Effects on gonadal function by lesioning tanycytes in the median eminence of the rat and Japanese quail

Summary The ependymal linings of the median eminence were destroyed by electric cautery or intraventricular injection of picric acid in the rat and Japanese quail. In these animals the ventricular lumen near the median eminence disappeared due to adhesion of lesioned walls on both sides of the third ventricle. Electric lesions of the ependymal layer containing tanycytes did not induce appreciable disturbance in the estrous cycles. Rats in which tanycytes were lesioned by picric acid displayed 4-day estrous cycles after prolonged diestrus (10–22 days). After destruction of tanycytes in the quail, a photostimulated gonadal growth was observed. It is concluded that the tanycyte transport of the ventricular fluid to capillaries of the portal vessels appears unnecessary for maintenance of adenohypophysial gonadotrophic activities.Supported by a grant from the Ministry of Education of Japan and by a grant from the Ford Foundation. The authors wish to express their gratitude to Dr. B.K. Follett for providing chicken LH (IRC2) and to the late Mr. T. Asai for assay of serum LH     

The tanycyte of the rat median eminence

Summary The present ultrastructural study proves the existence of nerve terminals closely apposed to the plasmalemmata of tanycytes in the rat median eminence. Several of these axo-tanycytic endings display remarkable accumulations of agranular endoplasmic reticulum in the form of pleomorphic vesicles which are closely apposed on either side of the plasma membrane of each cell compartment. Some of these vesicular profiles give the impression of structural continuity across both membrane systems. This phenomenon is discussed in the context of being a potential substratum for communication between both cell compartments.Supported by NIH Grant NS13717 and NSF Grant BNS 78-11820     

Transmission and scanning electron-microscopic observations on tanycytes in the mediobasal hypothalamus and the median eminence of adrenalectomized rats

Summary Tanycytes in the median eminence (ME) of the rat exhibit morphological features suggesting their possible participation in transport phenomena. After adrenalectomy, which modifies the hypothalamo-hypophyseal axis, they undergo morphological changes characterized by an accumulation of lipid droplets, an increased number of bleb-like protrusions at their apex, as well as an increased pinocytosis of intraventricularly injected horseradish peroxidase (HRP). In addition, after adrenalectomy an increased number of vacuoles appears at the level of the tubero-infundibular sulci. Their intracellular location in the tanycytes is demonstrated by an intraventricular injection of HRP. The significance of these vacuoles is discussed in relation to the hydroelectrolytic modifications associated with the state of adrenalectomy.Supported by a Fonds Lekime-Ropsy AwardSupported by grants: 20472 from the Fonds de la Recherche Scientifique Médicale; and 40025 from the Fonds National de la Recherche Scientifique     

Cellular organization of the lateral and postinfundibular regions of the median eminence in the rat

Summary The structural organization of the rostral, lateral and postinfundibular regions of the median eminence (ME) of 5-day cyclic diestrous rats was studied with light and electron microscopic methods. The ependymal cells lining (i) the floor of the infundibular recess (IR) at rostral levels, (ii) the lateral extensions of the IR, and (iii) the floor of the premammillary recess appear to represent the same type of tanycyte ependyma (1 tanycytes). In the entire width of the rostral and postinfundibular palisade regions, as well as in the lateral palisade region of the preinfundibular ME, the processes of the 1 tanycytes form a continuous cuff. This cuff separates the nerve endings from the blood vessels and the pars tuberalis. At this level, synaptoid contacts between neurosecretory axons and the ependymal cuff can be observed. The ultrastructural characteristics of the 1 tanycytes are described and their ependymal endings tentatively classified into three types. In the lateral regions of the ME, the Golgi study revealed the presence of two fiber systems: (i) one possessing a latero-medial trajectory and distributed in the subependymal region; (ii) the other formed by a loose longitudinal tract originating from neurons of the arcuate nucleus. Some functional implications of the cellular organization of the rat ME are discussed.Supported by Grants from PLAMIRH (92.171.2.77) and from the Dirección de Investigaciones, Universidad Austral (S-77-28)The authors wish to thank Miss Rosario Andrade, Mrs. Elizabeth Santibáñez and Mr. Armando Bilbao for their assistance     

