Renal Na+-K+-ATPase in weanling and adult spontaneously hypertensive rats

The interrelationships among plasma renin activity (PRA, ng AI/ml plasma/hr), aldosterone concentration (ng%), and renal Na+-K+-ATPase activity (mumole PO4/mg protein/hr) were studied in 9 weanling normotensive spontaneously hypertensive rats (SHR), 9 adult hypertensive SHR, and 9 weanling and 9 adult normotensive Wistar-Kyoto rats (WKY). All groups were placed on a normal (0.4% sodium) diet. PRA and plasma aldosterone, measured in samples drawn from the ether-anesthetized rat, were higher in weanling SHR (15.2 +/- 2.0, 37 +/- 4.2) than in WKY. PRA measured in samples collected from a separate group of unanesthetized weanling SHR was also greater than in age-matched WKY. In adult SHR, PRA (6.1 +/- 0.9) and plasma aldosterone (20.0 +/- 2.7) were decreased. During the weanling period Na+-K+-ATPase activity in SHR was not only greater than in age-matched WKY but was also increased compared to adult normotensive and hypertensive rats (137 +/- 9 weanling SHR, 89 +/- 7 weanling WKY, 73 +/- 11 adult SHR, 84 +/- 17 adult WKY). Thus, during the weanling period the renin-angiotensin-aldosterone (R-A-A) system and renal Na+-K+-ATPase activity are activated in SHR. The elevation of Na+-K+-ATPase activity may be due to increased aldosterone levels. It was noted, however, that plasma aldosterone was similar in adult WKY and weanling SHR, while Na+-K+-ATPase activity was higher in SHR. These findings involving R-A-A and renal Na+-K+-ATPase activity prior to the elevation of blood pressure suggest that the kidneys may play a role in the initiation of hypertension in SHR.     

Ontogeny of glutamine transport by rat liver plasma membrane vesicles.

Glutamine metabolism in the liver is essential for gluconeogenesis and ureagenesis. During the suckling period there is high hepatic protein accretion and the portal vein glutamine concentration is twice that in the adult, whereas hepatic vein glutamine concentration is similar between adult and suckling rats. Therefore, we hypothesized that glutamine uptake by the liver could be greater in the suckling period compared to the adult period. The present studies were, therefore, designed to investigate the transport of glutamine by plasma membranes of rat liver during maturation (suckling--2-week old, weanling--3-week old and adult--12-week old). Glutamine uptake by the plasma membranes of the liver represented transport into an osmotically sensitive space in all age groups. Inwardly directed Na+ gradient resulted in an "overshoot" phenomenon compared to K+ gradient. The magnitude of the overshoot was greater in suckling rats plasma membranes compared to adult membranes. Glutamine uptake under Na+ gradient was electrogenic and maximal at pH 7.5, whereas uptake under K+ gradient was electroneutral. Glutamine uptake with various concentrations of glutamine under Na+ gradient was saturable in all age groups with a Vmax of 1.5 +/- 0.1, 0.7 +/- 0.1 and 0.5 +/- 0.06 nmoles/mg protein/10 seconds in suckling, weanling and adult rats, respectively (P < 0.01). Km values were 0.6 +/- 0.1, 0.5 +/- 0.1 and 0.5 +/- 0.1 mM respectively. Vmax for Na(+)-independent glutamine uptake were 0.6 +/- 0.1, 0.55 +/- 0.07 and 0.54 +/- 0.06 nmoles/mg protein with Km values of 0.54 +/- 0.2, 0. +/- 0.1 and 0.5 +/- 0.2 mM, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)     

Central infusion of aldosterone synthase inhibitor prevents sympathetic hyperactivity and hypertension by central Na+ in Wistar rats

