Fine structure of neurons of the arcuate nucleus and median eminence of the hypothalamus of the golden hamster following immobilization

Summary The fine structure of arcuate neurons of the arcuate nucleus, the ependymal tanycytes and the contact zone of the median eminence was examined following immobilization, an acute stress which significantly activated the hypothalamo-pituitary-adrenal (HPA) axis. Arcuate neurons of immobilized adult male hamsters displayed morphological indications of heightened activity; the number of lysosomes and dense core vesicles (80–120 nm) was increased. A markedly greater number of dense core vesicles was present in axon terminals of the contact zone of the mid-central median eminence and the ventral proximal stalk.Tanycytes of the median eminence exhibited an augmented number of electron dense bodies in both perikarya and end processes. These results indicate that the arcuate neurons, the axons of the contact zone, and the ependymal tanycytes of the hamster medial basal hypothalamus (MBH) may be involved in the response to immobilization.This work was supported by Program Project Grant #NS-11642     

大鼠下丘脑内一氧化氮合酶阳性神经元的分布

用ＮＡＤＰＨ－ｄ组织化学方法观察了大白鼠下丘脑内一氧化氮合酶（ＮＤＳ）阳性神经元的分布及形态特征。结果显示：在视上核、室旁核的大细胞部、环状核、穹窿周核、下丘脑外侧区、下丘脑腹内侧核、下丘脑背内侧核、乳头体区大部分核团均可见一氧化氮合酶阳性神经元聚集成团。在视前内侧区、视前外侧区、下丘脑前区、下丘脑背侧区、下丘脑后区、室周核、室旁核小细胞部及穹窿内可见散在的一氧化氮合酶阳性神经元。室周核内可见呈阳性反应的接触脑脊液神经元的胞体及突起。一氧化氮合酶阳性神经元大多可见突起,有的突起上可见１～２级分支,并可见膨体。下丘脑大部分区域内可见阳性神经纤维。弓状核内可见许多弧形纤维连于第三脑室室管膜和正中隆起。     

Uptake of peroxidase from the third ventricle by ependymal cells of the median eminence

Summary Peroxidase injected into the subarachnoid space in mice is absorbed by ependymal cells of the median eminence. The ependymal cells of the median eminence of the rat and Japanese quail absorb peroxidase injected into the third ventricle. The processes of these ependymal cells terminate at the capillaries of the primary plexus or those surrounding the ventromedial nucleus of the hypothalamus. In all three species, peroxidase is absorbed by the ependymal cells of the paraventricular organ and by those in close proximity to it. Some ependymal cells send processes to the capillaries in the lateral nucleus of the hypothalamus. These phenomena are discussed in relation to adenohypophysial function.A part of this investigation was effected while the senior author held a Visiting Professorship at the University of Giessen [Department of Anatomy (Professor A. Oksche, Director)].     

A system of intraependymal cisternae along the margins of the median eminence in the rat: Structure,three-dimensional arrangement and ontogeny

Summary Structure, three-dimensional arrangement and ontogeny of large intracellular cisternae located in the median eminence region of the rat hypothalamus were studied using toluidin-blue stained semithin sections and electron microscopy. The cisternae occur along the projections of ependymal cells lining the ventral portion of the third ventricle (infundibular recess). Small cisternae can be seen close to the ventricle, whereas larger ones, divided into smaller compartments by thin septa, cluster near the surface of the hypothalamus. The cisternae are encompassed by a thin layer of cytoplasm to which axon terminals containing synaptic and dense core vesicles are closely related. Cisternae are arranged around the median eminence in a characteristic pattern. They occupy the midline in the retrochiasmatic area, flank both margins of the median eminence and extend caudally behind the origin of the pituitary stalk. The cisternae appear first between the 15th and 17th postnatal days. At about the 30th day their size and distribution resemble the situation observed in adult animals. The ependymal cisternae are suggested to be closely related to the luteinizing-hormone releasing-hormone (LH-RH)-containing fibers.     

