On the fine structure of the perivascular cells in the neural lobe of rats

Summary The purpose of this study is to investigate the ultrastructural features of perivascular cells as found in the neural lobe of the hypophysis. In particular, an inquiry was made into the nature of the relationship of such cells to neurosecretory fibers and endings. The latter, in fact, are often invaginated within the perivascular cells and enveloped by their processes; furthermore, they often reveal a certain number of empty granules as well as characteristics of degenerative nature. In the course of this study the localization of the perivascular cells has been investigated as well as that of their processes within the extensive interlobular network typical of the hypophysial neural lobe of rodents. Based on the data gathered, the hypothesis is put forward that the perivascular cells play an important role in the turnover of neurosecretory endings, both under physiological and experimental conditions, contributing thereby to the release of post-hypophysial hormones.With the technical assistance of Vincenzo Panetta.     

The distribution of orthogonal arrays and their relationship to intercellular junctions in neuroglia of the freeze-fractured hypothalamo-neurohypophysial system

Summary Using freeze-fracture techniques, we have investigated membrane specializations of the glia associated with the hypothalamo-neurohypophysial system of the rat. In the paraventricular (PVN) and supraoptic (SON) nuclei, astrocytes in areas of high neuronal density (i.e., magnocellular regions) display orthogonal arrays of 6–7 nm particles soley near gap junctions, while astrocytes in areas of lower neuronal density (i.e., parvocellular regions) contain additional arrays in membranes not displaying gap junctions. Arrays are especially numerous on astrocytic perivascular end-feet in both nuclei and in the laminations of the pial-glial limitans ventral to the SON. Ependymal cells near the PVN show arrays both on their lateral surfaces (displaying gap junctions) and on their apical surfaces (facing the CSF). Tight junctions are not noted on astrocytes or ependymal cells, but are noted on both the somas and myelin lamellae of oligodendroglia. Both of these latter membranes occasionally contain gap junctions as well; however, orthogonal arrays are never noted on oligodendroglia.The plasma membranes of pituicytes in the neurohypophysis display gap junctions, complex junctions, and tight junctions. Orthogonal arrays are noted near the first two of these, but not near the last. Arrays in the neural lobe appear most dense on membranes adjacent to subpial or perivascular spaces. Pituicyte membranes containing orthogonal arrays appear infrequently near the neural stalk, increasing towards the distal end of the neural lobe. The distribution of orthogonal arrays in this system, as well as in other systems in which they have been noted, suggests a polarization of membrane activity.     

Ultrastructural features of rat pituicytes in organotypic culture

Summary Neural lobes of adult rats have been explanted and organ-cultured for 5–10 days. While perivascular cells and neurosecretory fibers undergo a progressive degeneration, scattered pituicyte-like cells are observed mostly associated with each other in number of two or three cells for group. Cultured pituicytes are quite similar to in vivo pituicytes except for some particular features as the shape of the nucleus and the number of dense bodies. Furthermore they share with the in vivo pituicytes the phagocytic capacity which appears even increased as far as neurosecretory fibers are concerned. Finally, degenerating pituicytes and free lipid droplets are observed in the intercellular spaces: the significance of these facts is discussed and tentatively interpreted. The organotypic culture of the neural lobe seems to represent an experimental model useful to obtain a relatively pure population of pituicytes.With the technical assistance of Vincenzo Panetta.     

Effects of hormone-releasing stimuli on the area of the perivascular space in the neural lobe of the rat

Summary Neural lobes from rats subjected to neurohypophysial hormone-releasing stimuli were examined electron microscopically following fixation in 4 % tannic acid in 2.5 % glutaraldehyde. This fixation allowed the delineation of the perivascular space in the neural lobe tissue. Measurement of the area of the perivascular space showed that it was significantly increased in the rats subjected to vagal stimulation and intraarterial calcium ions compared to the control rats. The rats which had been subjected to haemorrhage as a hormonereleasing stimulus did not show any significant change in the area of the perivascular space. The significance of these findings in relation to hormone release is discussed.     

