Immunohistochemical demonstration of a CRF-associated neurophysin in the external zone of the rat median eminence

Cell and Tissue Research - In picric acid-formalin fixed, paraplast embedded median eminence sections from normal and bilaterally adrenalectomized rats, the amount of “Gomori-positive”...     

On the relations between the hypothalamus and the anterior pituitary in Rana temporaria

Summary Dawson (1952) showed that, in Rana, many nerve terminals end round the capillaries of the venous portal system of the median eminence. Our observations not only confirm these facts but show as well that, besides the posterior lobe, the outer zone of the median eminence is also a centre of accumulation of neurosecretory material derived from the pre-optic nucleus. The amount of this neurosecretory material is moreover subject to a certain variation in connection with the seasons and with experimental conditions. The idea is put forward that the outer zone of the median eminence of Rana has to be considered as an analogical organ of the posterior lobe. Indeed, in the median eminence, the same morphological facts can be found, which are advanced for the conception that the posterior lobe is a centre of accumulation and a place of release of neurosecretory material originating from the pre-optic nucleus. On the basis of our observations we presume that the hypothalamo-hypophyseal neurosecretory system of Rana can be divided in two sub-units. System I ends in the posterior lobe; system II in the outer zone of the median eminence. The data of the literature indicate that the two systems also probably exist in the higher vertebrates. From these facts it is obvious that the neurosecretory material is transported from the pre-optic nucleus, via the tract, towards the outer zone of the median eminence.     

Electronmicroscopic immunocytochemical study on the vasopressin-containing neurons of the thirsting rat

Summary Whereas in thirsting animals the perikarya of the nucleus supraopticus are nearly empty of neurosecretory granules as evidenced by electron microscopic observation, the perikarya are heavily stained by light microscopic immunohistochemical staining. In an attempt to discover the substrate responsible for the positive immunohistochemical staining in thirsting rats, the neurons of the supraoptic nucleus of normal and long-term thirsting animals were compared by electron microscopic immunocytochemistry (indirect PAP-method). In controls all parts of the vasopressin-synthesizing neuron are filled with elementary granules which render a positive and uniform reaction after immunostaining with the indirect PAP-method. The positively reacting fibers in the external zone of the median eminence contain smaller granules than those of the tractus supraoptico-hypophyseus. Within the nucleus suprachiasmaticus, no positive reaction after immunostaining was found. In long-term thirsting animals PAP-complexes as markers of vasopressin are located over the ergastoplasm and over the few small elementary granules. The processes within the nucleus supraopticus and the ballooned axons in the internal zone of the median eminence exhibit free, i.e. non granule-bound, PAP-complexes. Findings in the nucleus suprachiasmaticus and the median eminence of thirsting animals correspond to those in controls. The neurohypophysis is almost completely devoid of PAP-labeled elementary granules.From these results it can be concluded that during thirst vasopressin synthesis is increased in the ergastoplasm and that the hormone is transported partly in a non granule-bound form. Direct contacts between neurosecretory cells and the basal lamina are found more often in thirst-stressed animals and are typical of neurohemal regions. It is discussed whether these neurohemal regions may develop transitionally under stress.Supported by the Deutsche Forschungsgemeinschaft (Grant Nr. Kr 569/1) and Stiftung Volkswagenwerk. This work was presented in part at the 72nd meeting of the Anatomische Gesellschaft, Aachen 1977     

Identification of adenohypophysiotropic neurohormone producing neurosecretory cells in Rana temporaria

Summary By means of electron microscopy, in the median eminence of Rana temporaria, the terminal arborizations of axons of six different types of neurosecretory cells, located in the pars ventralis of the tuber cinereum, were identified. In addition, phenomena connected with the release of neurosecretory material from the axon terminals of these neurosecretory cells into the blood capillaries of the median eminence are described.Preliminary results suggested the existence, in the median eminence, of additional different neurosecretory axon types which could also belong to corresponding neurosecretory cell types probably located in the apical part of the pars ventralis of the tuber cinereum. Moreover, in the external region of the normal median eminence, separate monoaminergic nerve fibres were tentatively identified. Arguments are adduced which plead (1) against the assumption that the ependyma or the pituicytes of the median eminence could produce adenohypophysiotropic hormones; (2) against the inference that the ependymal cells of the median eminence might be involved in the transport of adenohypophysiotropic hormones from the cerebrospinal fluid into the blood capillaries of the median eminence.     