Tanycyte absorption affected by the hypothalamic deafferentation in Japanese quail,Coturnix coturnix japonica

In Japanese quail, Coturnix coturnix japonica the tanycytes of the median eminence absorbed peroxidase injected into the third ventricle. The number of tanycytes showing peroxidase reaction was greater in the posterior median eminence than in the anterior median eminence. Following hypothalamic deafferentation, the tanycyte absorption was augmented both in the posterior and anterior median eminence. These findings suggest that axons of some neurons, which have inhibitory action on the tanycyte absorption, were transected by deafferentation resulting in augmentation of tanycyte absorption. A considerable number of ependymal cells lining the upper portion of the third ventricle and those of the pars nervosa also absorbed peroxidase. In birds with a deafferented hypothalamus, photostimulated ovarian growth was completely inhibited.     

Ultrastructural changes in the median eminence of the rat following deafferentation of the basal hypothalamus

Summary This report concerns a light and electron microscopic investigation of the median eminence and dorsal infundibular stem of the rat following surgical isolation (deafferentation). Using a modification of the Halász technique, the basal hypothalamus, including the arcuate nucleus and median eminence were surgically isolated from surrounding structures. Special attention was directed to the contact (external) zone of the median eminence and rostral infundibulum where tuberohypophyseal axons as well as ependymal cell processes abut upon the abluminal basement membrane of the portal perivascular space. The results of this study to date suggest that 9, 20, and 40 days following surgical isolation, there is a distinct increase in the population of tuberohypophyseal dense core vesicles. It is suggested that deafferentation abolishes inhibitory and excitatory input that serves to modify the cellular dynamics of tuberohypophyseal neurosecretory elements. Comments are also made on the presence of cistern-like structures in the lateral median eminence; the presence of vesicle-like inclusions in terminal ependymal processes is discussed in relationship to the role that ependyma may play in linking the third ventricle with the adenohypophysis.This research was supported by USPHS Grants NB 08171 and AM 10002. The authors are indebted to the excellent technical assistance provided by G. Krobisch Dudley. Further, the authors wish to express their thanks to Dr. Adolph Weindl for his valuable advice and criticism, and to Matilde Holzwarth for her helpful assistance.     

Mitotic Figures in the Median Eminence of the Hypothalamus

The median eminence of the hypothalamus is part of the avenue by which neurosecreted hormones from the hypothalamic nuclei reach the pars nervosa (neural lobe) of the pituitary and eventually the bloodstream. Lithium treatment and osmotic stress increases the transport of neurosecretory hormones to the pituitary in the adult rat. Specialized astrocytes termed pituicytes in the pars nervosa of the pituitary participate in the secretory process and also develop considerable mitotic activity. The present work reveals similar mitotic figures in cells within the median eminence following 3 days of lithium treatment. The location and appearance of these mitoses add to the evidence that pituicytes are present in the median eminence. Moreover, mitoses occur within the ependymal (tanycyte) layer of the median eminence. Thus, the present results suggest that the tanycyte layer may contain pituicytes, indicating that the hypothalamus possesses specialized cells for modulating neurosecretion in response to osmotic challenges.     

Immunocytochemical localization of neuropeptides in the hypothalamus of the Japanese long-fingered bat,Miniopterus schreibersii fuliginosus