Organization and development of the perivascular space system in the neurohypophysis of the laboratory mouse

Summary The organization of the system of perivascular space around the capillaries in the neurohypophysis was studied in the adult and developing laboratory mouse by the use of histological silver impregnation and electron microscopical techniques.In the median eminence short and long extensions, arising mainly from the shallow space around capillary loops of the primary plexus of the portal system, formed radiations into the adjacent neural tissue of the external zone. The tissue of the neural lobe was separable into non-vascular regions dominated by undilated portions of neurosecretory nerve fibres and pituicytes, and neurovascular regions with perivascular space extensions forming an extensive system of connections between neighbouring capillaries.In the median eminence, the system of extensions of the perivascular space was estimated to increase the neurovascular contact surface area by at least 50%, implying an increased efficiency of the organ without a notable increase of its volume. The possibility that the ramifications of the perivascular space imply an enhanced uptake rate into the bloodstream and a subsequent increased concentration of the neurohormones in the portal blood, was discussed.During development of the median eminence, differentiation of perivascular space extensions of the adult type started in the juvenile of about three weeks of age, when shallow capillary loops had been formed. In the neural lobe, perivascular space ramifications were already present when the internal capillaries were formed and were fairly frequent in ten-day young. At the age of three to four weeks the organization of the system was similar to that of the adult animal.     

Light- and electron-microscopic studies on the secretory cytology of the adenohypophysis of the Japanese quail,Coturnix coturnix japonica

The effects of thyroidectomy, adrenalectomy, and castration on the pars distalis of male Japanese quail, and of injection of LH-RH on sexually inactive females, were investigated by light and electron microscopy. Correlation between light and electron microscopy was attained by use of alternate thin and thick sections. Six types of secretory cells were identified and the ultrastructural characteristics described. Putative endocrine functions have been designated on the basis of responses to experimental interventions and on other criteria. The putative STH cells are characterized by the presence of large dense secretory granules (250-300 nm) that are stained with orange-G by the trichrome method. They occur only in the caudal lobe and appear to be unchanged by castration, thyroidectomy, adrenalectomy and LH-RH injection. The putative prolactin cells are characterized by large (400-600 nm), spherical or polmorphic, dense secretory granules stainable with acid fuchsin and aniline blue; prominent Golgi apparatus and well developed endoplasmic reticulum with densely packed, regularly parallel lamellae. They are found mainly in the cephalic lobe. The prolactin cells develop some vacuolization after adrenalectomy and undergo some degeneration after castration. The ACTH cells, which are restricted to the cephalic lobe, are identified by the dense, spherical granules (250-300 nm) that are stained with acid fuchsin. After adrenalectomy, they lose their secretory granules and are transformed into large, chromophobic adrenalectomy cells. TSH cells are so designated by their response to thyroidectomy including loss of their fine secretory granules and transformation to large, vacuolated thyroidectomy cells. We have found TSH cells and thyroidectomy cells only in the cephalic lobe. Basophilic cells, considered to be gonadotropes, occur in both the cephalic and caudal lobes. The gonadotropes of the cephalic lobe appear to have slightly larger (120-200 nm) granules than the caudal lobe (120-150 nm). However, after castration, the gonadotropes in both lobes become hypertrophied and vacuolated and are transformed into mutually indistinguishable castration cells. Twenty minutes after injection with LH-RH, the gonadotropes of both lobes increase in size and number, degranulate, develop vacuoles in the cytoplasm, and appear very similar to castration cells.     

