS) to the preoptic area and the hypothalamus were identified in 20 female guinea pigs after iontophoretic injection of the anterograde axonal tracer Fluoro-Ruby. Tubero-infundibular (TI) neurons of the preoptic area and the hypothalamus were retrogradely labeled after intracardiac injection of Granular Blue or Fluoro-Gold. Magnocellular neurons of the supraoptic and paraventricular nuclei were also labeled. The double labeling procedure allowed an estimation of the extent of the direct relationship between LS efferents and TI neurons. Contacts between lateral septal fibers and TI cell bodies were mainly observed at the light-microscopical level in the preoptic area. A group of labeled fibers coursing along the third ventricle established sparse connections with hypothalamic periventricular TI neurons. A few appositions was observed in the infundibular (arcuate) nucleus, suggestive of a monosynaptic regulation of TI neurons by a septo-arcuate tract. Close association with labeled magnocellular neurons was also noted at the edge of the supraoptic and paraventricular nuclei. The sparse but direct connections between LS and TI neurons may be involved in the neuroendocrine functions of the LS.     

1例雄性朱鹮脑内脑啡肽的免疫组织化学分布

用免疫组织化学方法研究脑啡肽(ENK)在极危物种朱(Nipponia nippon)脑内的分布,结合计算机图像分析仪检测免疫阳性细胞和末梢的灰度值。ENK阳性细胞、纤维和终末分布如下:发声核团有原纹状体中间区腹部、丘脑背内侧核外侧部、中脑丘间核、中脑背内侧核、延髓舌下神经核。听觉中枢有丘脑卵圆核壳区、中脑背外侧核壳区、脑桥外侧丘系腹核、上橄榄核、耳蜗核等。内分泌核团有视前区前核、旧纹状体增加部、下丘脑外侧核、下丘脑腹内侧核等。结果表明,朱脑内ENK可能对发声、听觉和下丘脑内分泌的生理活动有一定的调制作用。     

Neuronal inputs from the hypothalamus and brain stem to the medial preoptic area of the ram: neurochemical correlates and comparison to the ewe

The retrograde tracer, FluoroGold, was used to trace the neuronal inputs from the septum, hypothalamus, and brain stem to the region of the GnRH neurons in the rostral preoptic area of the ram and to compare these imputs with those in the ewe. Sex differences were found in the number of retrogradely labeled cells in the dorsomedial and ventromedial nuclei. Retrogradely labeled cells were also observed in the lateral septum, preoptic area, organum vasculosum of the lamina terminalis, bed nucleus of the stria terminalis, stria terminalis, subfornical organ, periventricular nucleus, anterior hypothalamic area, lateral hypothalamus, arcuate nucleus, and posterior hypothalamus. These sex differences may partially explain sex differences in how GnRH secretion is regulated. Fluorescence immunohistochemistry was used to determine the neurochemical identity of some of these cells in the ram. Very few tyrosine hydroxylase-containing neurons in the A14 group (<1%), ACTH-containing neurons (<1%), and neuropeptide Y-containing neurons (1-5%) in the arcuate nucleus contained FluoroGold. The ventrolateral medulla and parabrachial nucleus contained the main populations of FluoroGold-containing neurons in the brain stem. Retrogradely labeled neurons were also observed in the nucleus of the solitary tract, dorsal raphe nucleus, and periaqueductal gray matter. Virtually all FluoroGold-containing cells in the ventrolateral medulla and about half of these cells in the nucleus of the solitary tract also stained for dopamine beta-hydroxylase. No other retrogradely labeled cells in the brain stem were noradrenergic. Although dopamine, beta-endorphin, and neuropeptide Y have been implicated in the regulation of GnRH secretion in males, it is unlikely that these neurotransmitters regulate GnRH secretion via direct inputs to GnRH neurons.     

雌激素受体在老年和青年猕猴下丘脑中表达的比较

为了观察老年猕猴下丘脑中雌激素受体的表达及变化情况,运用免疫组织化学SABC 法,对青年和老年猕
猴下丘脑雌激素受体的表达特点进行了研究。结果发现,雌激素受体免疫阳性细胞主要分布于下丘脑的室周灰
质、室旁核、腹外侧核、腹内侧核、弓状核等,雌激素受体阳性产物主要定位于细胞核和细胞质中。与青年猕
猴比较,老年猕猴下丘脑各个核团的雌激素受体在表达强度及阳性细胞数量上均显著或极显著低于青年猕猴
(P<0.05 或 P<0.01)。表明,老年猕猴下丘脑雌激素受体的表达显著低于青年猕猴,可能为老年猕猴雌激素缺
乏所致。     