The functional and structural border of the neurohemal region of the median eminence

Summary In stressed rats the tanycytes of the ventrolateral wall of the third ventricle exhibit by light microscopic immunohistochemistry a positive staining for neurohormones which is distinctly limited to the distal perivascular end of the tanycyte process. Since by electron microscopic immuncytochemistry the tanycyte cytoplasm does not show any reaction product, the light microscopic reaction most likely results from a labeling of the intercellular space in the direct vicinity of the subendothelial cleft. Whether this subendothelial space is permeable to neurohormones was tested by injection of HRP¹. In the region of the arcuate nucleus 30 min after intravenous application, the marker is affixed to the membranes of the perivascular tanycyte processes in the subendothelial cleft of capillaries possessing non-fenestrated endothelia. Occasionally, HRP penetrates for a short distance between the tanycytes. Then the labeling of the intercellular cleft ends abruptly. Here, several parallel ridges of tight junctions between the perivascular distal tanycyte processes are found by the freezeetching technique. Since HRP cannot reach the subendothelial clefts of this region by passing through capillary walls due to the presence of a blood-brain barrier, it is suggested that the marker penetrates from the median eminence this far via the subendothelial extracellular space. It is prevented from spreading further by the tight junctions of the perivascular tanycyte endings. The same way may be taken by the neurohormones. Hence, a border area exists adjacent to the dorsolateral aspect of the neurohemal region of the median eminence where the tanycytes isolate the neuropil from the cerebrospinal fluid not only by their apical tight junctions, but also by basal tight junctions from the subendothelial cleft. This communicates with the perivascular space of the portal vessels.Supported by the Deutsche Forschungsgemeinschaft (Grant Nr. Kr. 569/2) and Stiftung VolkswagenwerkDedicated to Professor Dr. R. Ortmann on the occasion of his 65th birthday.The skilful technical assistance of Miss K. Bielenberg, Mrs. A. Hinz and Mrs. H. Prien is thankfully acknowledged     

Prenatal and early postnatal development of the glial cells in the median eminence of the rat

Summary The development of the glial cells of the rat median eminence (ME), including the supraependymal cells, was investigated from embryonic day (ED) 14 through postnatal day (PD) 7, and pituicyte development from ED 12 through ED 17. The anlage of the ME and neurohypophysis shows a neuroepithelial-like structure at ED 12. From ED 13 to 15, the cells of both regions start to differentiate. At the ultrastructural level, only one cell type appears. At the beginning of ED 16, glioblasts of the oligodendrocyte and astrocyte series migrate laterally (from the region of the arcuate nucleus) into the ME. Also at this time the first distinctive structural features appear in the neurohypophysial anlage, the cells of which later develop into pituicytes. Starting at ED 18, tanycytes and astrocytic tanycytes arise in the ME from local glial cells, and somewhat later oligodendroblasts and astroblasts are formed from immigrant glioblasts. Due to their common features, the pituicytes, tanycytes and astrocytic tanycytes apparently represent different forms of the same parent cell type. Microglial and supraependymal cells are first seen at ED 12. Initially, they resemble the prenatal phagocytic connective tissue cells and mature in the fetus into typical electron-dense microglia and macrophage-like supraependymal cells. Both cell types are apparently of mesodermal origin. The microglial elements of the ME probably migrate from the mesenchyma through the basement into the nervous tissue. The intraventricular macrophages of the infundibular region may originate from microglia, epiplexal cells and subarachnoid macrophages.Dedicated to Prof. I. Törö, Budapest, on the occasion of his 80th birthday     

The adsorptive and transport capacity of tanycytes during the perinatal period of the rat

