Immunocytochemical and morphometrical study of thyrotropic cells of Rana ridibunda

Summary Indirect immunoflorescence and PAP techniques for light microscopy as well as the immunogold complex technique for electron microscopy were used to localize and identify thyrotropic (TSH) producing cells in the pars distalis of Rana ridibunda. A double immunostaining procedure was used to distinguish TSH cells from other glycoprotein hormone producing cells. Rabbit anti-human--TSH was used as the primary antiserum and revealed a basophil, PAS and alcian blue positive cell type in the ventro-central zone of the gland. Under the electron microscope, TSH cells show irregular morphology, polymorphic secretory granules with diameters ranging between 120 and 375 nm and poor development of the endoplasmic reticulum and Golgi complex; they are usually polarized towards capillaries. Ultrastructural morphometry (point-counting method) was used to evaluate stereological parameters of rough endoplasmic reticulum, Golgi complex, secretory granules and mitochondria.This work has been supported by grant 2184-83 from the Comisión Asesora (CAICYT) of Spain     

Immunohistochemical detection of adenohypophyseal cells containing hormones in Rana ridibunda

Summary The pars distalis of the pituitary of Rana ridibunda captured throughout the spring and summer was examined with immunofluorescence techniques using antisera to mammalian pituitary hormones. On the basis of their immunoreactivity, four different cell types are recognized: 1) cells immunoreactive to anti-bovine LTH, 2) cells immunoreactive to anti-bovine STH, 3) cells immunoreactive to anti-ovine LH, and 4) cells immunoreactive to anti-synthetic ACTH (1–24). Their distribution and morphology as well as their staining characteristics (classical histological techniques) are reported in this study.     

The autonomic innervation of Rana ridibunda intestine

1. The innervation of Rana ridibunda intestine has been studied by the following methods: (a) demonstration of cholinesterase activity; (b) FIF method for catecholamines; (c) immunohistochemistry for VIP, SP and SOM, and (d) conventional electron microscopy. 2. The intrinsic intestinal innervation is represented by cholinergic-, VIPergic-, SP- and SOM-like plexuses. The intestinal adrenergic component is of extrinsic origin. 3. The intestinal peptidergic innervation is the most developed, the large intestine being the portion where the studied peptidergic plexuses are more widely distributed. 4. Against a poorly developed cholinergic/adrenergic innervation, it seems that there is a predominant peptidergic innervation in the amphibians intestine wall. 5. Taking into account that amphibians sacral parasympathetic as well as sympathetic innervation development are limited, it could be considered that in vertebrates the intestinal peptidergic innervation is phylogenetically earlier and hence better developed.     

Acetylcholinesterase-containing nerve cells in the pineal complex and subcommissural area of the frogs,Rana ridibunda and Rana esculenta

Summary In Rana esculenta and Rana ridibunda the frontal organ and the pineal organ (epiphysis cerebri) form a pineal complex. Approximately 60 nerve cells of the frontal organ and 220–320 nerve cells of the pineal organ display a positive acetylcholinesterase reaction (Karnovsky and Roots, 1964). The dorsal wall of the pineal organ is considerably richer in acetylcholinesterase-positive neurons than the ventral wall (ratio 31); a group of unusually large-sized nerve cells occurs in the rostral portion of the frog pineal. Two different types of nerve cells were observed in the pineal complex: multipolar and pseudounipolar elements. The former are embedded in the pineal parenchyma and their processes penetrate radially into the plexiform layer, whereas the latter are distributed along the roots of the pineal tract near the basal lamina. The ratio of the multipolar to pseudounipolar neurons is 14 for the frontal organ and 35 for the pineal organ. The multipolar elements may be interneurons; the pseudouni-polar cells send one of their processes into the pineal tract. At the caudal end of the pineal organ 30–50 unipolar nerve cells are clustered in juxtaposition with the pineal tract, and other 30–50 unipolar neurons are scattered along the basis of the subcommissural organ. Some of these nerve cells emit their processes toward the mesencephalon and others toward the pineal organ via the pineal tract. The results are discussed with respect to previous physiological and morphological findings on the pineal complex of Anura.Supported by a fellowship from the Alexander von Humboldt Foundation, Federal Republic of Germany, to K. Wake. Completed November 22, 1973.Supported by the Deutsche Forschungsgemeinschaft.     

Histochemical study on the tongue of Rana ridibunda (anuran amphibian)

A structural and histochemical study of the tongue in the Anuran Amphibian Rana ridibunda was carried out. Histochemical analysis of the filiform and fungiform papillae of the dorsal epithelium has shown a variety of cellular types which may be characterized cytochemically. All of them, except the goblet cells, show a remarkable amount of neutral mucins. The intensity of histochemical positive reaction for sulphomucins, sialomucins and protein is variable according to the cell type. Other histochemical reactions show that the lingual glands are rich in neutral mucins, but not in sialomucins. Histochemically in the component the basic proteins with sulphydryl groups are demonstrated. This sulphydryl groups are more abundant in the glands located in the deep regions.     