Ultrastructure of the neurosecretory cells of the anterior commissural nucleus of the hypothalamus in rats

The ultrastructure of neurosecretory cells of the anterior commissural nucleus of rat hypothalamus is similar to that of the supraoptic nucleus and of the "magnocellular" part of the paraventricular nucleus. The only difference is a less expressed granular endoplasmatic reticulum and a smaller diameter of elementary neurosecretory granules (80-150 nm in diameter). Such elementary granules are characteristic of neurosecretory terminals located in the external zone of the median eminence. It is suggested that neurosecretory cells of the anterior commissural nucleus project to this neurohemal region.     

The origin of the vasopressinergic and oxytocinergic fibres of the external region of the median eminence of the rat hypophysis

Summary The origin of the vasopressinergic and oxytocinergic nerve fibres of the external region of the rat median eminence was investigated by means of hypothalamic lesions, adrenalectomy and immunocytochemistry. The results obtained in bilaterally adrenalectomized animals with complete, or incomplete, destruction of the suprachiasmatic nuclei showed that, at least, the great majority of the vasopressinergic and oxytocinergic nerve fibres of the external region of the rat median eminence do not originate from the suprachiasmatic nuclei. From the observations obtained in bilaterally adrenalectomized animals with total or subtotal destruction of both paraventricular hypothalamic nuclei, it appears that the paraventricular nuclei must be the origin of (nearly) all the vasopressinergic and oxytocinergic nerve fibres of the external region of the rat median eminence. The results strongly suggest that both types of fibres originate from all parts of the paraventricular nuclei.This investigation was supported by a grant from the Belgian Nationaal Fonds voor Geneeskundig Wetenschappelijk OnderzoekThe authors are much indebted to Prof. Dr. E. Kühn (Leuven) in whose laboratory the stereotactic operations were done     

Organization of tyrosine hydroxylase-immunoreactive neurons in the di-and mesencephalon of the American bullfrog (Rana catesbeiana) during metamorphosis

Summary We examined the immunocytochemical distribution of tyrosine hydroxylase, the rate-limiting enzyme in catecholamine synthesis, in the di-and mesencephalon of developing bullfrog tadpoles. Special attention was given to catecholaminergic innervation of the median eminence and pituitary. In premetamorphic tadpoles, tyrosine hydroxylase-immunoreactive neurons were visualized in the suprachiasmatic and infundibular hypothalamus, the ventral thalamus, and midbrain tegmentum by Taylor-Kollros stage V. The number of labeled neurons in all these areas increased as metamorphosis progressed. By mid-prometamorphosis, labeled neurons appeared in the preoptic recess organ as well as in the posterior thalamic nucleus. The majority of cells in the preoptic recess organ, as well as occasional neurons in the suprachiasmatic nucleus, exhibited labeled processes which projected through the ependymal lining of the preoptic recess to contact cerebrospinal fluid. The modified CSF-contacting neurons of the nucleus of the periventricular organ were devoid of specific staining. By late prometamorphosis, labeled fibers from the suprachiasmatic nucleus were observed projecting caudally to enter the hypothalamo-hypophysial-tract en route to innervating the median eminence and pituitary. Labeled fibers arising from the dorsal infundibular nucleus projected ventrolaterally to contribute to catecholaminergic innervation of the median eminence and pituitary. Immunoperoxidase staining of tyrosine hydroxylase-immunoreactive fibers and terminal arborizations in the median eminence were restricted to non-ependymal layers, while labeled fibers in the pituitary were observed in the pars intermedia and pars nervosa. Staining of tyrosine hydroxylase-immunoreactive fibers in the median eminence and pituitary was sparse or absent in premetamorphic tadpoles, but became increasingly more intense as metamorphosis progressed.     