Summary To elucidate the role of hypothalamic neuropeptides in regulation of reproductive phenomena of seasonally breeding feral mammals, we used Japanese long-fingered bats, Miniopterus schreibersii fuliginosus, for immunocytochemical study of distribution of the following neuropeptides in the hypothalamus: arginin vasopressin, oxytocin, luteinizing hormone-releasing hormone, somatostatin, corticotropin-releasing factor, and growth hormone-releasing factor. The size, shape and location of supraoptic, paraventricular, suprachiasmatic, and arcuate nuclei of the bat were determined. Arginin vasopressin-and oxytocin-immunoreactive magnocellular neurons were found in the supraoptic and paraventricular nuclei, where they exhibited separate distribution into two distinct groups. Parvocellular arginin vasopressin neurons occurred only in the suprachiasmatic nucleus. The hibernating bats exhibited slightly increased numbers of vasopressin and oxytocin neurons in the supraoptic and paraventricular nuclei. The pregnant bat displayed further increased numbers of vasopressin and oxytocin neurons in both nuclei. Somatostatin-immunoreactive neurons in the paraventricular nucleus were also immunopositive to anti-oxytocin serum, while those in the ventromedial and arcuate nuclei reacted solely to anti-somatostatin serum. They projected to the anterior median eminence and infundibular stalk. Luteinizing hormone-releasing hormone-immunoreactive perikarya were scattered throughout the basal hypothalamus, being particularly abundant in the arcuate nucleus. They were larger in size in hibernating bats than those in normal (non-pregnant) and pregnant females. They projected fibers mainly to the internal layer of the median eminence and infundibular stalk. A few luteinizing hormone-releasing hormone-reactive fibers were also observed in the organum vasculosum laminae terminalis, lateral habenular nuclei, pineal stalk, retroflexus fasciculus, and olfactory tubercle. Corticotropin releasing factor-immunoreactive perikarya were distributed in the paraventricular nucleus and medial preoptic area and projected into the external layer of the anterior median eminence, while growth hormone-releasing factor-immunoreactive perikarya occurred only in the arcuate nucleus and projected into the posterior part of the median eminence.     

Sites of origin of growth hormone-releasing factor-containing neurons projecting to the stalk-median eminence of the rat

The topographical location of neurons containing GRF which project to the median eminence were studied with immunofluorescence for GRF in combination with the retrograde transport of True blue. After the injection of True blue into the median eminence, retrogradely-labeled GRF neurons were identified in the arcuate nucleus and the lateral basal hypothalamus. GRF neurons in the perifornical area contained no positive dye. We concluded that the location of neurons containing hypophysiotrophic GRF are confined within the arcuate nucleus and the lateral basal hypothalamus.     

Immunocytochemical evidence for anti-LHRH and anti-ACTH activity in the "F" antiserum.

An antiserum which has previously been thought to be specific for LHRH-like immunoreactive material was shown to contain two populations of antibodies, one demonstrating anti-LHRH activity and the other anti-ACTH(1-24) activity. In rat and mouse, ACTH(1-24)-like immunoreactive substance is present in perikarya within the basal hypothalamus and in fibers in arcuate, periventricular and dorsomedial nuclei. LHRH-like immunoreactivity is present in fibers within the median eminence and arcuate nucleus, in a few fibers running along the ventral border of the hypothalamus, and in a small number of cell bodies within the medial basal hypothalamus.     

Localization of proopiomelanocortin (POMC) immunoreactive neurons in the forebrain of the pig

The distribution of proopiomelanocortin (POMC)-immunoreactive neurons was examined in the forebrains of nine sexually mature female pigs by indirect biotin-avidin horseradish peroxidase immunocytochemistry. Primary antiserum against ovine beta-endorphin (Bioflex #BF-EP-3-1) yielded positive staining of neuronal perikarya and processes. Adjacent control sections treated either with primary antiserum preabsorbed with beta-endorphin or substituted with normal rabbit serum lacked specific staining. POMC-immunoreactive cells were located in the anterior and intermediate lobe of the pituitary gland. POMC-immunoreactive perikarya were located in the arcuate nucleus and periarcuate area. The pituitary stalk/median eminence contained sparsely distributed POMC-immunoreactive fibers, which were confined to the zona interna. POMC-immunoreactive fibers were located in the arcuate nucleus and extended rostrally from the arcuate nucleus into the telencephalon coursing adjacent to the wall of the third ventricle as well as through the anterior hypothalamus, suprachiasmatic, supraoptic nuclei and preoptic areas to the nucleus accumbens, diagonal band of Broca, olfactory tubercle, bed nucleus of the stria terminalis and the ventro-lateral aspect of the septum. Caudal projections extended along the wall of the third ventricle to the level of the mammillary bodies and also coursed dorsally, passing through the periventricular, paraventricular, and dorsal medial nuclei of the hypothalamus to the midline thalamic nuclei and habenular nucleus. Lateral projections extended from the arcuate nucleus along the dorsal aspect of the optic tract and terminated in the amygdaloid complex. The distribution of POMC-immunoreactive perikarya and fibers is similar to that of the luteinizing hormone-releasing hormone (LHRH) fiber network. Therefore the opportunities exist, anatomically, for interactions between the POMC and the LHRH systems.     