In Wistar rats, increasing cerebrospinal fluid (CSF) Na+ concentration ([Na+]) by intracerebroventricular (ICV) infusion of hypertonic saline causes sympathetic hyperactivity and hypertension that can be prevented by blockade of brain mineralocorticoid receptors (MR). To assess the role of aldosterone produced locally in the brain in the activation of MR in the central nervous system (CNS), Wistar rats were infused ICV with artificial CSF (aCSF), Na+ -rich (800 mmol/l) aCSF, aCSF plus the aldosterone synthase inhibitor FAD286 (100 microg x kg(-1) x day(-1)), or Na+ -rich aCSF plus FAD286. After 2 wk of infusion, rats treated with Na+ -rich aCSF exhibited significant increases in aldosterone and corticosterone content in the hypothalamus but not in the hippocampus, as well as increases in resting blood pressure (BP) and sympathoexcitatory responses to air stress, and impairment of arterial baroreflex function. Concomitant ICV infusion of FAD286 prevented the Na+ -induced increase in hypothalamic aldosterone but not corticosterone and prevented most of the increases in resting BP and sympathoexcitatory and pressor responses to air stress and the baroreflex impairment. FAD286 had no effects in rats infused with ICV aCSF. In another set of rats, 24-h BP and heart rate were recorded via telemetry before and during a 14-day ICV infusion of Na+ -rich aCSF with or without FAD286. Na+ -rich aCSF without FAD286 caused sustained increases ( approximately 10 mmHg) in resting mean arterial pressure that were absent in the rats treated with FAD286. These data suggest that in Wistar rats, an increase in CSF [Na+] may increase the biosynthesis of corticosterone and aldosterone in the hypothalamus, and mainly aldosterone activates MR in the CNS leading to sympathetic hyperactivity and hypertension.     

Effect of cellular differentiation on 11beta-hydroxysteroid dehydrogenase activity in the intestine.

11beta-Hydroxysteroid dehydrogenase (11betaHSD) converts endogenous glucocorticoids to their biologically inactive 11-dehydro derivatives and is therefore able to determine, at least in part, the biological action of glucocorticoids. Type 1 11betaHSD has both oxidase and reductase activities interconverting corticosterone and 11-dehydrocorticosterone, whereas type 2 11betaHSD has only oxidase activity converting corticosterone to 11-dehydrocorticosterone. Since 11betaHSD expression is regulated during development and by hormones in a tissue-specific manner and since glucocorticoids play an important role in postnatal intestinal maturation, we investigated the role of corticosteroids and cytodifferentiation in the regulation of intestinal 11betaHSD. Using rat intestinal organ cultures and epithelial cell lines derived from rat small intestine (IEC-6, IEC-18) and from human colon adenocarcinoma (Caco-2, HT-29), we analyzed the effect of corticosteroids and cytodifferentiation on 11betaHSD. Screening of the clonal cell lines showed that Caco-2 cells expressed by far the greatest 11betaHSD2 oxidase activity, lower activity was observed in HT-29 cells, and lowest activity was seen in IEC cells. Treatment with dexamethasone (50 nM) increased the activity of 11betaHSD2 in IEC-6 cells (+59%) and HT-29 cells (+31%), whereas aldosterone (50 nM) stimulated 11betaHSD2 in IEC-6 cells only (+31%). Caco-2 cells and IEC-18 cells did not respond to corticosteroids. Growth of IEC-6 cells on Matrigel, treatment of HT-29 cells with butyrate, and postconfluency of Caco-2 cells increased not only the markers of cytodifferentiation, such as alkaline phosphatase and sucrase, but also the activity of 11betaHSD2 in all of these cell lines (IEC-6, +96%; HT-29, +139%; Caco-2, +95%). Addition of corticosteroids to these more differentiated cell cultures did not enhance 11betaHSD2 activity. In intestinal organ cultures of suckling rat small intestine, dexamethasone and aldosterone stimulated 11betaHSD by more than 300%. We conclude that corticosteroids markedly and differentially regulate intestinal 11betaHSD2 and that cytodifferentiation of intestinal epithelial cells is associated with upregulation of 11betaHSD2 activity that is independent of corticosteroids.     

Quercetin and NPPB-induced diminution of aldosterone action on Na+ absorption and ENaC expression in renal epithelium

