Glucocorticoid receptor of frog (Rana esculenta) liver

The presence of a glucocorticoid soluble receptor is demonstrated in frog liver cytosol. The kinetic characterization of frog liver cytosolic receptor for glucocorticoids is reported and its steroid specificity assessed. Results indicate a gross similarity between frog liver and mammalian glucocorticoid receptor, being a major difference the reduced binding capacity.     

Fiber-type morphology and function of the triads in frog (Rana esculenta) skeletal muscle)]

1. Owing to differing structural characteristics of the contractile substance, the muscle fibres have been divided into the three types A, B and C in former papers. This distinction seems to be corroborated by our investigations into the different structure regarding the traids. As for the A-fibres, they are structured in terms of the T-system being connected in its entire length with the SR-cisternae and circling the myofibrils at the level of the Z-layer. In the B-fibres, this permanent couping of the two membrane systems is partially interrupted so that the T-tubules are arranged round the myofibrils in such a way that they are isolated or only coupled on one side with the SR-cisternae. Apart from the triads we also find diads. There is a totally different arrangement of the membrane systems in the C-fibres. In this instance the T-tubules are not only arranged transversally but also vertically along the contractile elements. They are surrounded by an "SR-labyrinth" which forms individual minor cisternae which are lateraly coupled with the T-tubules. So the axis of these triads is turned by 90 degrees as compared to the A and B fibres. As a result of this different arrangement, these triads always appear in cross-sections, not however, in longitudinal sections as is the case with the A and B fibres. The tirads have a varying shape in the cross-sections depending on the level of the section and due to the fact that the cisternae are not always coupled congruently with the T-tubules. 2. In our discussion we have tried to related these differing shapes and arrangements of the triads in the fibre types A, B and C to known physiological findings. Therefore we deduced that the excitation transmittance and calcium release can be correlated with the anomal rectification of the triads, which has been localized in the region where the T-tubules and SR-cisternae are coupled. However, we can only reckon with a solution once the morphology and function of the "feet" and the eletronmicroscopically "blank" spaces which fill the gap-junction between the T-tubules and the SR-cisternae have been explained. Whatever function the free surface of the T-tubules has remains open. It is directly adjoining the sarcoplasm and we are tryping to relate it to the delayed rectification which appears on the fibre membrane. 3. Moreover from the arrangement of the SR-cisternae i- the individual fibre types we can deduce th intrafibrillar directions of expansion of the calcium after its release and thus the process of the excitation in the A, B and C fibres. It appears that calcium is being directed homogeneously from the SR-cisternae of the A-fibres to the actinfilaments. here the morphological appearance of the twitch fibre presents itself to us. In principle this pattern of expansion of calcium in the B-fibres remains consistent. Owing to the interruption between the T-system and the SR-cisternae we may assume that the process of contraction is delayed in contrast to the A-fibres...     

Synthesis and intracellular transport of proteins in the exocrine pancreas of the frog (Rana esculenta)

Summary Frog pancreatic tissue was pulse-labelled in vitro with ³H-leucine and protein transport was studied in exocrine cells by electron microscope autoradiography. The proteins appeared to be synthesized in the RER and transported to the secretory granules along a similar route and with the same velocity as previously described under in vitro conditions.Evidence was obtained for the involvement of the vesicular and tubular elements at the periphery of the Golgi system in transferring protein from the RER to the Golgi cisternae.Kinetics of the release of newly synthesized proteins from the RER and their appearance in the condensing vacuoles are discussed and related to results reported from other tissues.The transport velocity in this poikilothermic system was studied in relation to the incubation temperature and compared with results reported from its mammalian counterpart. At temperatures between 20 and 30° C intracellular protein transport occurs faster in the frog than in the Guinea pig pancreas. At higher temperature the transport process was severely disturbed in the frog.     

Tyrosine aminotransferase activity of frog (Rana esculenta) liver

1. The presence of tyrosine aminotransferase is reported both in particulate and soluble fractions of frog liver. 2. The activity of the soluble enzyme of frog liver was investigated with regard to its dose and time dependence, its substrate specificity and concentration dependence, its thermal sensitivity as well as pH and temperature dependence. 3. It appears that the properties of the soluble tyrosine aminotransferase of frog liver are in close agreement with those reported for the mammalian liver enzyme.     