Fine structure of ependymal cells in the median eminence of the frog and mouse revealed by freeze-etching

Summary Freeze-etched preparations of the ventricular surfaces of ependymal cells clearly reveal the presence of pinocytotic vesicles opening into the third ventricle and large vacuoles formed by broad cell projections. The density of the vesicular openings is approximately 20 per m². The ependymal cells in the median eminence of the frog are adjoined by tight junctions comprised of five to eight interconnected junctional strands, whereas near the median eminence in the mouse only one to two such strands form the tight junction of the ependymal cells. Gap junctions between the adjacent ependymal cells are detected near the median eminence in the mouse but not in the frog.This study was supported in part by a grant from the Japanese Ministry of Education (No. 067670)     

Distribution of arginine vasotocin in the brain of the lizard Anolis carolinensis

Summary The distribution of immunoreactive arginine vasotocin (AVT-ir) was determined in the brain of the lizard Anolis carolinensis. Cells and fibers containing AVT-ir were found in the medial septal region, lamina terminalis, lateral forebrain bundle, preoptic area, supraoptic nucleus, anterior hypothalamus, paraventricular nucleus, periventricular nucleus, arcuate nucleus, and ventromedial nucleus of the thalamus. Occasional AVT-ir cells were found in the interpeduncular nucleus. Fibers containing AVT-ir were found in the cortex, around the olfactory ventricle, in the diagonal band of Broca, amygdala area, dorsal ventricular ridge, striatum, nucleus accumbens, septum, ventromedial hypothalamus, lateral hypothalamus, medial forebrain bundle, median eminence, pars nervosa, nucleus of the solitary tract, locus coeruleus, cerebellar cortex (granular layer), dorsal part of the nucleus of the lateral lemniscus, substantia nigra, and myelencephalon. The intensity of AVT-ir staining was, in general, greater in males than in females. Comparison of AVT-ir distribution in A. carolinensis with those previously published for other reptilian species revealed species-specific differences in distribution of AVT.     

Structure of the neurohypophysis and the hypothalamo-hypophysial vascularization in the teleost Channa punctatus Bloch.

In C. punctatus the median eminence includes the subterminal region of the hypothalamus and the anterior neurohypophysis. It is formed of ependymal, fibrous and reticular layers as in the tetrapods. Primary capillary plexus extends from the subterminal region to the extremity of the anterior neurohypophysis. Only few portal vessels from the hypothalamus enter in the pars distalis. All the components of pituitary including the pars intermedia are irrigated by the secondary plexus formed from the portal vessels emerging out of the anterior neurohypophysis. The neurosecretory axons and the ependymal cells are in close morphological contact with the primary plexus. Several axons have perivascular endings at the median eminence. Some axons were found to be only silver or aldehyde fuchsin positive whereas some others take up both. The silver positive axons were abundant in the pars distalis and the AF positive ones were more concentrated in the pars intermedia with greater accumulation of neurosecretory material.     