In renal epithelial A6 cells, aldosterone applied for 24 h increased the transepithelial Cl- secretion over 30-fold due to activation of the Na+/K+/2Cl- cotransporter and stimulated the transepithelial Na+ absorption, activity of epithelial Na+ channel (ENaC), and alpha-ENaC mRNA expression. The stimulatory action of aldosterone on the transepithelial Na+ absorption, ENaC activity, and alpha-ENaC mRNA expression was diminished by 24h-pretreatment with quercetin (an activator of Na+/K+/2Cl- cotransporter participating in Cl- entry into the cytosolic space) or 5-nitro 2-(3-phenylpropylamino)benzoate (NPPB) (a blocker of Cl- channel participating in Cl- release from the cytosolic space), while 24h-pretreatment with bumetanide (a blocker of Na+/K+/2Cl- cotransporter) enhanced the stimulatory action of aldosterone on transepithelial Na+ absorption. On the other hand, under the basal (aldosterone-unstimulated) condition, quercetin, NPPB or bumetanide had no effect on transepithelial Na+ absorption, activity of ENaC or alpha-ENaC mRNA expression. These observations suggest that although aldosterone shows overall its stimulatory action on ENaC (transepithelial Na+ transport), aldosterone has an inhibitory action on ENaC (transepithelial Na+ transport) via activation of the Na+/K+/2Cl- cotransporter, and that modification of activity of Cl- transporter/channel participating in the transepithelial Cl- secretion influences the aldosterone-stimulated ENaC (transepithelial Na+ transport).     

Developmental changes in glutamine transport by rat jejunal basolateral membrane vesicles

The ontogeny of glutamine uptake by jejunal basolateral membrane vesicles (BLMV) was studied in suckling and weanling rats and the results were compared with adult rats. Glutamine uptake was found to represent a transport into an osmotically active space and not mere binding to the membrane surface. Temperature dependency indicated a carrier-mediated process with optimal pH of 7.0. Transport of glutamine was Na+ (out greater than in) gradient dependent with a distinct "overshoot" phenomenon. The magnitude of the overshoot was higher in suckling compared with weanling rats. The uptake kinetics and inhibition profile indicated the existence of two major transport pathways. A Na(+)-dependent system correlated with System A showed tolerance to System N and System ASC substrates, and a Na(+)-independent system similar to the classical L system that favors leucine and BCH. The Vmax for the Na(+)-dependent system was higher in suckling compared with weanling and adult rats. The Vmax for the Na(+)-dependent system was 0.86 +/- 0.17, 0.64 +/- 0.8, and 0.41 +/- 0.9 nmol.mg protein-1.10 sec-1 for suckling, weanling, and adult rats, respectively. The Vmax for the Na(+)-independent system was 0.68 +/- 0.08, 0.50 +/- 0.03, and 0.24 +/- 0.03 nmol.mg protein-1.10 sec-1 for suckling, weanling, and adult rats, respectively. We conclude that glutamine uptake undergoes developmental changes consistent with more activity and/or number of glutamine transporters during periods of active cellular proliferation and differentiation.     

The hormonal regulation of mRNA Na,K-ATPase expression in rat kidneys in postnatal ontogeny]

A study was made of effects of aldosterone, aldosterone+dexamethasone, and aldosterone+spironolactone on Na,K-ATPase mRNA expression in renal cortex of adult and 10 day old rats, when kidney is not sensitive to the hormone injection. It is shown that hormonal induction of synthesis of Na,K-pump mRNA occurs in the early postnatal period apart from mineral corticoid receptors. It seems probable that aldosterone exerts its action in 10 day old rats by interaction with glucocorticoid receptors inducing synthesis of different amounts of alpha- and beta-subunits of Na,K-ATPase.     

Developmental pecularuties of aldosterone regulation of rat epithelial Na channel expression and functional activity

Developmental changes of mRNA alfa-subunit ENaC abundance in renal cortex of 10-day old and in adult rats and aldosterone (10 nM) influence on it were studied. The mRNA level was lower in young rat renal cortex and there was no aldosterone effect on it within 5 hours and 30 minutes, in contrast to adult rats. Intracellular sodium concentration [Na+ i] measured by fluorescent dye Na+ Green in CCD fragments micro-dissected from the kidneys was lower in fragments from immature kidneys, whereas fast nongenomic of aldosterone (10 nM) was the same at both ages. Aldosterone significantly raised [Na+ i] by 40 and 50% in conditions of low sodium concentration (14 mM) in outer medium in epithelial cells of both type of CCD fragments from 10-day old and adult animals, respectively (p < 0.05). We assume that heterochrony of fast and long time genomic effect takes place in aldosterone molecular mechanism maturation. Fast nongenomic effect on the [Na+ i] appeared earlier compared to delayed genomic effect of aldosterone mediated through the changes of the mRNA alpha-subunit ENaC abundance.     