Immunohistochemical and electron microscopic study of the rat hypothalamic nuclei and cell clusters under various experimental conditions

Summary In untreated, pregnant and thirsting rats the neurosecretory hypothalamic areas were investigated by means of the immunoperoxidase technique in order to demonstrate vasopressin- and oxytocin containing elements at the light- and electron microscopic level. In addition, chromalum-hematoxylinphloxin (CHP) staining and conventional double staining of ultrathin sections were used. The areas investigated included the anterior and posterior supraoptic nuclei, the paraventricular nuclei, the numerous accessory cell clusters in the region between the tractus opticus and the third ventricle as well as the median eminence. In all nuclei and in the accessory cell clusters, the number of vasopressin-reactive neurons exceeds that of oxytocin-reactive neurons. Compared with the anterior supraoptic nucleus, the posterior supraoptic nucleus and the accessory cell clusters react more heavily to prolonged thirst. In the median eminence the neurosecretory axons display close contacts with the portal vessels not only in its lateral portion but in thirsting animals also around the mid-line. There the internal layer is broadened and vasopressin-positive tanycytic processes reach the external zone. Parasagittally, fine vasopressin-positive material can be traced from the internal layer to small deposits at the portal vessels. In long term thirsting animals the typical feature of swollen axons exhibits a characteristic distribution in the median eminence and renders a distinct positive reaction to anti-vasopressin. The release of peptide hormones from the perikarya and from the axons within the nuclei as well as the mode of release within the median eminence are discussed. The significance of the positive immunostaining of the ependymal tanycytes and of some perikarya of the suprachiasmatic nucleus must be reconsidered by further studies.Supported by the Deutsche Forschungsgemeinschaft (Grant Nr. Kr 569/1) and Stiftung VolkswagenwerkDedicated to Professor Berta Scharrer on the occasion of her 70th birthdayThe author wishes to express her special gratitude to Dr. L.A. Sternberger for supplying the peroxidaseantiper oxidase-complex and to Dr. H. Stein (Pathologisches Institut der Universität Kiel) for supplying Anti-IgG. The skilful technical assistance of Mrs. H. Prien and Mrs. H. Schöning is thankfully acknowledged     

Ontogeny of the hypothalamo-neurohypophysial system in rats--an immunohistochemical study.

In fetal rats neurophysin has been visualized immunohistochemically first at the 16th gestation day in perikaryons of the supraoptic nucleus, followed by the median eminence and the neurohypophysis at the 17th day, and the paraventricular neurons at the 19th day. The external zone of the median eminence contains abundantly immunoreactive fibres at the first days post partum. In the perikaryons of the suprachiasmatic nucleus immunoreactive material appears after the 3rd day of postnatal development.     

Reaction of the hypothalamo-hypophyseal system of cats to stimulation of the cervical sympathetic nerve and the afferent fibers of the vagus nerve]

Data are presented on the functional morphology of the hypothalamo-hypophysial neurosecretory system of cats in stimulation of the cervical sympathetic nerve and of the afferent fibers of the vagus. Stimulation of the sympathetic nerve selectively activated the supraoptic nucleus and caused the discharge of the neurohormones from the posterior lobe of the hypophysis, whereas its infundibular portion contained much neurosecretory material. In response to the stimulation of the vagus all the portions of the neurohypophysis were released of the Gomori-positive substance; both the supraoptic and the paraventricular nucleus were activated.     

Fine structure of the median eminence and the pars nervosa of the bullfrog,Rana catesbeiana

Summary The hypothalamic neurosecretory system of the bullfrog, Rana catesbeiana, was studied with light- and electron microscopy. The median eminence is roughly divided into two portions. The upper portion mostly consists of ependymal cells, glial cells and preoptico-hypophysial nerve tract, whereas in the lower portion, neurosecretory axons, glial cells, processes of glial and ependymal cells, and fine blood vessels of the hypothalamic portal vein are located. A part of the neurosecretory axons of the preoptico-hypophysial tract proceeds to the lower portion of the median eminence. These axons are arranged perpendicularly to the capillaries of the hypothalamic portal vein. The glial cells are densely located in the area of the median eminence where neurosecretory material is abundant. The neurosecretory material in the neurosecretory cells, their axons, the median eminence and the pars nervosa of the bullfrog shows a positive reaction to PAS treatment.The neurohemal area of the median eminence is occupied by many neurosecretory and non-neurosecretory axons, containing neurosecretory granules and/or synaptic vesicles. The axonal portions with the synaptic vesicles which are considered to be the nerve endings abut on the capillaries of the portal system. The size of synaptic vesicles in the axon terminals containing few neurosecretory granules is larger than those in the endings with many neurosecretory granules. Infrequently glial and ependymal processes are interposed between the nerve endings and the capillary wall.In the hilar region of the infundibulum, synapses are frequently observed between the thin fibers with or without neurosecretory granules and dendrites of non-neurosecretory neurons. The probable functions of these synapses are briefly discussed on the basis of our findings. Both in the hilar region of the infundibulum and in the pars nervosa, electron-dense neurosecretory granules of two different sizes were observed. The median eminence contains only one type of granules.The fine structure of the pars nervosa shows similar structures to those of the median eminence. Both in the median eminence and the pars nervosa, the fenestrated endothelium of the capillaries was frequently observed. The thick perivascular connective tissue space containing fibroblasts and collagen fibrils was observed both in the median eminence and the pars nervosa. Vesicles in the cytoplasm of the endothelial cells which appear to take a part in the transendothelial transport were observed.This investigation was supported in part by United States Public Health Service Research Grant, No. A-3678, to Hideshi Kobayashi from the National Institute of Arthritis and Metabolic Diseases and partly by a grant for Fundamental Scientific Research from the Ministry of Education of Japan. The authors wish to express their thanks to Prof. K. Takewaki for his kind encouragement.     