Summary Transport of ferritin and horseradish peroxidase from the 3rd ventricle to the median eminence was examined in rats during the perinatal life, the time when functional interrelations between hypothalamus and hypophysis are established. Protein tracers injected into the lateral ventricle are adsorbed on the apical surface of the tanycyte, mainly on its protrusions or in indentations. On the 18th day of prenatal life a few small bleblike protrusions are observed. After birth microvilli appear. In time their concentration increases to result in an increase of adsorbed substances. They are taken up by smooth and coated pinocytotic vesicles and transported to the basal portion of the cell or to the intercellular space bypassing junctional complexes. In addition to pinocytotic vesicles protein tracers fill channels of smooth ER or Golgi complex and multivesicular bodies illustrating a process probably involved in metabolic or secretory processes.     

Antibodies obtained by xenotransplantation of organ-cultured median eminence specifically recognize hypothalamic tanycytes

Tanycytes are specialized ependymal cells lining the infundibular recess of the third ventricle of the cerebrum. Early and recent investigations involve tanycytes in the mechanism of gonadotropin-releasing hormone (GnRH) release to the portal blood. The present investigation was performed to obtain a specific immunological marker of tanycytes and to identify the compound(s) responsible for this labeling. After 30 days of organ culture, explants of bovine median eminence formed spherical structures mostly constituted by tanycytes. These tanycyte spheres were xenotransplanted to rats, and the antibodies raised by the host animals against the transplanted living tanycytes were used for immunochemical studies of the bovine and rat median eminence. This antiserum immunoreacted with two compounds of 60 kDa and 85 kDa present in extracts of bovine and rat median eminence. The individual immunoblotting analysis of rat medial basal hypothalami showed a decrease in the amount of the 85-kDa compound in castrated rats as compared to control rats processed at oestrus and dioestrus. The antiserum, labeled as anti-P85, when used for immunostaining of sections throughout the rat central nervous system, immunoreacted specifically with the hypothalamic tanycytes. Within tanycytes, P-85 immunoreactivity was exclusively present in the basal processes. It is suggested that the 85-kDa and 60-kDa compounds correspond to two novel proteins selectively expressed by tanycytes. The possibility that they are secretory proteins involved in GnRH release is discussed. Anti-P85 appears to be the first specific marker of hypothalamic tanycytes.     

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     

Polarized endocytosis and transcytosis in the hypothalamic tanycytes of the rat

Four types of tanycytes can be distinguished in the rat hypothalamus: ₁ and ₂ tanycytes establish an anatomical link between the ventricular cerebrospinal fluid (CSF) and the arcuate nucleus, whereas ₁ and ₂ tanycytes establish a link between CSF and portal blood. Endocytosis and transcytosis in these cells have been investigated by (1) immunocytochemistry with antibodies against molecular markers of the endocytotic and transcytotic pathways; (2) the administration of wheat germ agglutinin (WGA) into the ventricular or subarachnoidal CSF and following its internalisation by and its routing through tanycytes. The four populations of tanycytes show marked differences concerning the expression and subcellular location of proteins involved in endocytosis and transcytosis, such as clathrin, caveolin-1, Rab4 and ARF6. Thus, _1,2 tanycytes express caveolin-1 at the ventricular cell pole and at their terminals contacting the portal capillaries, whereas _1,2 tanycytes do not, suggesting that caveolae-dependant endocytosis does not occur in the latter and that, in _1,2 tanycytes, it may occur at both cell poles. In _1,2 tanycytes, clathrin is only expressed at the ventricular cell pole indicating that clathrin-dependant endocytosis operates for compounds present in the ventricular CSF and not for those exposed to the terminals. This agrees with the property of _1,2 tanycytes of internalising WGA through the ventricular cell pole but not through the terminals. The subcellular distribution in _1,2 tanycytes of WGA and of the proteins clathrin and Rab4 indicates that part of the internalised WGA follows the degradative pathway and part is sorted to a transcytotic pathway and that the transcytotic and the secretory pathways might intersect. Financial support was provided by grants 01/1050, from FIS, Spain (to J.L.B.) and 1030265, from FONDECYT, Chile (to E.M.R.)     