Ontogeny of the Na(+)-H+ exchanger in rat ileal basolateral membrane vesicles.

This study was designed to examine the activity of the Na(+)-H+ exchanger across the basolateral membranes of the ileal enterocyte and its developmental pattern. The function of the Na(+)-H+ exchanger was studied using a well validated basolateral membrane vesicle technique. Na+ uptake represented transport into the vesicle rather than binding as validated by initial rate studies. Na+ uptake represented an electroneutral process as shown by studies in which negative membrane potential was induced by the ionophore valinomycin. Various outwardly directed pH gradients significantly stimulated Na+ uptake compared with no pH gradient conditions at all age groups. However, the magnitude of stimulation was significantly different between the age groups with more marked stimulation of amiloride-sensitive Na+ uptake occurring in adolescent rats as compared to weanling or suckling rats. The amiloride sensitivity of the pH stimulated Na+ uptake was investigated using [Amiloride] = 10(-2)-10(-5) M at pHi/pHo = 5.2/7.5. At 10(-2) M amiloride concentration, Na+ uptake was inhibited by 80%, 70%, 77%, in the basolateral membranes of adolescent, weanling and suckling rats, respectively. Dixon plot analysis in both adolescent and weanling rats was consistent with two amiloride binding sites, a low affinity system and a high affinity system. In the suckling rat, on the other hand, the data supported a single high affinity binding site. Kinetic studies revealed a Km for amiloride-sensitive Na+ uptake of 12.6 +/- 6.6, 10.2 +/- 1.77, 9.46 and Vmax of 4.83 +/- 1.22, 4.47 +/- 0.36 and 8.08 +/- 1.92 n.mol.mg.protein-1.7 s-1 in suckling, weanling and adolescent rats, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)     

Glucocorticoid metabolism and Na+ transport in chicken intestine

The role of aldosterone in regulation of electrogenic Na+ transport is well established, though mineralocorticoid receptors bind glucocorticoids with similar binding affinity as aldosterone and plasma concentration of aldosterone is much lower than glucocorticoids. In mammals, the aldosterone specificity is conferred on the low-selective mineralocorticoid receptors by glucocorticoid inactivating enzyme 11beta-hydroxysteroid dehydrogenase (11HSD) that converts cortisol or corticosterone into metabolites (cortisone, 11-dehydrocorticosterone) with lower affinity for these receptors. The present study examined the chicken intestine, whether changes in 11HSD activity are able to modulate the effect of corticosterone on Na+ transport, and how the metabolism of this hormone is distributed within the intestinal wall. This study shows that not only aldosterone, but also corticosterone (B), was able to increase the electrogenic Na+ transport in chicken caecum in vitro. The effect of corticosterone was higher in the presence of carbenoxolone, an inhibitor of steroid dehydrogenases, and was comparable to the effect of aldosterone. The metabolism of B in the intestine was studied; results showed oxidation of this steroid to 11-dehydrocorticosterone (A) and reduction to 11-dehydro-20beta-dihydrocorticosterone (20diA) as the main metabolic products at low nanomolar concentration of the substrate. In contrast, 20beta-dihydrocorticosterone and 20diA were the major products at micromolar concentration of B. Progesterone was converted to 20beta-dihydroprogesterone. The metabolism of corticosterone was localized predominantly in the intestinal mucosa (enterocytes). In conclusion, the oxidation at position C11 and reduction at position C20 suggest that both 11HSD and 20beta-hydroxysteroid dehydrogenase (20HSD) operate in the chicken intestine and that the mucosa of avian intestine possesses a partly different system of modulation of corticosteroid signals than mammals. This system seems to protect the aldosterone target tissue against excessive concentration of corticosterone and progesterone.     