The corticotropin releasing factor (CRF) neurosecretory system in intact, adrenalectomized, and adrenalectomized-dexamethasone treated rats. An immunocytochemical analysis

In general, antisera generated against ovine CFR do not reveal immunopositive neuronal perikarya in the rat. If animals are adrenalectomized significant amounts of immunoreactive CFR are present in the hypothalamus. By using this model, we have visualized the CFR system of the rat. Intact, intact pretreated with dexamethasone, adrenalectomized, and adrenalectomized pretreated with dexamethasone animals were used in the present study. In adrenalectomized and adrenalectomized plus dexamethasone treated animals the CFR-immunopositive neurons were observed in the parvocellular portion of the paraventricular nucleus. Distinct pathways of CRF fibers could be seen emerging from this hypothalamic nucleus. The greatest number of these fibers exited the PVN laterally and crossed either superior to or beneath the fibers of the fornix. The fibers then turned ventrally and cascaded to form a bundle of fibers above the superio-lateral margin of the optic chiasm. They turned caudally and followed the optic tract. As these fibers reached the level of the anterior median eminence, they turned medially to run along the inferior margin of the hypothalamus and enter the median eminence. A few fibers emerged from the PVN along the periventricular margin of the third ventricle, traveled caudally in the periventricular nucleus and entered the median eminence. Adrenalectomized and adrenalectomized-dexamethasone treated rats had very dense accumulations of immunoreactive CRF in the median eminence when compared with controls. Immunoreactive neurons and fibers were also observed in the central nucleus of the amygdala in the adrenalectomized and adrenalectomized-dexamethasone treated animals.     

Ultrastructural characteristics of immunolabelled,corticotropin releasing factor (CRF)-synthesizing neurons in the rat brain

Summary The corticotropin releasing factor (CRF)-synthesizing perikarya and neural processes were detected at ultrastructural level in the hypothalamic paraventricular nucleus and in the median eminence of control and colchicine-pretreated rats. The unlabelled antibody peroxidase-antiperoxidase complex (PAP) immunohistochemical method was used in a pre-embedding manner, on thick, non-frozen sections. In CRF-perikarya, neurosecretory granules (80–120 nm in diameter), free ribosomes, and the rough endoplasmic reticulum were labelled. Unlabelled axon terminals formed asymmetric synapses on CRF-containing perikarya and dendrites. Immunolabelled axons terminated in the palisadic zone of the median eminence.     

Effects of adrenalectomy and excess hydrocortisone on the incorporation of 35S-labelled cysteine in the hypothalamic-neurohypophyseal neurosecretory system of the rat

Summary The incorporation of ³⁵S-labelled cysteine in the hypothalamic-neurohypophyseal system was studied in normal and adrenalectomized rats and in rats treated with excess hydrocortisone. Labelled cysteine was intraperitoneally administered and grain counts were made of autoradiographs produced from sections of the supraoptic and paraventricular nucleus, median eminence and neurohypophysis of animals killed 45 min., 4 hours and 24 hours after administration of the labelled substance. On the whole, lower incorporation levels of the label were noted in the adrenalectomized rats, compared with the controls. In the rats treated with excess hydrocortisone, the grain counts at 45 min and 4 hours after injection were higher and those at 24 hours were lower than those of the controls.The findings are discussed, among other things, in terms of rate of uptake vs. time and related to previous reports on the cysteine uptake and neurosecretory activity of the hypothalamic-neurosecretory sytem.This study was financially supported by the Sigrid Jusélius Foundation, Helsinki, Finland and the National Research Council for Medical Sciences, Finland.     