Golgi-like immunostaining of pituicytes and tanycytes positive for glial fibrillary acidic protein in the neurohypophysis of the Mongolian gerbil (Meriones unguiculatus)

Glial cells that contain the glial fibrillary acidic protein (GFAP; the major protein constituent of glial filaments) were stained immunohistochemically in thick frozen sections of the neurohypophysis of the Mongolian gerbil (Meriones unguiculatus). The resulting Golgi-like images provided informations on cytological features and distributional patterns of tanycytes and pituicytes. In the proximal median eminence, numerous bundled processes of tanycytes were revealed. They emerged from the ependymal and sub-ependymal layer and mostly reached the brain surface. Several tanycytic processes extended into the anatomical neural stalk. In the whole neural lobe, a dense network of GFAP-immunoreactive pituicyte processes was visualized. Stained pituicytes were highly asymmetric and exhibited a great morphological variability. Immunopositive fibers which were encountered in the intermediate lobe might be derived from pituicytes. Electron-microscopically further evidence was obtained that GFAP-positive pituicytes correspond to filament-rich fibrous pituicytes at the ultrastructural level.     

Golgi-like immunostaining of pituicytes and tanycytes positive for glial fibrillary acidic protein in the neurohypophysis of the Mongolian gerbil (Meriones unguiculatus)

Summary Glial cells that contain the glial fibrillary acidic protein (GFAP; the major protein constituent of glial filaments) were stained immunohistochemically in thick frozen sections of the neurohypophysis of the Mongolian gerbil (Meriones unguiculatus). The resulting Golgi-like images provided informations on cytological features and distributional patterns of tanycytes and pituicytes. In the proximal median eminence, numerous bundled processes of tanycytes were revealed. They emerged from the ependymal and subependymal layer and mostly reached the brain surface. Several tanycytic processes extended into the anatomical neural stalk. In the whole neural lobe, a dense network of GFAP-immunoreactive pituicyte processes was visualized. Stained pituicytes were highly asymmetric and exhibited a great morphological variability. Immunopositive fibers which were encountered in the intermediate lobe might be derived from pituicytes. Electron-microscopically further evidence was obtained that GFAP-positive pituicytes correspond to filament-rich fibrous pituicytes at the ultrastructural level.     

Ultrastructure of the APUD-like cells of the frog thymus gland

Within the thymus gland of the European common frog, Rana temporaria, cells with endocrine- like appearance have been found. At the ultrastructural level the most characteristic feature of their cytoplasm is the presence of secretory granules. Some cells possess irregular electron lucent granules with an eccentrically located dense core while others possess smaller electron dense granules. The cytoplasm contains also cisterns of rough endoplasmic reticulum, free ribosomes, and small mitochondria. The cells possess irregular nuclei with the pronounced nucleoli. These endocrine-like cells are connected by desmosomes with neighbouring non-granulated epithelial cells. Ultrastructural features of the cells described here resemble those seen in polypeptide hormone-secreting cells belonging to the family of cells of the APUD (Amine Precursor Uptake and Decarboxylation) series.     

Control of the pars intermedia of the lizard,Anolis carolinensis

Summary The distribution of the tracer substance horseradish peroxidase (HRP, Mw 40,000) in the neuro-intermediate lobe of the lizard, Anolis carolinensis, was studied at various time intervals (13 min to 24 h) after vascular injection. HRP rapidly entered the extracellular lumen of the neural lobe, but did not penetrate into the third ventricle. The tracer was found in micropinocytotic vesicles (MPVs) of ependymal cells within 13 min after injection. The number of cellular inclusions containing HRP increased during the period of observation (24 h). The tracer was sparsely taken up by aminergic and peptidergic nerve terminals of the external layer. After transection of the hypophysial stalk, numerous dense, labelled droplets were found in the peptidergic terminals, and the number of labelled inclusions in ependymal cells increased.MPVs were frequently found in extensions of stellate cells of the intermediate lobe, and endocytotic vacuoles (EVs) developed especially in the perikaryon. HRP was also found in large cisternae of the secretory cells, appearing predominantly towards the perivascular septum (PVS). These cisternae were found to communicate with the extracellular lumen, probably representing a system of the extracellular space extending into the secretory cell. After transection of the hypophysial stalk, there was an increase in the number of small EVs in secretory cells of the intermediate lobe.The results are discussed in terms of MSH-release regulation and possible participation of the extracellular lumen, glial and stellate cells in the transport of regulating factors and secretory material.Supported by grants from the Swedish Natural Science Research Council (to Dr. Patrick Meurling) and the Royal Physiographic Society of LundThe author is indebted to Mrs. Lena Sandell and Miss Inger Norling for excellent technical assistance and photographic aid; and to Dr. Rolf Libelius and Dr. Ingmar Lundquist for generous advice and stimulating discussions concerning the tracer technique     

