David Zambrano

The secretory cycle of supraoptic neurons in the rat

SummaryAfter perfusion with formaldehyde and glutaraldehyde the supraoptic nucleus and infundibular process of the neurohypophysis of the rat were dissected and prepared for electronmicroscope observation. This study was carried out in specimens under normal water balance, in others fed on dry food and in rats submitted to forced hydration.Two extreme types of neurons with intermediary stages were recognized in the normal supraoptic nucleus. The main difference between them is in the content of ribosomes, development and dilatation of the vacuolar system and in the number of elementary neurosecretory granules. In both types lysosome-like particles are observed. The volume of the elementary granules increases 1.7 times along the hypothalamic-hypophyseal tract while the increase in the dense core of the granule is of the order of 4.3 times.After forty-eight hours on dry food there is a general depletion of secretory granules from the perikaryon and nearby axons, the ribosomes are numerous and the endoplasmic reticulum is dilated in all cells and contains a macromolecular filamentous material. With more prolonged dehydration the neurosecretory granules reappear in relation to the Golgi complex and the vacuolar system becomes progressively flattened. With forced hydration the number of granules in the perikaryon increases considerably.These observations are interpreted as indicative that the early stages of synthesis take place at the level of the ribosomes. The product, in a dilute macromolecular form, is transferred into the cisternae of the endoplasmic reticulum and then condensed into granules within the Golgi complex. The increase in size of the granules along the axon is discussed in relation to the progressive increase in hormone content.

Ultrastructure of transitional epithelium of man

SummaryBlocks of human normal renal pelvis and ureter obtained at the time of surgery were fixed in glutaraldehyde and osmium with or without ruthenium red, for electron microscopic observations. The transitional epithelium is arranged in three cell layers: basal, intermediate and superficial. All epithelial cells show numerous microvilli and contain the characteristic vesicles of transitional epithelium, bundles of cytoplasmic filaments, microtubules and numerous free ribosomes. The epithelial extracellular compartment is notably large and appears as an intricate, tridimensional network of canaliculi and cisternae which are wider in the intermediate and superficial layers and in which microvilli and cytoplasmic folds of vicinal cells are often attached or interdigitated. At these sites there are desmosomes.The surface of all transitional epithelial cells is covered by a fibrillar mucous coat which is more developed at the plasmalemma of the free border of luminal cells in which microvilli are also seen. Ruthenium red stains selectively the plasmalemma and the mucous coat of the free surface of the epithelium, indicating the presence of an acid polysaccharide. With this technic (Luft, 1965), it is observed, radiating from the plasmalemma, branching filaments which measure 100 Å in diameter forming a zone of varying density which is about 400 mμ wide and which corresponds, at the light microscopic level, to the luminal border of the transitional epithelial cells in which a sialomucin has been identified. The slender filaments have a beaded appearance. At the free border, superficial cells are attached by functional complexes in which tight junctions seal the epithelial intercellular space, which is opened at the level of the basement membrane where only desmosomes are observed.The ultrastructure of human transitional epithelium of urinary tract resembles the duct cells of the salt gland of certain marine birds (Fawcett, 1962) and the amphibian epidermis (Farquhar and Palade, 1965) in which there are active processes of transport. The mucous surface coat, selectively stained by the ruthenium red, contains a sialomucin (Monis and Dorfman, 1965, 1967).The ultrastructure and histochemistry of the mucous fluffy coat of man transitional epithelium and the observations of Porter and Tamm (1955), on the ultrastructure of preparations of the Tamm and Horsfall mucoprotein (1952) are bases for suggesting that transitional epithelium of urinary tract of man is the site of biosynthesis of certain urinary mucoids. Present investigations are directed to obtain evidence to substantiate this hypothesis.