Secretory glycoproteins of the subcommissural organ of the dogfish (Scyliorhinus canicula): evidence for the existence of precursor and processed forms

The probable presence of oxytocin in the hypothalamo-hypophysial system of two reptilian species, the snake Natrix maura and the turtle Mauremys caspica, was re-investigated. A high-pressure liquid chromatographic analysis of the turtle neural lobe revealed the existence of vasotocin, mesotocin, and a third compound co-eluting with oxytocin. Brains from both species were fixed by vascular perfusion with Bouin's fluid. Adjacent paraffin sections were immunostained using antisera against the following substances: (1) bovine oxytocin-neurophysin; (2) a mixture of bovine oxytocin-neurophysin and vasopressin-neurophysin; (3) dogfish neurophysins; (4) oxytocin; (5) arginine-vasotocin; (6) mesotocin; (7) somatostatin. Immunoreactivity against oxytocin was found in parvocellular neurons of the snake suprachiasmatic nucleus and cerebrospinal-fluid contacting neurons of the medial nucleus of the infundibular recess of both species, the latter immunoreactivity being much more conspicuous in the turtle. Numerous fibers containing immunoreactive oxytocin extended between the medial nucleus of the infundibular recess, and the internal region of the medium eminence and the neural lobe. The oxytocin-immunoreactivity in all locations was completely abolished by preabsorption of the anti-oxytocin serum with three different oxytocin preparations. None of the neurons of the suprachiasmatic and medial nucleus of the infundibular recess, including the oxytocin-immunoreactive elements, reacted with either the antineurophysin sera used, or the anti-vasotocin or anti-mesotocin antibodies. The possible existence of a reptilian oxytocin-neurophysin is discussed. The alternative that, in the reptilian hypothalamus, neurons synthesize a compound closely related to, but different from oxytocin is also considered.     

The effect of castration upon the ultrastructure of the rat hypothalamus

Summary The neurosecretory hypothalamic nuclei and the inner zone of the median eminence of castrated rats were studied under the electron microscope. After one month of castration all the neurosecretory neurons of both nuclei show signs of hyperactivity characterized by dilated cisternae of the endoplasmic reticulum containing a macromolecular filamentous material and an increase in the number of ribosomes. After six months of castration, some neurosecretory neurons show an increased number of neurotubules and larger lysosomes than in the controls. Other neurons show a very significant hypertrophy of the endoplasmic reticulum, with large amounts of intracisternal filamentous material. These cells have few neurosecretory granules and in the adjacent synapses the number of granulated vesicles is increased. In the supraoptic nucleus there are two kinds of neurosecretory axons: the clear ones, which are similar to those that appear in control animals and the dark ones, which have smaller elementary granules. In the inner zone of the median eminence the axons show an increase in the number of neurosecretory granules with respect to the controls. After supplementary administration of sexual hormones, all the modifications produced by castration disappear. The ultrastructural changes observed in the neurosecretory nuclei after castration are discussed in relation to those previously described in the neurohypophysis under the same experimental conditions. A feedback regulatory action of sex hormones on hypothalamic neurosecretory neurons is postulated.Supported by grants from the Consejo Nacional de Investigaciones Científicas y Técnicas and by the Air Force Office of Scientific Research (AF-AFOSR 963-67).We are deeply indebted to Mrs. Defilippi-Novoa and Mr. Alberto Saenz for their skillful assistence.     

Morphology and function of capillary networks in subregions of the rat tuber cinereum