Origin of the met-enkephalinergic innervation of the lateral septum in the rat

Summary The location of the cells giving rise to the methionine-enkephalin (Met-Enk)-ergic innervation of the lateral septal nucleus has been investigated in the rat by combining immunohistochemistry and retrograde axonal tracing. Small volumes (0.06 l) of apo-horseradish peroxidase (Apo-HRP) conjugated to wheat-germ agglutinin (WGA) and coupled with colloidal gold particles (WGA-ApoHRP-gold) were injected into the lateral septum. The retrogradely labeled cell bodies were visualized by silver intensification of the gold particles on Vibratome sections that were subsequently processed for immunohistochemistry for Met-Enk. Cells labeled with WGA-ApoHRP-gold were observed in the septal area, throughout the hypothalamus (mainly in the perifornical and lateral nuclei) and in the mesencephalon. The localization of Met-Enk-immunoreactive cells was as previously described. With the exception of a few septal cells close to the injection site, doubly labeled cells were found only in the perifornical nucleus of the hypothalamus. Almost all perifornical magnocellular cells were doubly labeled ipsilateral to the injection site, whereas on the opposite side, only about 25% of the Met-Enk-immunoreactive cells contained WGA-ApoHRP-gold. Other brain regions containing retrogradely labeled or Met-Enk-immunoreactive cells (particularly the raphe nuclei) did not show double-labeled neurons. This study demonstrates, using a new and sensitive technique for specific neurochemical tracing of tracts, that the origin of the Met-Enk-ergic innervation of the rat lateral septal nuclei lies in the magnocellular perifornical nuclei of the hypothalamus. The precise involvement of this pathway in limbic functions remains to be determined.     

The formation of catecholaminergic structures in the mediobasal hypothalamus of the rat

A study was made of the formation of catecholaminergic system in the medial basal hypothalamus of Wistar rat fetuses using histofluorescent method modified by de la Torre. In the periventricular region of 16-day fetuses the cells with green fluorescence characteristic for catecholamines were found. In 18-day fetuses catecholaminergic cells were found in paraventricular, arcuate and dorsomedial hypothalamic nuclei. In all the nuclei studied the number of catecholaminergic fibers and terminals of various size is greatly increased. The data obtained suggest that medial basal hypothalamus of 18-day fetuses has a complex catecholaminergic system.     

Immunocytochemical distribution of FMRFamide-like substance in the brain of the cloudy dogfish,Scyliorhinus torazame

Summary The distribution of the molluscan cardioexcitatory tetrapeptide FMRFamide (Phe-Met-Arg-Phe-NH₂) in the brain of the cloudy dogfish, Scyliorhinus torazame, was examined by immunocytochemistry. FMRFamide-like immunoreactivity was demonstrated to occur extensively in various regions of the dogfish brain, except for the corpus cerebelli. Immunoreactive neuronal perikarya were located in the ganglion of the nervus terminalis, the preoptic area, and the hypothalamic periventricular gray matter consisting of the nucleus medius hypothalamicus, the nucleus lateralis tuberis, and the nucleus lobi lateralis. some of the immunoreactive cells in the hypothalamus were identified as cerebrospinal fluid-contacting neurons. The bulk of the immunostained fibers in the nervus terminalis penetrated into the midventral portion of the telencephalon and ran dorsocaudally toward the basal telencephalon and hypothalamus, showing radial projections or ramifications. The labeled fibers were abundant in the midbasal part of the telencephalon and in the hypothalamus, where some fibers were found in loose networks around the cell bodies of the nucleus septi and hypothalamic periventricular nuclei. The fibers demonstrated in the hypothalamus terminated around the vascular wall of the primary capillary plexus of the median eminence or penetrated deeply into the pars intermedia of the hypophysis. These results suggest that, in the dogfish, an FMRFamide-like substance participates in the regulation of adenohypophysial function. This molecule may have a role as a neurotransmitter and/or neuromodulator in the central nervous system.     