Accessibility of low‐molecular‐mass molecules to the median eminence and arcuate hypothalamic nucleus of adult mouse

Blood‐derived molecules are able to access to the median eminence (ME) and arcuate hypothalamic nucleus (Arc) due to the lack of the blood–brain barrier. In the present study, we examined the accessibility of low‐molecular‐mass (LMM) molecules into parenchyma in the ME and Arc of adult mice by administration of Dextran 3000 (Dex3k), Dex10k, Evans blue (EB) and fluorescein isothiocyanate (FITC). In the external zone of the ME, the fluorescence of Dex3k, EB and FITC tracers generated an intensity gradient from fenestrated capillary, but that of Dex10k was detected only between the inner and outer basement membrane of pericapillary space. The fluorescence of FITC in the external zone of the ME was closely associated with axonal terminals and surrounded by cellular processes of tanycytes‐like cells and astrocytes. In the ependymal/internal zone of the ME and Arc, the fluorescence of all LMM tracers was seen at tanycytes‐like cells and neurons. The fluorescence of EB and FITC in these regions was not detected when brains were fixed during or before the administration of tracers. The inhomogeneity of accessibility for fluorescent tracers depended on routes for tracer administration. Thus, the present study indicates that the accessibility of LMM blood‐derived molecules to parenchyma depends on fenestration of the capillary in the external zone of the ME and active transport of ependymal cells in the ependymal/internal zone of the ME and Arc. Copyright © 2013 John Wiley & Sons, Ltd.     

Immunocytochemical localization of growth hormone releasing factor (GHRF)-containing structures in the rat brain using anti-rat GHRF serum

The distribution of growth hormone releasing factor (GHRF) immunoreactive structures in the rat hypothalmus was studied after colchicine treatment with PAP immunocytochemistry in vibratome sections using an antiserum directed to rat hypothalamic GHRF. The majority of the GHRF-immunoreactive cell bodies were found in the arcuate nucleus, the medial perifornical region, and the ventral premammillary nuclei of the hypothalamus. Scattered cells were seen in the lateral basal hypothalamus, the medial and lateral portions of the ventromedial nucleus, and the dorsomedial and paraventricular nuclei. Immunoreactive fibers were observed in all the regions mentioned above. GHRF terminals were located in the central region of the median eminence. In addition, GHRF-immunoreactive neuronal processes were seen in the ventral region of the dorsomedial nucleus, the medial preoptic and suprachiasmatic regions, dorsal portion of the suprachiasmatic nucleus, bed nucleus of the stria terminals and the hypothalamic portion of the stria terminals. The localization of GHRF-immunoreactive terminals in the median eminence reinforces the view that GHRF plays a physiological role in the regulation of pituitary function. In addition, the localization of GHRF-immunoreactive structures in areas not usually considered to project to the median eminence suggest that GHRF may act as a neuromodulator or neurotransmitter.     

Immunoreactivity to vasoactive intestinal polypeptide (VIP) and thyreotropin-releasing hormone (TRH) in hypothalamic neurons of the domesticated pigeon (Columba livia). Alterations following lactation and exposure to cold

Summary The distribution of VIP- and TRH-immunoreactivity in neurons and processes within the hypothalamus of the pigeon was investigated with light-microscopic immunocytochemical techniques. Most of the VIP-containing neurons are concentrated in the middle and caudal parts of the hypothalamus, with the greatest concentration of perikarya occurring in the medial and lateral part of the ventromedial hypothalamic nucleus and the infundibular nucleus. These cells give rise to axons that seem to extend into the median eminence. An extensive network of VIP-immunoreactive fibers and varicosities occupy the external layer of the median eminence. The majority of TRH-containing neurons is found in the anterior hypothalamus with the greatest concentration of cells in the magnocellular preoptic, medial preoptic, suprachiasmatic and paraventricular nuclei. TRH-immunoreactive fibers and varicosities form a dense arborization in the external layer of the median eminence. Lactation seems to induce substantial changes in VIP as well as in TRH-immunostaining in the median eminence and other hypothalamic regions as compared to control, sexually active animals. Furthermore, TRH-immunoreactivity decreased in the median eminence following 60-min exposure to cold. These results suggest that VIP- and TRH-containing pathways in the pigeon hypothalamus are involved in the mediation of neuroendocrine responses.     