Summary The differentiated cytology, cytochemistry, and functions within subdivisions of the tuber cinereum prompted this morphometric and physiological investigation of capillaries in the median eminence and arcuate nucleus of albino rats. Morphometric studies established that the external zone of the median eminence had 3–5 times the number and surface area of true and sinusoidal capillaries than the internal or subependymal median eminence zones, or either of two subdivisions examined in the arcuate nucleus. Type-I true capillaries, around which Virchow-Robin spaces comprise 1% of arcuate tissue area, were situated proximally to the median eminence border. This finding is consistent with a premise that confluent pericapillary spaces enable infiltration of arcuate neurons by factors from capillary blood from the median eminence or Virchow-Robin spaces. Physiologically, the rate of penetration across the median eminence capillaries by blood-borne [¹⁴C]-aminoisobutyric acid (a neutral amino acid used as a capillary permeability tracer) was 142 times greater than for capillaries in the distal arcuate nucleus within 12 s of tracer administration. A new finding was that the proximal arcuate nucleus had a permeability x surface area product of 69 l g^–1 min^–1, 34 times greater than that in more distal aspects of the tuber where blood-brain barrier properties exist. We also found that the microcirculatory transit time of a plasma space marker, [¹⁴C]sucrose, was considerably longer (1.2 s) in the median eminence and proximal arcuate nucleus than in the distal arcuate or ventromedial nucleus (0.4 s). By virtue of its high capillary permeability and extensive blood-tissue surface area, including the wide Virchow-Robin spaces, the median eminence external zone could be a gateway for flooding other tuberal compartments with blood-borne factors. This effect may be compounded by capillary bed specializations in the proximal arcuate nucleus where Type-I true capillaries, Type-III sinusoids, and pericapillary spaces are confluent with those in the median eminence. The results indicate that the proximal arcuate parenchyma could be exposed to circulating neuroactive substances on a moment-to-moment basis.Dedicated to Dr. Milton W. Brightman of Bethesda, Maryland, USA on the occasion of his 67th birthday and tribute as Craigie scholar at the First Craigie Conference on Brain Capillaries, Toronto, Ontario, June 24, 1990     

Regeneration of the supraoptico-hypophyseal and paraventriculo-hypophyseal tracts in the hypophysectomized rat

Summary The effect of hypophysectomy on the nerve fiber pattern in the median eminence and infundibular stem of the rat has been investigated by a slightly modified Bodian technique. Postoperative changes in the distribution of neurosecretory material and connective tissue and changes in vascularity have also been studied.Extensive regeneration of the fibers of the supraoptico-hypophyseal and paraventriculo-hypophyseal tract could be demonstrated. It is most pronounced at the distal extremity of the infundibular stem but occurs also in the rostral part of the infundibular stem and in the median eminence. Regeneration starts in the second postoperative week and is completed about four weeks later. The nervous regeneration observed in the pituitary area after hypophysectomy is more extensive than is usually encountered after lesions elsewhere in the central nervous system.It could be demonstrated moreover that neurosecretory material accumulates at the same sites in which the terminals of the regenerated nerve fibers can be found. Hypophysectomy also causes an increase in capillary density and connective tissue content of the infundibular stem. Accumulations of neurosecretory material are always found in areas showing a high capillary density and a considerable amount of connective tissue.Factors which might be responsible for the extensive nervous regeneration in the pituitary area are discussed as are the factors determining the pattern of outgrowth of the regenerating nerve fibers. Morphological aspects of storage of posterior pituitary hormones are considered in the light of data in the literature and the results of the present work.Partly supported by a U.S. Public Health grant to Dr. E. Scharrer and a travel grant to the author from the Netherlands Organization for pure scientific Research (ZWO).     

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.     

The zinc iodide-osmium tetroxide (ZIO) reaction on nerve endings in the median eminence of the rat under normal and experimental conditions

Summary The reaction of nerve endings in the median eminence of the rat to zinc iodide-osmium tetroxide (ZIO) staining was examined electron microscopically under normal and experimental conditions. The experimental condition of catecholamine exhaustion in the nerve endings was induced by the administration of H44/68 and reserpine. Vesicles in the terminals of catecholaminergic nerves reacted similarly to ZIO staining in both normal and experimental material. The majority of synaptic vesicles in various terminals gave a positive ZIO reaction. The neurosecretory elementary granules, however, failed to react with ZIO. On the other hand, some nerve terminals in the external layer of the median eminence showed a strong positive reaction in the cytoplasmic matrix, in mitochondria as well as in synaptic vesicles. These findings strongly suggest that the ZIO-positive substance in nerve terminals is not the transmitter itself, i.e. the monoamine, but rather represents a range of substances commonly found in various kinds of synaptic vesicles and is probably proteinaceous in nature. A brief discussion is also given on the difference in ZIO reactivity between neurosecretory elementary granules and small vesicles in the hypothalamo-hypophyseal tract.This work was supported in part by a research grant from the Ministry of Education, Japan