NEUROSECRETORY ACTIVITY IN SUPRAOPTIC NUCLEUS OF NORMAL RATS

SummaryThe secretory rhythm of neurosecretory cells was studied in supraoptic nuclei of normally drinking rats. Both nuclei were divided in five zones, and in each one, the nuclear and nucleolar volumes of 50 neurons were measured. Parallel observations were made on the Nissl material and neurosecretory granules. Comparative effects of different fixatives showed Carnoy and Zenker to be the best fluids for this type of investigation.Two extreme types of neurosecretory neurons with intermediary stages are observed. Type I neuron has a small nucleus and nucleolus, well developed Nissl bodies and a greater content of neurosecretory material. Type II has larger nucleus and nucleolus, scanty or packed Nissl bodies, and little Gomori positive material. The nuclear and nucleolar volumes differ between the different zones of the supraoptic nucleus and the zone of maximal activity varies in each particular case. To explain this asynchronic behavior a “neurosecretory wave” of activity moving along the supraoptic nucleus is postulated.

Ultrastructural aspects of the inhibition of neurosecretion by puromycin

SummaryThe supraoptic nuclei and the infundibular process of the neurohypophysis were studied under the electron microscope in rats fed on dried food for 1 to 3 days and in similarly dehydrated animals but injected subarachnoidally with puromycin 4 to 24 hours prior to fixation. With 4 to 8 hours of puromycin the intracisternal filamentous material found in the control is greatly diminished. After 24 hours this inhibitory effect disappears in the perikaryon but the neurotubular material of the neuronal processes within the supraoptic nucleus and the neurosecretion in the infundibular process are greatly reduced. These results are discussed taking into consideration the inhibition of protein synthesis caused by puromycin and the neurosecretory cycle which characterizes the supraoptic neurons.

Ultrastructure of the pars distalis of the lizard Anolis carolinensis with special reference to the identification of the gonadotropic cell

SummaryFive categories of granulated cells were distinguished by their ultrastructural features, and quantitative analyses were made of the pars distalis cells in normal and castrated lizards. The gonadotropin-producing cell was identified on the basis of its uniform distribution in the gland as well as from cytological changes resulting from castration. The secretory granules of the gonadotropic cell vary in size (100–500 mμ) and density, and lipid bodies are commonly present. Following castration, the endoplasmic reticulum proliferates, forming many small, rough-surfaced, dilated cisternae which do not coalesce greatly as in other vertebrate species. Degranulation is accompanied by hypertrophy and hyperplasia of the mitochondria and by the appearance in the cytoplasm of conspicuous clusters of microfilaments. The designated gonadotropic cell was the only class of secretory cell showing consistent changes following three weeks of castration.In addition to the uniformly distributed gonadotrope cell, two secretory cells occur mainly in the rostral half of the gland, and two in the caudal half. Tentative identification of the cell types is discussed in the light of available information on the localization of the hormones in the pars distalis of this species.

Transitional epithelium of urinary tract in normal and dehydrated rats

SummaryThis report is a light microscopic histochemical and fine structural study of transitional epithelium of the urinary tract of normal and dehydrated rats. Four types of cells were recognized: basal, intermediate, squamous or luminal and bundle cells. The transitional epithelium of normal rat ureter and bladder shows distinct cytoplasmic staining of the squamous cells layer by PAS. The luminal free border stains more intensely with PAS. With the electron microscope, abundant cytoplasmic tonofilaments, free ribosomes and the characteristic thick-walled fusiform and round vesicles are observed, which were in greater number in the squamous cells. Lysosomes are identified with PAS, and Toluidine Blue 0, by their content of acid phosphatase and non-specific carboxylic esterase, and by their ultrastructural appearance. The bundle cell (Hicks, 1965) is characterized by histochemical technics. These cells form about 2.5% of the total cell population of normal transitional epithelium. The bundle cell contains basophilic metachromatic granules, which indicates the presence of a weakly acid mucosubstance. It is suggested that bundle cell granules are released in the intercellular spaces of transitional epithelium and that the mucosubstance may regulate flow of ions and metabolites in the epithelial intercellular channels.Several ultrastructural changes occur in the transitional epithelium of dehydrated rats: marked increase in number of thick-walled vesicles, development of polysomes, relative increase of cytoplasmic filaments and greater number of enlarged lysosomes. Bundle cells decrease in number. These ultrastructural changes promptly regressed by allowing the animal to drink water.It is suggested that the rate of formation of the characteristic vesicles of transitional epithelium, a function of membrane synthesis, may be under the control of the antidiuretic hormone.