Corticosteroid induction of renal and intestinal K+-dependentp-nitrophenylphosphatase in young and adult rats

Summary The post-natal development of the K⁺-dependentp-nitrophenylphosphatase (K-NPPase) activity of the Na, K-ATPase complex and its regulation by corticosteroids was studied in renal and intestinal epithelia of the rat using thep-nitrophenylphosphatecerium capture method. The distribution of the phosphatase was analysed in detail in the renal epithelia of the medullary thick ascending limb of Henle's loop and distal convoluted tubule and in the surface epithelial cells of the distal colon. The convoluted tubule and Henle's loop segments showed a stronger reaction for K-NPPase than the colon epithelium both in adult and young animals (suckling and weanling pups). The intensity of staining rose progressively in all three epithelia during early postnatal development and reached the highest levels during the weaning period and in adulthood. The most distinct change was observed between days 10 and 16. Adrenalectomy significantly reduced the density of the final reaction product in weanling and adult rats. Replacement hormone therapy of adrenalectomized weanling rats with the glucocorticoid dexamethansone restored the K-NPPase activity in the two renal epithelia, whereas the mineralocorticoid deoxycorticosterone acetate had no effect on the activity in the medullary thick ascending limb, a very slight effect in distal convoluted tubules, and a strong effect on the distal colon epithelial activity. The observed small effect of the mineralocorticoid in distal convoluted tubule activity may reflect a cross-over into glucocorticoid receptors. We conclude that the postnatal development of Na,K-ATPase is regulated by glucocorticoids in nephron epithelia and predominatly by mineralocorticoids in the surface enterocytes of the distal colon.     

Regulation of epithelial Na+ channels by adrenal steroids: mineralocorticoid and glucocorticoid effects

Epithelial Na+ channels (ENaC) can be regulated by both mineralocorticoid and glucocorticoid hormones. In the mammalian kidney, effects of mineralocorticoids have been extensively studied, but those of glucocorticoids are complicated by metabolism of the hormones and cross-occupancy of mineralocorticoid receptors. Here, we report effects of dexamethasone, a synthetic glucocorticoid, on ENaC in the rat kidney. Infusion of dexamethasone (24 μg/day) for 1 wk increased the abundance of αENaC 2.26 ± 0.04-fold. This was not accompanied by an induction of Na+ currents (I(Na)) measured in isolated split-open collecting ducts. In addition, hormone treatment did not increase the abundance of the cleaved forms of either αENaC or γENaC or the expression of βENaC or γENaC protein at the cell surface. The absence of hypokalemia also indicated the lack of ENaC activation in vivo. Dexamethasone increased the abundance of the Na+ transporters Na+/H+ exchanger 3 (NHE3; 1.36 ± 0.07-fold), Na(+)-K(+)-2Cl(-) cotransporter 2 (NKCC2; 1.49 ± 0.07-fold), and Na-Cl cotransporter (NCC; 1.72 ± 0.08-fold). Surface expression of NHE3 and NCC also increased with dexamethasone treatment. To examine whether glucocorticoids could either augment or inhibit the effects of mineralocorticoids, we infused dexamethasone (60 μg/day) together with aldosterone (12 μg/day). Dexamethasone further increased the abundance of αENaC in the presence of aldosterone, suggesting independent effects of the two hormones on this subunit. However, I(Na) was similar in animals treated with dexamethasone+aldosterone and with aldosterone alone. We conclude that dexamethasone can occupy glucocorticoid receptors in cortical collecting duct and induce the synthesis of αENaC. However, this induction is not sufficient to produce an increase in functional Na+ channels in the apical membrane, implying that the abundance of αENaC is not rate limiting for channel formation in the kidney.     

Steroid-induced protein synthesis in giant-toad (Bufo marinus) urinary bladders. Correlation with natriferic activity.

We have identified a group of proteins (Mr approximately 70 000-80 000; pI approximately 5.5-6.0) in giant-toad (Bufo marinus) urinary bladders whose synthesis appears to be related to aldosterone-stimulated Na+ transport. Spironolactone, a specific mineralocorticoid antagonist in renal epithelia, inhibits the synthesis of these proteins as well as the natriferic effect of the hormone. Since a variety of other steroids (some of which are traditionally considered to be glucocorticoids) also stimulate Na+ transport in toad urinary bladders, we examined whether their natriferic activity was expressed in a fashion similar to that of aldosterone. Short-circuit current was used to measure Na+ transport, and epithelial-cell protein synthesis was detected with high-resolution two-dimensional polyacrylamide-gel electrophoresis and autoradiography. At a concentration of approximately 100 nM, dexamethasone, corticosterone and aldosterone were equinatriferic. Dexamethasone and aldosterone had identical dose-response curves, maximal and half-maximal activity being evident at concentrations of approximately 100 nM and 10 nM respectively. In contrast, at a concentration of approximately 10 nM, corticosterone had no effect on Na+ transport. The natriferic activities of these three steroids correlate with their known affinities for the putative mineralocorticoid receptor in toad urinary bladders. Natriferic concentrations of dexamethasone and corticosterone (140 nM) induced the synthesis of proteins with characteristics identical with those induced by aldosterone. Spironolactone, at an antagonist/agonist ratio of 2000:1, inhibited steroid-induced Na+ transport and the synthesis of these proteins. Thus it appears that all natriferic steroids share a common mechanism of action in toad urinary bladders. Natriferic activity can be correlated not only with relative steroid-receptor affinity but also with the induction of a specific group of epithelial-cell proteins.     