青紫蓝兔体内Ghrelin的免疫组化定位

采用免疫组织化学方法研究了Ghrelin在青紫蓝兔(Oryctolagus cuniculus)体内的分布定位。结果显示,Ghrelin免疫阳性细胞分布于下丘脑、大脑皮质、延髓、脊髓、胃、小肠和大肠。下丘脑内的阳性神经元胞体主要分布于弓状核、室旁核、腹内侧核、背内侧核和下丘脑外侧区。胃肠道内的Ghrelin免疫阳性细胞存在两种类型,即"闭合型"细胞和"开放型"细胞。实验结果表明,兔体内Ghrelin的分布与人和其他动物的分布基本相似,但也存在一些差异。     

Estrogen receptor and progesterone receptor-immunoreactive cells are not co-localized with gonadotropin-releasing hormone in the brain of the female mink (Mustela vison)

The distribution of gonadal steroid (estrogen, progesterone) receptors in the brain of the adult female mink was mapped by immunocytochemistry. Using a monoclonal rat antibody raised against human estrogen receptor (ER), the most dense collections of ER-immunoreactive (IR) cells were found in the preoptic/anterior hypothalamic area, the mediobasal hypothalamus (arcuate and ventromedial nuclei), and the limbic nuclei (amygdala, bed nucleus of the stria terminalis, lateral septum). Immunoreactivity was mainly observed in the cell nucleus and a marked heterogeneity of staining appeared from one region to another. A monoclonal mouse antibody raised against rabbit uterine progesterone receptor (PR) was used to identify the PR-IR cells in the preoptic/anterior hypothalamic area and the mediobasal hypothalamus (arcuate and ventromedial nuclei). This study also focused on the relationship between cells containing sex-steroid receptors and gonadotropin-releasing hormone (GnRH) neurons on the same sections of the mink brain using a sequential double-staining immunocytochemistry procedure. Although preoptic and hypothalamic GnRH neurons were frequently in close proximity to perikarya containing ER or PR, they did not themselves possess receptor immunoreactivity. The present study provides neuroanatomical evidence that GnRH cells are not the major direct targets for gonadal steroids and confirms for the first time in mustelids the results previously obtained in other mammalian species.     

Different distributions of nitric oxide synthase-containing neurons in the mouse and rat hypothalamus.

The distribution of nitric oxide synthase-containing neurons was studied in the rat and mouse hypothalamus by immunohistochemistry and NADPH-diaphorase histochemistry. Immunostaining and NADPH-diaphorase staining of hypothalamic neurons were comparable in all hypothalamic nuclei of either species except in the arcuate nucleus that stained positive for nitric oxide synthase immunoreactivity but negative for NADPH-diaphorase reactivity. The presence of nitric oxide synthase-immunopositive neurons in the arcuate nucleus was confirmed by nitric oxide synthase immunofluorescence viewed under the confocal microscope at 1 microm thickness. Cross-species comparison showed that, in general, the number and intensity of nitric oxide synthase-containing neurons were much higher in the rat than in the mouse hypothalamus. Differences in the distribution of nitric oxide synthase-containing neurons between these two rodents were found in most hypothalamic nuclei. In particular, two dense clusters of nitric oxide synthase-containing neurons were found in the paraventricular and supraoptic nuclei of the rat hypothalamus in contrast to their scarcity in the same nuclei of the mouse hypothalamus.     

Morphology of neuroglia in the hypothalamus of the opossum (Didelphis virginiana), armadillo (Dasypus novemcinctus mexicanus) and cat (Felis domestica)

The hypothalamus of the opossum (Didelphis virginiana), the armadillo (Dasypus novemcinctus mexicanus), and the cat (Felis domestica) was studied using Del Rio Hortega's silver carbonate technique, as modified by Scharenberg ('60). This technique demonstrates astrocytes, oligodendroglia, and neuronal perikarya, but does not impregnate microglia. The morphology of macroglia was observed in ten comparable nuclei in each of the three species. The subpial and subependymal areas were also examined. Astrocytes display more cell body angularity and have more processes in most hypothalamic regions of the cat when compared to similar regions of the opossum and armadillo. In the anterior hypothalamic nucleus, the ventromedial and the dorsomedial hypothalamic nuclei, and the medial mammillary nucleus of all three species, astrocytes send processes to neurons, but neuronal and astrocytic perikarya are usually not directly contiguous. However, oligodendrocytes in a perisomatic position on neurons are a consistent feature in these nuclei. A closer relationship appears to exist between astrocytes and neurons in the neurosecretory nuclei. In the supraoptic nucleus and paraventricular nucleus of all three species a basket-like structure, designated a ?pericellular envelope”? was observed surrounding neuronal perikarya. This structure is composed of astrocytic and oligodendroglial cell bodies and processes, and is most highly developed in the cat. A dense astrocytic plexus was observed in the suprachiasmatic nucleus of the cat, and in the comparable nuclei of the armadillo and opossum. The most prominent macroglial cell type of the lateral hypothalamic and lateral mammillary nuclei of all three species is the interfascicular oligodendrocyte. The posterior hypothalamic nucleus of each species has many perisomatic oligodendrocytes, and in the armadillo and cat astrocytes are closely related to the larger neurons. A subpial plexus, consisting of a palisade of small glial cells with many processes, is present in the hypothalamus of the three species. Ependymal cells have long projecting processes throughout the length of the third ventricle in the armadillo hypothalamus, but such processes are only apparent in the region of the infundibular nucleus and median eminence in the opossum and cat.     