Histology and ultrastructure of the neurohypophysis of the south american lungfish, Lepidosiren paradoxa

SummaryThe neurohypophysis of the South American lungfish Lepidosiren paradoxa has been studied with light and electron microscopy, including the Falck-Hillarp technique for catecholamines. The pars nervosa hypophyseos is a well-marked, dorsally located subdivision of the pituitary gland composed of lobes or follicles, each one constituted of a central core of ependymal cells, a subependymal hilar region made up of nerve fibers and a peripheric palisade zone of nerve endings which contact capillary vessels. Four types of neurosecretory axons can be distinguished under the electron microscope. Type I, the most common, contains spherical elementary granules of high electron density, 1500–1800 Å in diameter. The scarce type II axons contain irregularly-shaped elementary granules. Type III contains only small clear vesicles, 400–600 Å in diameter. Type IV, mostly present in regions of the gland contacting the pars intermedia, contain large granulated vesicles, 900–1000 Å in diameter. The Falck-Hillarp technique revealed axons with a positive reaction for catecholamines at sites corresponding approximately to the location of type IV of the electron microscope.Ependymal cells are of large size, linking the cerebrospinal fluid, the nerve endings and the blood vessels. A conspicuous membrane-bound, spherical dense material, 1400–2000 Å in diameter, is observed in both the apical and vascular processes of these cells. The ependymal processes which traverse the hilar and palisade regions contain structures resembling degenerated neurosecretory axons. These results are discussed in relation with the currently available information on the comparative anatomy of the pars nervosa. The possible functional significance of ependymal cells and of each type of axon are also discussed.This study was aided by the following grants: NIH NS 06953 to Prof. De Robertis, Consejo Nacional de Investigaciones Científicas y Técnicas to Prof. Zambrano, Comisión de Investigaciones Científicas de la Provincia de Buenos Aires and Comisión de Investigaciones Cientificas de la Universidad Nacional de la Plata: to Prof. Iturriza.The authors are indebted to Prof. De Robertis for his generosity in granting us his laboratory facilities, and to Dr. F. J. J. Risso and Mr. A. Fernández (Resistencia, Chaco) who provided the specimens used in this study. The able microtechnical assistance of Miss L. Riboldazzi and Mrs. R. Raña and the photographic work of Mr. A. Saenz are much appreciated.

Ultrastructure of rat pituitary gonadotrophs following incubations of the gland with synthetic LH-RH

SummaryThe action of luteinizing releasing hormone (LH-RH) on FSH and LH-gonadotrophs from the anterior pituitary glands of normal rats has been studied with electron microscopy. After 15 min. incubation with LH-RH, LH cells showed secretory granules undergoing exocytosis, but FSH did not appear to release secretory products. On the other hand, incubations with the releasing hormone for 3 and 6 hr stimulated exocytosis from both FSH and LH-gonadotrophs. Of more interest was the finding that FSH cells appeared as typical “signet cells” of castration when glands were incubated for 3 and 6 hr with the releasing hormone. The appearance of such “signet cells” was prevented if puromycin was added to the medium together with LH-RH.

Appearance of granules in the Purkinje cells of dehydrated toads (Bufo arenarum Hensel).

SummarySeven days of dehydration produce alterations in the Purkinje cells of the toad, Bufo arenarumHensel. Aldehyde-fuchsin-positive granules appear in the cytoplasm of the cells and at the same time modifications take place in cytoplasmatic and nuclear ribonucleoproteins. These modifications consist of an increase in nucleolar volume and appearance of a nuclear cap, polarized towards the granules.