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Vascular and biochemical responses to pancreatic sympathetic nerve stimulation were investigated in the blood-perfused pancreas of anesthetized dogs. During sympathetic nerve stimulation, pancreatic perfusion pressure and norepinephrine release increased, whereas insulin secretion decreased. The latter effect did not occur after pretreatment with the alpha 2-adrenoceptor antagonist idazoxan. However, after beta-adrenoceptor blockade with propranolol, neither single administration of idazoxan nor the alpha 1-adrenoceptor antagonist prazosin or glibenclamide, a blocker of ATP-modulated K+ channels, affected the decrease in insulin secretion induced by sympathetic nerve stimulation. In contrast, the combination of glibenclamide with idazoxan markedly antagonised the decrease in insulin release evoked by the latter procedure. After depletion of catecholamines with syrosingopine, the stimulation-induced inhibition of insulin secretion remained unchanged even though no increases in pancreas perfusion pressure or norepinephrine release were observed. In this preparation, glibenclamide inhibited the decrease in insulin release by 50%. In animals pretreated with the neuronal blocking agent bretylium, all of the responses to sympathetic nerve stimulation were abolished. These results indicate that the inhibitory effects exerted by the sympathetic nervous system on insulin secretion are mediated not only by the classical neurotransmitter norepinephrine acting on alpha 2-adrenoceptors but also by a nonadrenergic cotransmitter that can maintain transmission under conditions of catecholamine deficiency. The postulated nonadrenergic cotransmitter(s) acts, at least partly, via the opening of ATP-modulated K+ channels blockable by glibenclamide, and its release can be prevented by the neuronal blocking agent bretylium.

International audience; The recently synthesized progesterone (P) derivatives, 18-vinylprogesterone (18VP) and 18-ethynylprogesterone (18EP), are potent inhibitors of aldosterone synthesis by adrenal glands. To evaluate the potential interest of these compounds as antihypertensive drugs, we determined whether they also interact with renal mineralocorticosteroid receptors (MR) in kidney and, if so, whether they mimic or antagonize aldosterone action. For this purpose, we evaluated the potency of 18VP and 18EP 1) to displace [3H]aldosterone binding in cytosolic fractions of kidney from adrenalectomized rats and 2) to interfere with aldosterone-induced stimulation of Na(+)-K(+)-ATPase in the collecting tubule of adrenalectomized rats. The properties of 18VP and 18EP were compared with those of their precursor progesterone and of the antimineralocorticosteroid spironolactone. The binding of [3H]aldosterone was restricted to cytosolic MR by presaturating glucocorticosteroid receptor with RU 38486. All compounds tested displaced [3H]aldosterone binding with the following efficiency: spironolactone greater than aldosterone greater than P greater than 18VP greater than 18EP; apparent Kd varied between 0.66 and 16.4 nM. Spironolactone, P, and 18VP antagonized aldosterone-induced stimulation of Na(+)-K(+)-ATPase in the collecting tubule, whereas 18EP mimicked the mineralocorticosteroid action. The different steroids tested altered Na(+)-K(+)-ATPase stimulation and aldosterone binding with the same order of potency.

The mechanism of aldosterone uptake in the epithelial cells of toad bladder was studied using mathematical modeling. Two complementary approaches were used. The first involved analysis of cytosolic aldosterone binding at steady state according to models defined by the sum of independent noninteractive binding sites. The best model describing the experimental data corresponded to two specific binding sites with mean dissociation constant values of 0.20 and 60 nM for types 1 and 2, respectively. The second approach was based on the analysis of cytoplasmic and nuclear aldosterone uptake kinetics at 25 and 0 degrees C in intact bladder. Two models (A and B) were studied. They both implied the existence of two types of aldosterone binding sites as precursors of the corresponding chromatin bound complexes. In model A, nuclear translocation of the two types of receptors was assumed to obey first-order kinetics. In model B, the translocation process for type 1 sites involved a time lag leading to delayed binding to chromatin. Both models were found to fit the experimental data satisfactorily. The fit obtained for model B appeared to be better at low aldosterone concentrations.

The A6 cell line is derived from the kidney of Xenopus laevis. Aldosterone increases sodium transport across A6 cell epithelia. In the present study, aldosterone binding characteristics were studied in A6 cell cytosol. Both type I (mineralocorticoid) and type II (glucocorticoid) receptors are present in the cytosolic fraction of these cells. Aldosterone and corticosterone had a high affinity for type I sites (Kd = 1.25 and 0.16 nM, respectively) and a lower affinity for type II sites (Kd = 39 and 10 nM, respectively). Testosterone and estradiol did not compete for aldosterone binding. RU 26988, a highly specific glucocorticoid agonist, competed with aldosterone for type II but not for type I sites. Hydrodynamic parameters of both type I and type II corticosterone receptor complexes were identical. Their Stokes radius was approximately 6 nm, as estimated by high-performance size-exclusion chromatography, and their sedimentation coefficient determined by ultracentrifugation on glycerol gradients was approximately 9s. The molecular mass calculated from these parameters was approximately 200 kDa, a value that is very close to the value estimated for nontransformed mineralocorticoid and glucocorticoid receptors of other species. The [3H]aldosterone labeling of intact A6 cells was examined by autohistoradiography. At every concentration tested (2, 20, and 50 nM), all cells were found to be specifically labeled in both cytoplasm and nucleus. At 20 nM, in the presence of an excess of RU 26988, labeling was also detected. At every concentration the labeling data was compatible with a Gaussian distribution, indicating that A6 cells correspond to a homogeneous population with regard to aldosterone binding and that probably both type I and type II sites are present in the same cells.