Minimal latency of calcium release in frog twitch muscle fibres

Intracellular release of calcium in frog skeletal muscle fibres was monitored by the use of arsenazo III, in response to voltage clamped depolarizing pulses. A latency of a few milliseconds was evident between the onset of depolarization and the first detectable rise in the arsenazo-calcium signal, and this decreased logarithmically as the depolarization was increased. The minimal latency with strong depolarization (to +20 to +100 mV) was about 2 ms at 5 degrees C. This delay appears to be sufficiently long to be compatible with a chemically mediated coupling mechanism between depolarization and calcium release from the sarcoplasmic reticulum.     

Ultrastructure of venom glands in the frog (Rana esculenta)

Electron microscopic study of skin venom glands in the frog, Rana esculenta, revealed the syncytial structure of the inner (secretory) wall which presents two distinct zones: a basal (juxtamuscular) one, which contains nuclei and major cytoplasmic organelles, and an apical one where large electron-dense granules form and accumulate. Granules are seen to arise inside clusters of tightly packed smooth endoplasmic reticulum (SER) elements, which suggests that the SER system is mainly involved in synthesis of this material. A high glutaraldehyde concentration (5%) also reveals a poorly defined material filling the intergranular cytoplasm. No apical limits to the syncytium could be traced, which suggests massive holocrine secretion. Nerves insinuate between the muscle cells and occur all along the internal face of the muscular layer, sometimes in close contact with the syncytium. The gland duct, the wall of which consists of epidermal cells, is blocked, in contact with the gland, by an epidermal bud linked externally to the muscle layer surrounding the gland. Thus, only strong muscle tension such as to expel all or part of the epidermal bud can trigger granule release. This phenomenon can be induced by the subcutaneous injection of epinephrine, but the high and distressing dose needed to provoke appreciable changes in venom glands renders unlikely any natural intense venom release triggered by epinephrine in the frog.     

High resolution scanning electron-microscopic study on the three-dimensional structure of the sarcoplasmic reticulum in the slow (tonic) muscle fibers of the frog,Rana nigromaculata

Summary The three-dimensional structure of the sarcoplasmic reticulum (SR) in the slow (tonic) fibers of the reclus abdominis muscle of the Japanese meadow frog (Rana nigromaculata nigromaculata Hallowell) was examined by high resolution scanning electron microscopy, after removal of the cytoplasmic matrices by the osmium-DMSO-osmium procedure. The SR forms a repetitive network throughout these fibers. At the level of the Z-line, a slender transverse tubule (T-tubule) runs transversely to the longitudinal axis of the myofibril. Small, spherical or ovoid terminal cisternae couple laterally with the T-tubule at intervals of 0.4–1.0 m, and form a terminal cisterna-T-tubule complex on whose surface tiny indentations are occasionally seen. Each terminal cisterna gives rise to a few sarcotubules that run in various directions, divide frequently and form circular or oval meshes of diverse sizes in front of the A- and I-bands. The sarcotubules usually form small meshes in the middle of the A-band, but occasionally fuse and form a poorly developed H-band (fenestrated) collar.     

Circannual variations in mevalonate utilization in frog (Rana esculenta)

1. The fate of mevalonate, the product of HMGCoA reductase, was studied in male and female frogs (Rana esculenta) in order to explain the circannual variations of enzyme activity. 2. The incorporation of 2-14C MVA into unsaponifiable lipids, cholesterol and dolichol in liver, plasma and eggs was followed. 3. Labeled MVA shows a different utilization depending on season and sex. In spring and summer cholesterol synthesis is related to hepatic reserve storage in both sexes, while the peak of enzyme activity, present only in females in fall, seems committed to cholesterol export into the blood and uptake by the oocytes. 4. The presence of a MVA-derived protein identifiable with vitellogenin and labeled on the lipid moiety, suggests that HMGCoA reductase activity in fall is committed to the lipidation of this protein essential for oocyte maturation.     