Failure to demonstrate serotonergic neurons in the myenteric plexus of the rat

Summary Several recent studies suggested that serotonergic neuron-like elements are present in the guinea pig ileum. The present paper reports an extensive study of the digestive tract of the rat with the use of a histofluorescence technique. Administration of the serotonin precursor, tryptophan, associated with a monoamine oxidase inhibitor, did not allow histochemical demonstration of rapidly fading, yellow fluorescent, 6-hydroxydopamine-resistant neurons; conversely such neurons were readily detected in the brain. It is concluded that serotonergic neuron-like elements cannot be detected histochemically in the rat myenteric plexus area after chemical sympathectomy.     

Ultrastructure of the median eminence of the rat

Summary The ependymal cells bordering the median eminence to the third ventricle are characterised by many microvillus-like projections and bulbous cell processes of the luminal plasma membrane. The latter contain many vesicles 500–1,000 Å in diameter. Cilia with 9+2 fibrillar pattern are seen occasionally. Adhesive devices in the from of zonula adhaerens and zonula occludens are found in the apical part of the intercellular junction. Unmyelinated nerve fibres with a mean diameter of 1 and containing many electron dense granules of 830–1,330 Å are often seen between the ependymal cells.Two types of glial cells are found in the median eminence. One is characterised by a nucleus with dense blods of chromatin and dense cytoplasm, and it is associated chiefly with the nerve fibres in the region of the hypothalamo-hypophysial tract. The other type of glial cell is characterised by fine, uniformly distributed chromatin in the nucleus and a relatively pale cytoplasm and branched processes which terminate perivascularly in the base of the median eminence.Myelinated nerve fibres are seen only in the region of the hypothalamo-hypophysial tract. Only a part of them contain electron dense granules 1,330–2,330 Å in diameter.Three types of unmyelinated nerve fibres can be distinguished in the median eminence according to the size of the electron dense granules they contain: 1. Nerve fibres containing granules 1,330–2,330 Å in diameter. They are seen primarily in the hypothalamo-hypophysial tract, but also in the zona externa; 2. those containing granules with a mean diameter of 1,330 Å; and 3. those containing granules with a mean diameter of 1,000 Å. The last two types are both encountered in the hypothalamo-hypophysial tract, the zona externa and the perivascular region of the base of the median eminence. Under high magnification, the membrane of the granules show evidence of a trilaminar structure and the content of the granules with a low electron density appeares to consist of small microvesicles or globular components. Besides granules, these nerve fibres contain vesicles mostly 420 Å in diameter whose relative number increases towards the perivascular nerve endings. 53 per cent of the inclusions in the hypothalamo-hypophysial tract are granules and 47 per cent vesicles, while the corresponding percentages for the zona externa are 40 and 60 and for the perivascular nerve endings 20 and 80.The mean width of the pericapillary space is 1 , but it varies greatly. It containes many collagen fibrils and fibroblasts. The capillary endothelium is frequently fenestrated and contains many vesicles of various sizes.Two types of granules-containing cells are found in the pars tuberalis depending on the size of the electron dense granules: 1. cells containing granules with a mean diameter of 1,330 Å: and 2. cells containing granules with a mean diameter of 2,000 Å. In addition, there are occasional follicular cavities filled with amorphous material, microvilli and cilia of 9+2 fibrillar pattern.Aided by a grant from the Sigrid Jusélius Stifteise.     