A light and electron microscopic study of the pineal gland of the ground squirrel, Citellus tridecemlineatus.

The pineal gland of the 13-lined ground squirrel (Citellus tridecemlineatus) has been examined at the light and electron microscopic level. This gland is composed of low-density parenchymal cells interspersed among which are occasional glial, vascular and neural elements. Punctuating the glandular parenchymal mass are prominent perivascular and intercellular spaces. The parenchymal cells possess numerous mitochondria and less prominent profiles of rough and smooth-surfaced endoplasmic reticulum. Golgi apparatus, microtubules and lipid droplets of varying size and electron density constitute regular cytoplasmic features, with dense-core vesicles being present occasionally. The parenchymal cells have numberous processes. One among these in each cell extends for several micra to terminate in a bulbous expansion containing both clear and dense-core vesicles and occasional electron-dense inclusions. These bulbous terminals are found within the perivascular and intercellular spaces where they course in close proximity to both other parenchymal elements and axon terminals. Glial cells and their processes invest the pineal periphery and incompletely separate the parenchymal cells.     

THE NATURE OF HYPOTHALAMO-NEUROHYPOPHYSEAL NEUROSECRETION IN THE RAT : A Study by Light- and Electron Microscope Autoradiography

The nature of hypothalamo-neurohypophyseal neurosecretion was examined in the rat by means of intraventricular injections of tritiated amino acids. Quantitation of autoradiographs was used at the light microscope level to study the sites of synthesis of proteins and their time of arrival in the neural lobe. Electron microscope autoradiographs were used to study the labeling of neural lobe tissue. It was concluded that the great majority of the labeled material was translocated inside dense-cored granules and was probably composed mostly of neurophysins. The effect of ether anesthesia was also examined. It was found to remove the dense cores from about 20% of the granules in the neural lobe tissue, a process accompanied by the loss of most of their labeled material. The mechanism of the ether effect is discussed and compared to the normal secretion process.     

Control of the pars intermedia of the lizard,Anolis carolinensis

Summary The ultrastructure of the neural lobe of the lizard, Anolis carolinensis, was studied after fixation in a threefold aldehyde solution. The neural lobe appeared as narrow vertical diverticula separated from one another and from the pars intermedia by a continuous vascular septum. No nerves passed through this septum. The ependymal, fibrous and external layers were readily recognized. Peptidergic fibres were the main component of the fibrous layer. The peptidergic endings were in intimate contact with the ependymal cells, suggesting that the ependyma mediates the release of neural lobe peptides. The external layer contained ependymal end-feet and numerous aminergic terminals, ending directly on the perivascular basal lamina and/or on the ependymal end-feet. The functional aspects are discussed in terms of intermediate lobe control. The findings suggest that aminergic substances take part in the control of the intermedia, but do not exclude the involvement of peptide hormones.Supported by grants from the Swedish Natural Science Research Council and the Royal Physiographic Society of LundThe authors are indebted to Mrs. Lena Sandell for valuable technical assistance and to Miss Inger Norling and the late Mr. Lajos Erdös for photographic aid     

Immunohistochemical study of cells positive for glial fibrillary acidic protein (GFAP) in the human pituitary gland, with special reference to folliculo-stellate cells