Urea tolerance and osmoregulation in Bufo viridis and Rana ridibunda

1. The tolerance and adaptation to urea solutions by terrestrial green toads (Bufo viridis) and semi-aquatic frogs (Rana Ridibunda) were studied. 2. the green toad showed tolerance to urea solution of 800 mM and the frogs showed tolerance only to about 400 mM urea solution. 3. The plasma concentrations of both species was hyperosmotic to the external medium in all the different urea solutions. 4. Blood osmolality, urea, Na+ and Cl- concentrations of B. viridis were always higher than in R. ridibunda. 5. The urea concentration in muscle of R. ridibunda was higher than the urea concentration in muscle of B. viridis. 6. The muscle tissue weight loss of B. viridis was significantly lower than R. ridibunda.     

The karyosphere during late oogenesis in Rana ridibunda.

The organization of the nucleus in the oocytes of Rana ridibunda was examined during late diplotene at the light and electron microscopic level. At this stage the chromosomes are relatively condensed and assembled in the centre of the nucleus, constituting a karyosphere. The chromosomes here are associated with the central "protein sphere" (15--20 microns in diameter), obviously at their telomeres. Numerous nucleoli are accumulated around the chromosomes, forming a karyosphere capsule and contain segregated fibrillar and granular components; structures resembling perinucleolar chromatin and fibrillar bodies (spherules) are associated with the nucleoli. Granules 30 to 40 nm in diameter are seen to surround the fibrillar spherules. "Nucleolus-like bodies" consisting of granules 10 to 15 nm in diameter which are embedded in finely fibrillar material are often associated in contact with the chromosomes. The central sphere is an accumulation of annular structures similar to those of the pore complexes of the nuclear envelope. These structures are bound to the chromosome material, the "nucleolus-like bodies" and the fibrillar bodies. A participation of "nucleolus-like bodies" in the formation of the central sphere is suggested. A possible role of the nuclear protein matrix in the construction of the karyosphere elements is discussed.     

Cardiac minute volume and its pattern in Rana ridibunda frogs

In frogs with an average body mass 56 g, the minute volume of the heart is equal to 4.5 ml/min X 100 g, which is approximately an order lower than in mammals with the same body mass. Pulmonary fraction constitutes 52% of the minute volume of the heart. The main bulk of systemic fraction of the minute volume of the heart (78%) passes to locomotor system and skin, whereas 19% of this volume are adressed to vegetative organs. This pattern of distribution significantly differs from that in mammals with a similar body mass, in which the vegetative and locomotor fractions are approximately equal. Differentiation in muscular blood supply was noted--there is a threefold difference in the volume of blood flow between gastrocnemius and submandibular muscles.     

The autonomic innervation of the liver and gallbladder of Rana ridibunda

1--The innervation of the liver and gallbladder of Rana ridibunda has been studied by the following methods: (a) demonstration of cholinesterase activity; (b) FIF method for catecholamines; (c) immunohistochemistry for VIP and (d) electron microscopy. 2--The hepatocytes are arranged in regular rows of hepatic cords, very little connective tissue is distributed in the parenchyma, the innervation being restricted to the big branches of blood vessels. 3--Well defined cholinergic and adrenergic plexuses surround the hepatic arteries, portal veins and biliary ducts. The VIPergic innervation is scarce in the liver but a richly branched plexus spreads in the wall of the gallbladder. 4--Cholinesterase-positive cells are widely distributed accompanying the nerve trunks of the gallbladder. The innervation distribution is prominent in the portion of the gallbladder next to the hepatic hilus. 5--A population of melanin-storing cells besides free melanin granules are present in the liver parenchyma and are prominent in the gallbladder where the melanocytes are disposed in close contact with blood vessels and nerve structures. We have observed that the number of these visceral melanocytes considerably increases in winter, particularly in the liver.     

Purification, catalytic and regulatory properties of Rana ridibunda erythrocyte pyruvate kinase

Pyruvate kinase of Rana ridibunda erythrocytes is one of the regulatory enzymes of glycolysis. PK was purified about 7800-fold. The purified enzyme showed on SDS-electrophoresis three protein bands with an apparent molecular weight of between 60 and 65 kD. The enzyme is subject to activation by FDP and to inhibition by ATP. It showed Km values for PEP and ADP of 0.095 and 0.98 mM respectively. It was activated by K+, Mg2+ and Ca2+ ions whereas it was inhibited by Na+ ions. The role of PK of Rana ridibunda erythrocytes, as a key and rate controlling enzyme of the glycolytic flux is discussed.