Quantitative radioautographic light and electron microscopic analysis of the localization of monoamines in the median eminence of the rat

Summary Serotonin containing structures in the median eminence of the rat have been studied by quantitative light and electron microscopic radioautography following intraventricular infusion of tritiated 5-hydroxytryptophan. One hour after injection of the tracer the highest density of silver grains was recorded in the ependymal and external zones, especially in the lateral palisade zone. The proportion of labelled neurosecretory terminals was also larger in the lateral palisade zone (29%) as compared with the medial palisade zone (13%), although the mean number of developed silver grains per one terminal was higher in the latter. On the average, 16% of neurosecretory terminals sequestered radiolabelled 5-hydroxytryptophan in the external zone of the rat median eminence. It is suggested that serotonin, like catecholamines, is discharged from neurosecretory terminals localized in the external zone and via the portal circulation affects the function of the anterior pituitary. The sites of origin of serotoninergic structures of the median eminence as well as the possible role of monoamine (catecholamine and indolamine) neurohormones in a dual peptidergic and monoaminergic control of anterior pituitary functions are discussed.This work was presented at the joint session of the Society of Anatomists, Histologists and Embryologists, the Society of Physiologists, and the Society of Neuroendocrinologists dedicated to the memory of Professor Wolfgang Bargmann, held in Leningrad, January 17, 1979     

Ultrastructure of the rat median eminence after superfusion

Summary Isolated medio-basal hypothalami of adult rats were continuously superfused in a chamber with controllable inputs and outputs, for periods from 30 to 240 min. The median eminence was prepared for transmission electron microscopy under carefully controlled conditions by immersion fixation with osmium tetroxide. The ultrastructure of superfused median eminence was compared with that of directly fixed, non-superfused median eminence. Even after 4h of superfusion, the median eminence displays remarkably well preserved histological and cytological patterns; cytomembranes, cell organelles, intercellular relationships, and extracellular spaces were remarkably similar in superfused and non-superfused tissues. As a consequence of osmium tetroxide fixation, microtubules were not observable. The ultrastructural information obtained from unstimulated rat median eminence superfused in vitro provides a basis for future morphofunctional correlations in the study of neurosecretion.     

A Golgi study of third ventricle tanycytes in the adult rodent brain

Summary Tanycytes along the third ventricle have been studied in adult rat and mouse brains with the rapid Golgi method. A tanycyte can be divided into three portions: somatic, neck, tail. The somatic portion is in the ependymal layer and frequently has thin cytoplasmic extensions. The neck portion originates from the soma and sticks into the periventricular layer. It, too, has numerous fine lamellar processes radiating from it. The neck contacts a blood vessel. Distal to this contact, the neck becomes thin and devoid of its cytoplasmic processes. This is the tail portion, which courses through hypothalamic nuclei to terminate as small bulbous swellings either on a vessel or at the pial surface.Although tanycytes occur throughout the dorsoventral extent of the ventricle, they are especially numerous ventrally. Midway down the ventricular wall, the neck processes interdigitate and form a moderately loose fabric beneath the ependyma. Proceeding ventrally, this becomes denser and thicker.Because the tails have no apparent associations with cells in the hypothalamic nuclei, the functional interactions of tanycytes with hypothalamic neuropil are probably confined to the periventricular layer.Supported by: NINDS Grants 5 RO1 NS 09001-02 NEUA, 5 TO1 NB 5309, and GM 00958, and by the Eleanor Roosevelt Cancer Foundation Research Institute.It is a pleasure to acknowledge the expert photographic assistance of Mr. Keith Johnson.     

Distribution of LHRH in the rat and mouse brain with special reference to the tanycytes

Summary The distribution of luteinizing hormone-releasing hormone (LHRH) was studied in the rat and mouse brain by means of light and electron microscopic immunohistochemistry using the peroxidase-antiperoxidase method. An immunoreactive product to LHRH antiserum was found near the blood vessels of the vascular organ of the lamina terminalis. In the arcuate nucleus-median eminence region, an immunoreactive material occurred bilaterally in the hypothalamic tissue around the tuberoinfundibular sulci. Electron microscopy revealed that immunoreactive fibers observed light microscopically contain numerous granules 100–130 nm in diameter. No immunoreactive product was located in the tanycytes of the median eminence, the perikarya of hypothalamic neurons, and the parenchyma of several circumventricular organs (subfornical organ, subcommissural organ, pineal organ, area postrema).Supported by grants from the Ministry of Education of Japan and the Ford Foundation