Steroid receptors in the brain: Topography and some functional implications

The anatomical localization of brain cells which concentrate steroid hormones or their metabolites was carried out by radioautographic procedures. Ovariectomized or adrenalectomized animals were injected with the appropriate tritiated hormones, and brain tissue was processed through procedures which minimize the removal or displacement of steroids. Target cells were characterized by the concentration and retention of radioactive hormone in their nuclei. For each mammalian steroid hormone, nuclear binding sites exist in populations of cells with a specific regional localization in the brain and in the pituitary. The distribution of estrogen target cells was remarkably similar in the brains of rodents and primates although some minor species differences existed. Heavily labeled cells were present in the preoptic region, the septum, the amygdala and the mediobasal hypothalamus. The localization of progestagen-concentrating cells in the rodent and galago brain was limited to two hypothalamic areas: the preoptic region and the mediobasal hypothalamus. Corticosterone target cells were situated in extrahypothalamic regions of the rat central nervous system such as the hippocampus, the septum, the amygdala and certain regions of the brain cortex. However, the synthetic glucocorticosteroid, dexamethasone, was mainly found in the pituitary cells and in some neurons and glial cells of the mediobasal hypothalamus. The distribution pattern of steroid-sensitive cells within the brain and the pituitary gland corresponds to sites which are involved in the neuroendocrine processes regulating reproduction, including gonadotropin secretion and sexual behavior.     

Organization of hypothalamic area of diencephalon in the sturgeons

Cytoarchitectonics of hypothalamic area of diencephalon of the sturgeons was studied in serial sections by techniques of Nissl staining and Bielschowski impregnation in Viktorov’s modification. The hypothalamus was shown to be the most expanded area of diencephalon and forms its the most ventral part. The hypothalamic area of four studied sturgeons, the hausen, Huso huso L., Kura sturgeon, Acipenser guldenstaedtii persicus n. Kurensis Belyaeff, Caspian sturgeon, Ac. stellatus Pall. and the barbel sturgeon Ac. nudioventris Lov. was found to have similar structure. Eleven nerve structures are identified and described in the hypothalamic area: dorsal, ventral, and caudal periventricular zones, rostral and dorsal hypothalamic nuclei, ventral and ventrolateral hypothalamic nuclei, diffuse and central nuclei of the inferior lobes, nucleus of the vascular sac, and mammillary nucleus. Peculiarities and common features of organization of four major parts of hypothalamus of the sturgeons are considered in comparison with those of other ray-finned fish. The performed analysis indicates a high level of development of hypothalamus of the sturgeons.     

Innervation of the superficial pineal of the rat using retrograde tracing methods

Fast blue (FB), rhodamine microspheres (RH), horseradish peroxidase (HRP), and wheat germ agglutinin-horseradish peroxidase conjugate (WGA-HRP) were used as retrograde tracers to study the innervation of the rat superficial pineal gland (SP). One of the tracers was injected into the gland of each animal. All four retrograde tracers injected into the gland always labeled neurons in the superior cervical ganglia (SCG). No retrograde labeling was ever seen in the suprachiasmatic nuclei, paraventricular hypothalamic nuclei, lateral hypothalamus, habenular nuclei, amygdalar nuclei, or superior salivatory nuclei. Retrograde labeling was seen in the anterior hypothalamic nuclei, anterior thalamic nuclei, lateral geniculate bodies, and midbrain tectal structures when a tracer spread from the injection site to the overlying cortex, tectum, or commissures. Control studies included injection of tracer into the subarachnoid space around the SP or into structures adjacent to the SP. Only the injection of FB or WGA-HRP into the subarachnoid space labeled neurons in the SCG. This labeling was probably due to the spread of tracer to the choroid plexus. These results agree with recent work confirming the existence of a direct projection of the SCG into the interstitium around pinealocytes. The evidence does not substantiate an innervation originating in the habenular nuclei; the superior salivatory nuclei; or any other diencephalic, midbrain, pontine, or medullary structure.     