The cytology and the distribution of cells which contain glial fibrillary acidic protein (GFAP) were studied immunohistochemically in thick frozen sections of human pituitary glands. Immunoreactive cells were constantly demonstrated in both neuro- and adenohypophysis. In the neural lobe, an irregular network of long GFAP-positive pituicyte processes was revealed. Within this network, some asymmetric pituicytes became visible. A variable number of cells was stained in cell cords and follicles of the pars distalis and the intermediate zone. The morphology of these cells could be studied in detail, providing strong evidence to support the hypothesis that adenohypophyseal GFAP-immunoreactive cells belong to the folliculo-stellate (FS) cell system. Cells with similar cytological features in the pars distalis or the intermediate zone were found to share common immunoreactivities against GFAP and the presumable FS cell markers vimentin and S-100 protein. Our results corroborate the notion that, in the human pituitary, GFAP can be regarded as a marker protein of pituicytes and FS cells, which is expressed at varying degrees.     

Immunohistochemical study of cells positive for glial fibrillary acidic protein (GFAP) in the human pituitary gland,with special reference to folliculo-stellate cells

Summary The cytology and the distribution of cells which contain glial fibrillary acidic protein (GFAP) were studied immunohistochemically in thick frozen sections of human pituitary glands. Immunoreactive cells were constantly demonstrated in both neuro- and adenohypophysis. In the neural lobe, an irregular network of long GFAP-positive pituicyte processes was revealed. Within this network, some asymmetric pituicytes became visible. A variable number of cells was stained in cell cords and follicles of the pars distalis and the intermediate zone. The morphology of these cells could be studied in detail, providing strong evidence to support the hypothesis that adenohypophyseal GFAP-immunoreactive cells belong to the folliculo-stellate (FS) cell system. Cells with similar cytological features in the pars distalis or the intermediate zone were found to share common immunoreactivities against GFAP and the presumable FS cell markers vimentin and S-100 protein. Our results corroborate the notion that, in the human pituitary, GFAR can be regarded as a marker protein of pituicytes and FS cells, which is expressed at varying degrees.     

Electron microscopic study of subcapsular cell hyperplasia in the adrenal glands of IQI/Jic mice.

Hyperplastic cells in subcapsular cell hyperplasia (SCH) lesion in adrenal glands of female IQI/Jic mice were examined by electron microscopy. These cells were small and polygonal, and had irregular nuclei, elongated mitochondria with lamellar cristae and dense lipid droplets. While these cells showed different features, some of them had desmosomes and basement membranes, and a few round mitochondria with tubular cristae as endocrine cells. These findings suggest that hyperplastic cells in SCH lesions might originate from endocrine blastemic cells.     

Demonstration by light and electron microscopy of capsules on gonococci recently grown in vivo.

A study by light microscopy, using Leishman's stain alone or Leishman's stain followed by nigrosin, showed the presence of capsules on gonococci of two strains subcultured from subcutaneous chambers in guinea pigs. With the Alcian blue method of preparation for electron microscopy, gonococci of both strains recently grown in vivo showed densely stained capsules on some cells, while others in the same preparation showed only irregular masses of dense material on their surfaces with strands connecting adjacent bacteria. Treatment with antiserum, complement and conglutinin revealed irregular masses and strands of extracellular material with fixatives that did not contain Alcian blue.     