Hepatic respiratory compensation and blood volume in the frog (Rana esculenta)

In the frog, Rana esculenta (L.), the liver can change in volume by over 25%, depending on the respiratory conditions of the animal: in well-oxygenated specimens the organ can hoard about half of the total amount of erythrocytes in its sinusoids, and release them into the bloodstream under conditions of hypoxia. This phenomenon can be observed at a temperature of 6°C by comparing the liver volumes and haematic values of chlorobutanol-anaesthetized animals exposed to the air or submerged in still water (a condition which induces hypoxia): the blood volume remains constant, at about 5 ml per 100g of body weight, but red blood cell count and haematocrit value differ by as much as 50%. At 18°C there is an increase in oxygen demand and in anaesthetized animals, which rely totally on cutaneous respiration, the compensatory liver mechanism can no longer be observed, since all the available erythrocytes are already circulating in a blood volume which, depending on respiratory conditions, can vary between about 7 and 8ml/100g. At 30°C, cutaneous respiration alone does not allow the anaesthetized animals to survive long enough to stabilize their haematic parameters.     

Photoreceptor layer composition in the retina of the frog (Rana esculenta)

Retinas of Rana esculenta frogs were studied by light and electron microscopy in order to establish the photoreceptor layer composition. We found 56% red rods, 9% green rods, 19% single cones, and 16% double cones. This work provides the morphological basis for electrophysiological investigations concerning the mass receptor potential of isolated Rana esculenta retinas.     

Mechanical properties of frog muscle fibres at rest and during twitch contraction.

Data reported in the literature suggest that crossbridges in rapid equilibrium between attached and detached states (weakly binding bridges), demonstrated in relaxed skinned fibres at low ionic strength, could be present also in intact fibres under physiological conditions. In addition, it was suggested that the well known leading of stiffness over force during the tension development in stimulated muscle fibres could be due to an increased number of weakly binding bridges induced by the stimulation. The experiments reviewed in this paper were made to investigate these possibilities. Fast ramp length changes were applied to single frog muscle fibres at rest and during the early phases of activation. The corresponding force changes were analysed, searching for the components expected from the presence of weakly binding bridges. The results showed no mechanical indication for the presence of weakly binding bridges in both skinned and intact fibres, either at rest or during activation. It was also found that a portion of the fibre stiffness increase induced by stimulation leads the formation of crossbridges.     

Histochemical study of calcium on T-tubule membranes and in the sarcoplasmic reticulum, in frog twitch muscle fibres at rest and during activity

The trigger calcium hypothesis of signal transmission between T-tubules and terminal cisternae (TC) of the sarcoplasmic reticulum (SR) in twitch muscle fibres implies the presence of calcium along T-tubule membranes at rest and its release upon excitation. To test this hypothesis, calcium was immobilised using a fixing and precipitating solution of glutaraldehyde in phosphate buffer at pH 8.0 and the calcium was substituted for by lead. Simultaneous tension recordings revealed the occurrence of contractions or a burst of twitches upon perfusion with the fixative. Procaine or tetrodotoxin (TTX) was used to inhibit this activity. In fibres without fixative-induced activity, precipitates were observed along T-tubules and in adjoining parts of TC. In activated fibres, tubular and TC precipitates were absent. These results are consistent with the trigger calcium hypothesis. In fibres activated by depolarisation, calcium returned to TC after passing successively through different parts of the SR.     

Histochemical study of calcium on T-tubule membranes and in the sarcoplasmic reticulum,in frog twitch muscle fibres at rest and during activity

Summary The trigger calcium hypothesis of signal transmission between T-tubules and terminal cisternae (TC) of the sarcoplasmic reticulum (SR) in twitch muscle fibres implies the presence of calcium along T-tubule membranes at rest and its release upon excitation. To test this hypothesis, calcium was immobilised using a fixing and precipitating solution of glutaraldehyde in phosphate buffer at pH 8.0 and the calcium was substituted for by lead. Simultancous tension recordings revealed the occurence of contra tions or a burst of twitches upon perfusion with the tixative. Procaine or tetrodotoxin (TTX) was used to inhibit this activity. In fibres without fixative-induced activity, precipitates were observed along T-tubules and in adjoining parts of TC. In activated fibres, tubular and TC precipitates were absent. These results are consistent with the trigger calcium hypothesis. In fibres activated by depolarisation, calcium returned to TC after passing successively through different parts of the SR.