Transferrin degradation by gastrointestinal fluids of suckling and weanling rats

Dietary transferrins are postulated to play a number of biological roles in the developing gastrointestinal tract. A prerequisite for such roles is survival in the gastrointestinal lumen. To evaluate luminal transferrin digestion during development, 125I-transferrin was incubated in vitro with luminal fluid from the stomach and small intestine of 12-day old suckling and 31-day old weanling rats, followed by analysis of degradation products. At both ages, the rate of degradation to trichloroacetic acid soluble material was maximum in the mid-jejunum and lowest in the stomach. Transferrin hydrolysis by weanling fluid was 2-10 times greater than suckling depending upon the particular segment. Chromatography of small intestinal reaction mixtures on Sephacryl S-200 revealed label eluting between intact transferrin and free iodine: two such peaks were generated with suckling fluid and one with weanling. Electrophoresis on SDS-polyacrylamide gels showed two major bands of Mr 69K and 20K; the former was the predominant reaction product with suckling intestinal fluid and the latter with weanling. Both methods showed small amounts of apparently intact transferrin. Results indicate substantial yet incomplete luminal degradation of transferrin which is more pronounced in the weanling than in the suckling. This survival is compatible with potential biological functioning of dietary transferrin or one of its breakdown products within the gastrointestinal tract.     

Modulation of corticosterone availability to white adipose tissue of lean and obese Zucker rats by corticosteroid-binding globulin.

Corticosterone-binding (CB) capacity was determined in periovarian and subcutaneous white adipose tissue (WAT), as well as in plasma of lean and obese Zucker rats. In lean rats, plasma CB was twice the level of obese rats. In lean rat WAT, dexamethasone binding accounted for only 0.05-0.09% of corticosterone binding, and aldosterone bound even less; in the obese rats, dexamethasone accounted for 0.2 - 0.3 % of corticosterone binding. Scatchard plots showed that KD for corticosterone was 3.1 nM (WAT) or 3.4 nM (plasma) in lean rats and 1.8 nM (WAT) or 1.5 nM (plasma) in obese rats. The total CB capacity in WAT was lower in the obese than in lean rats (47-50%). Plasma non-esterified fatty acid levels were higher in obese rats. The results suggest that CBG may limit the access of glucocorticoids to adipocytes more weakly in obese rats because of the lower CBG. Fatty acids may increase the affinity of CBG for corticosterone, which would make WAT cells less accessible to circulating glucocorticoids. The modulation of CBG by fatty acids may protect fat reserves by decreasing the sensitivity of WAT to glucocorticoids.     

Identification of Biochemical Defects in Pancreatic Islets of fa/fa Rats: A Developmental Study

KIBENGE, MOLLY T AND CATHERINE B CHAN. Identification of biochemical defects in pancreatic islets of fa/fa rats: a developmental study. Obes Res. 1995;3:171–178. Adult obese (fa/fa) Zucker rats hypersecrete insulin in response to glucose and other secretagogues. Functional changes in islet ot2-adrenoceptors (8) and glycolytic regulation (9) have been reported. In this study, the development of these biochemical lesions in islets isolated from suckling (3 week old) and weanling (5 week old) lean and fa/fa rats was investigated and compared to results in adult animals. Glucose (15 mM)-induced insulin secretion was inhibited by mannoheptulose (MH) in lean (n=8) but not fa/fa (n=10) adult rats, indicating loss of sensitivity of glucokinase to competitive inhibition. Sensitivity to MH was somewhat reduced in the islets of 3- and 5-week-old fa/fa (n=7 and 12) compared to lean (n=15 and 9) rats, requiring 30–100 fold higher concentrations to achieve significant inhibition. At 3 weeks of age fa/fa rats did not differ from lean controls in either islet insulin content or body weight, but both parameters were increased in fa/fa rats by 5 weeks. The presence of altered α₂-adrenoceptor function in fa/fa rats could not be confirmed in this study. Unlike the previous report, prazosin did not antagonize α₂-agonist mediated inhibition of insulin secretion. The presence of defective regulation of the glycolytic pathway by mannoheptulose in suckling and weanling rats may contribute to development of hyperinsulinemia in fa/fa rats.     