THE FINE STRUCTURE OF MEISSNER's TOUCH CORPUSCLES OF HUMAN FINGERS

Thin slices of the finger pads of six individuals were fixed in buffered 1 per cent osmic acid, embedded in deaerated, nitrogenated methacrylate, and cut into thin sections for electron microscopic study. Before embedding, the slices were trimmed so as to include several digital tactile corpuscles. Some thin sections were stained in 10 per cent aqueous phosphotungstic acid solution. The principal part of Meissner's corpuscle is made up of flattened laminar cells stretching across the corpuscle in irregular layers. The perinuclear cytoplasm of these cells contains numerous small mitochondria, a sparse granular endoplasmic reticulum, and a large number of small vesicles. Nerve fibers enter the side or base of the corpuscle, lose their myelin sheaths, and follow a meandering course between the laminar cell plates. The nerve endings enter into a close appositional relationship with the flattened portions of the laminar cells. In some areas the apposed axolemma and cell membranes are slightly thickened with small vesicles located along the cell membrane or on both surfaces. These regions are interpreted as synapses. The most prominent feature of the nerve endings is an extraordinary accumulation of small mitochondria which vary in size and internal density. The nerve endings also contain vacuoles, groups of dense concentric membranes, and small dense vesicles of irregular distribution. The laminar cells are separated from one another by a dense intercellular substance of uniform thickness which also envelops the entire corpuscle. This material contains randomly oriented collagen fibers and fine fibrils bound together by a dense material at nodal points recurring at regular intervals of approximately 120 mµ. These findings are discussed in relation to the problems of the function of Meissner's corpuscle, neural material loss and replacement, and the presence of synapses.     

Ultrastructure of the Neurohypophysial Lobe of the Hagfish,Eptatretus stouti (Cyclostomata)

The neurohypophysial lobe is a thin-walled sac that, except for a few blood vessels, lacks any anatomical link with the adenohypophysis. Its wall consists of ependymal, fiber and palisade zones and is surrounded by blood vessels. The lobe is differentiated into distinct dorsal and ventral regions. The dorsal wall is doubly innervated by Gomori-positive axons arising in the anterior hypothalamus and by Gomori-negative fibers of unknown origin. Its surface is covered by an extensive vascular plexus. The ventral wall is innervated only by Gomori-negative fibers and is sparsely supplied with a few fine capillaries. All of the ependymal cells in both regions have the same ultrastructural appearance. The Gomori-positive or Type I axons are identified at the electron microscope level as fibers containing elementary granules with a diameter of 150–230 run. The Gomori-negative or Type II fibers contain dense-cored vesicles that vary from 80–125 nm in diameter. Both Type I and II fibers form synaptic-like complexes with the processes and end-feet of the ependymal cells. Type I axons also abut on the basal lamina bounding the perivascular spaces. It is suggested that the agranular reticulum of the ependymal cells may provide a transport pathway for neural products that are destined for release into the circulation. It is also possible that the ependyma itself is a target of neural activity.     

Ultrastructural demonstration of neurohaemal contacts in the internal zone of the median eminence of the Mongolian gerbil (Meriones unguiculatus): correlation with synaptophysin immunohistochemistry

Electron microscopy of the median eminence (ME) of the Mongolian gerbil (Meriones unguiculatus) revealed that, unlike most other mammalian species, abundant neurohaemal contacts were present not only in the external zone (EZ), but also in the internal zone (IZ) up to the subependymal layer. In the IZ, nerve terminals with dense core vesicles and/or small clear vesicles abutted on the outer basal lamina of the perivascular space of portal capillaries, alternating with tanycyte processes. In addition to these neurohaemal contacts, several layers of vesicle-filled varicosities surrounded the portal vasculature. An analysis of serial thin sections showed that the latter varicosities could also reach the perivascular basal lamina or contact it through small extensions in other planes of section. Apparently at least some of the nerve terminals making neurohaemal contacts were en passant in nature. A correlative investigation of synaptophysin (a major integral membrane protein of small synaptic vesicles) immunoreactivity at the light microscopical level demonstrated a conspicuously dense immunostaining around portal capillaries in both EZ and IZ of the proximal and distal ME (neural stalk). Since this perivascular accumulation of immunoreactivity coincides precisely with the ultrastructural accumulation of vesicle-filled axons which establish numerous neurohaemal contacts, it is concluded that this pattern of synaptophysin immunostaining indicates sites of neurohaemal contacts at the light microscopical level. During postnatal development, the perivascular concentration of synaptophysin immunoreactivity in the IZ appeared concomitantly with the early postnatal expansion of long portal capillary loops into the IZ.(ABSTRACT TRUNCATED AT 250 WORDS)