Hypothalamic and extrahypothalamic distribution of somatostatin-immunoreactive elements in the rat brain

Summary Using a highly sensitive antibody to somatostatin, its hypothalamic and extrahypothalamic distribution in the rat was re-examined by light microscopic immunohistochemistry (PAP-method). The scattered somatostatin-producing perikarya occur in multiple layers within the subependymal neuropil surrounding the third ventricle. They supply with short-distance projections the following hypothalamic nuclei: 1) preoptic nuclei (especially their suprachiasmatic and medial components), 2) the peripheral zones of the suprachiasmatic nuclei, 3) the ventromedial and 4) arcuate nuclei, and 5) the ventral premammillary nuclei. Furthermore, the following long-distance projections have been observed: In a rostral direction (A1) rostral of the anterior commissure to the lamina terminalis, (A2) to the OVLT, (A3) to the olfactory tubercle, and (A4) rostrally and caudally by-passing the anterior commissure to the dorsal part of the stria terminalis.More caudally, at the retrochiasmatic level an ascending dorso-lateral projection joins the ventral amygdalo-hypothalamic pathway in a reciprocal manner (B1). In addition, a descending ventrolateral tract projects to the optic tract bending dorsal to it in different directions: (C1) medial to the median eminence, (C2) lateral to the corticomedial amygdala, and (C3) caudal for additional support of the arcuate and ventral premammillary nuclei.The principal tract of somatostatin-containing fibers descends in the subependymal neuropil to the median eminence (D).The results are discussed with reference to a possible participation of the somatostatin fiber system in the afferent branch of the circuit connecting the hypothalamus with the amygdala via the stria terminalis.Supported by the Deutsche Forschungsgemeinschaft (Grant Nr. Kr. 569/2) and Stiftung Volkswagenwerk.     

Morphological aspects of the hypothalamic-hypophyseal system

Summary The reaction of neural structures of the medial basal hypothalamus (MBH) to its complete deafferentation was studied in male rats by means of enzyme-histochemical and histoautoradiographic methods. Particular attention was paid to nerve cells of the arcuate nucleus and to the tanycytes. The metabolic activity of these cells increased upon the whole.According to the indices of metabolic activity in normal conditions and following deafferentation, the authors distinguish among the ependymal cells of the recessus infundibularis in rats, -tanycytes, which correspond to the ependymal lining at the level of the arcuate nucleus, and -tanycytes, which correspond to the ependyma of the median eminence. In normal conditions both were marked by a sufficiently high level of metabolism, which did not exclude the possibility of protein synthesis. Following deafferentation, -tanycytes seemed most reactive. The most active elements among the -tanycytes were the ependymal cells of the lateral part of the median eminence (¹-tanycytes).The metabolic peculiarities of the nerve cells of the arcuate nuclei and the tanycytes, revealed in normal conditions and after deafferentation, are discussed in connection with the modern concepts of the role of these cells in hypothalamic-hypophyseal transmission.     

A specific radioimmunoassay for Cortricotropin Releasing Factor (CRF) using synthetic ovine CRF

This newly developed specific radioimmunoassay for corticotropin releasing factor (CRF) had a sensitivity range of 25 pg/tube to 4 ng/tube. Intra and interassay coefficient of variation were 4.6% and 9.8%, respectively. Rat median eminence extracts showed a parallel dose response curve with synthetic ovine CRF and a significant cross reaction was not evident with other tested neuropeptides. The highest mean levels of CRF were found in the median eminence (6.61 ng/mg protein). Considerable amounts of CRF were found in the arcuate nucleus, paraventricular nucleus, dorsomedial nucleus, suprachiasmatic nucleus and ventromedial nucleus. The immunoreactive CRF of the rat medial basal hypothalamus coeluted with bioassayable CRF and with iodinated CRF on Sephadex G-75 chromatography. The results indicate that rat hypothalamus contains a CRF similar to ovine CRF.