Na+,K+-ATPase activity of the subcellular fractions of the rat brain in aging

The Na+, K+-ATPase activity in the homogenate and in subcellular fractions of different parts of the brain of adult and old rats was studied in comparison. The content of cholesterol in the above fractions was also determined. In old age the Na+, K+-ATPase activity in the homogenate and microsomal fraction of the cerebral hemispheres' cortex decreases, while the Mg2+-ATPase activity in the cortex microsomal fraction increases. The age-related Na+, K+- and Mg2+-ATPase activity in the myelin of the stem in the synaptic plasma membranes of hemispheres and the brain stem remains unchanged whereas in the myelin fraction of hemispheres it grows. The content of cholesterol in the brain of old rats as compared with adult ones increases in the microsomal fraction and remains unchanged in synaptic membranes. The possible role of age-related modification of lipid component of plasma membranes in the above changes of Na+, K+-ATPase activity is discussed.     

Age-related specifics of aldosterone reception in distal segments of the rat nephrons and of Na(+), K(+)-ATPase expression induction

The reason of unresponsiveness of young 10-day rats kidney to aldosterone was explored. The aldosterone binding in distal segments of renal nephrons and the influence of hormonal induction on the mRNA of the alpha and beta subunits of the Na+, K(+)-ATPase in 10-day and 2-month old rats were investigated. There was no age related difference in the aldosterone specific binding in presence of RU-38486 (10(-7) M; Russel Uclaf) in the cortical collecting tubules: 0.26 +/- 0.04 (n = 9) and 0.22 +/- 0.03 (n = 8) mMol/mm of tubule lengths in 10 day and adult rats, respectively. By Nozern blot analysis and RT-PCR more then three and two fold increase of the mRNA abundance of both subunits was found in young and adult renal cortex compare to the adrenalectomized control after aldosterone induction (5 micrograms/100 g. v. b. w. 4 times i/p injections in 3 hour interval between injections) (p < 0.01). By RT-PCR no expression of the alpha 2, alpha 3 and beta 2 isoforms has been observed in all experimental conditions. The age difference was discovered when aldosterone was injected together with spironolactone (5 micrograms and 12 mg per 100 g. b. w. respectively). It was shown, that spironolactone inhibits the effect of aldosterone in adults whereas the latter was unaffected in young rats. The scheme of the age-related differences in the aldosterone regulation of the sodium pump induction in the target cell of the distal part of the rat nephrons is presented.     

The role of corticosteroids in the homeostasis of the eye

Three problems were studied on the human and rabbit eye: to what extent the mineralocorticoids contribute to the control of Na+ and K+ transport in the lens epithelium, how do the glucocorticoids influence the concentration of glucose in the aqueous humour and what is the effect of the pituitary-adrenal axis on the hemato-ocular barrier. Specific receptor-like proteins binding aldosterone were found in the lens epithelium. Na+ and K+ concentrations in the aqueous were influenced by both aldosterone and spironolactone administration. The aldosterone concentration in human cataracts was found to be higher in cases of cataracts complicated by arterial hypertension. In spite of some indication of anticataractogenous action of mineralocorticoids, aldosterone did not prevent the formation of cortisol-induced cataract in chick embryos. Glucose concentration in the aqueous was increased by glucocorticoid administration as well as by stimulation of their secretion by ACTH. Further, the contribution of the pituitary-adrenal axis to the breakdown of the hemato-ocular barrier was investigated by measuring the changes of the total protein content in the aqueous. ACTH1-24 caused a partial breakdown of the barrier, as well as ACTH4-10 or alpha-MSH. As the latter two peptides lack the stimulative effect on the corticoid secretion and glucocorticoids themselves fail to increase the protein content in the aqueous, the breakdown of the hemato-ocular barrier seems to be essential for ACTH-linked peptide fragments and is not mediated by corticoids.