Peptide immunoreactive neurons in the amygdala and the bed nucleus of the stria terminalis project to the midbrain central gray in the rat.

The central nucleus of the amygdala, bed nucleus of the stria terminalis, and central gray are important components of the neural circuitry responsible for autonomic and behavioral responses to threatening or stressful stimuli. Neurons of the amygdala and bed nucleus of the stria terminalis that project to the midbrain central gray were tested for the presence of peptide immunoreactivity. To accomplish this aim, a combined immunohistochemical and retrograde tracing technique was used. Maximal retrograde labeling was observed in the amygdala and bed nucleus of the stria terminalis after injections of retrograde tracer into the caudal ventrolateral midbrain central gray. The majority of the retrogradely labeled neurons in the amygdala were located in the medial central nucleus, although many neurons were also observed in the lateral subdivision of the central nucleus. Most of the retrogradely labeled neurons in the BST were located in the ventral and posterior lateral subdivisions, although cells were also observed in most other subdivisions. Retrogradely labeled neurotensin, corticotropin releasing factor (CRF), and somatostatin neurons were mainly observed in the lateral central nucleus and the dorsal lateral BST. Retrogradely labeled substance P-immunoreactive cells were found in the medial central nucleus and the posterior and ventral lateral BST. Enkephalin-immunoreactive retrogradely labeled cells were not observed in the amygdala or bed nucleus of the stria terminalis. A few cells in the hypothalamus (paraventricular and lateral hypothalamic nuclei) that project to the central gray also contained CRF and neurotensin immunoreactivity. The results suggest the amygdala and the bed nucleus of the stria terminalis are a major forebrain source of CRF, neurotensin, somatostatin, and substance P terminals in the midbrain central gray.     

Dynamics of spontaneous activity of hypothalamic neurons in the rat during emotional stress

Statistic parametres of discharges of separate cells in various hypothalamic nuclei were studied on Wistar rats in acute experiment under aethernembutal anesthesia. By logarithmic transformation of activity parameters a difference was established and some similar features of background firing of neurones in the frontal and posterior zones of the hypothalamus. Changes were shown in quantitative characteristics of impulse activity of hypothalamic neurones during development of emotional stress in conditions of long-term sexual conflict situations.     

Uptake of oestradiol by the rabbit hypothalamus. Specificity of binding by nuclei in vitro

The binding of [6,7-(3)H]oestradiol-17beta to subcellular fractions of the hypothalamus and the cerebellum of the rabbit was studied in vitro. Uptake of steroid was higher in hypothalamic nuclei than in cerebellar nuclei. Lower binding was observed in other fractions of both tissues. After dialysis of the fractions, hypothalamic nuclei retained a high percentage of oestradiol whereas cerebellar nuclei lost most of the bound steroid. Supernatant fractions of both tissues retained a significant proportion of label after dialysis and after gel filtration on Sephadex G-200. No specific binding was observed in these fractions when subjected to sucrose-density-gradient centrifugation. Purification of nuclei followed by incubation with labelled oestradiol in the absence of the supernatant fraction resulted in loss of binding of steroid by hypothalamic nuclei. Incubation of the purified hypothalamic nuclei with supernatant fraction maintained the binding specificity of hormone retention.     

Pituitary Adenylate Cyclase Activating Polypeptide (PACAP)-like immunoreactivity in human hypothalamus: co-localization with arginine vasopressin

Pituitary adenylate cyclase activating polypeptide (PACAP) is a novel hypothalamic peptide consisting of 38 amino acids (PACAP1–38) with a potent stimulatory action on adenylate-cyclase in rat pituitary. The presence of PACAP-like immunoreactivity in human brain was studied by radioimmunoassay. Co-localization of PACAP with arginine vasopressin and oxytocin was investigated by immunocytochemistry in the human hypothalamus. Immunoreactive PACAP was detected in all regions of human brain (cortex, thalamus, hypothalamus, pons and hemisphere of cerebellum) with the highest levels found in the hypothalamus (8.5±1.9 pmol/g wet weight, n=w, mean±S.E.M.). High performance liquid chromatography of the human hypothalamic ex approximately 50% of the immunoreactive PACAP was eluted in the position of PACAP1–38. Immunocytochemical studies showed the presence of PACAP immunoreactive neurons in the paraventricular and supraoptic nuclei of human hypothalamus. PACAP co-localized with arginine vasopressin in magnocellular cells of these nuclei. These findings suggest that PACAP1-38 plays important physiological roles in the human hypothalamus.