Andrew M. Michelakis

Hormonal effects of smoking--II: Effects on plasma cortisol, growth hormone, and prolactin.

Effect of smoking on the plasma levels of cortisol, growth hormone, and prolactin was evaluated in a group of smokers and nonsmokers. Plasma levels of these hormones were measured under basal conditions and following a short burst of smoking. In addition, to determine the mechanism of action of nicotine on the release of these hormones, rat renal cortical slices were incubated with nicotine and the generation of cyclic AMP was measured in vitro. Increasing concentration of nicotine in the incubation meidum resulted in increased generation of cyclic AMP. Basal levels of plasma cortisol were similar for both smokers and nonsmokers. After smoking, the cortisol levels increased significantly among smokers only and the levels achieved were significantly higher compared with nonsmokers. Mean prolactin curves were higher among nonsmokers compared with smokers, whereas growth hormone levels were similar in the two groups. There data suggest that the effects of smoking on pituitary/adrenal hormones differ among smokers and nonsmokers and that these effects may be mediated through increased generation of cyclic AMP induced by nicotine.

Effect of oral contraceptives on plasma glucose, insulin, and glucagon levels☆

Effects of oral contraceptive agents (mestranol and norethindrone) on carbohydrate metabolism were evaluated in a group of 18 healthy young women. Plasma glucose, insulin, and glucagon responses were evaluated after a glucose load (oral and intravenous) and an amino acid challenge (oral and intravenous). The oral glucose tolerance was normal and was unaltered by the use of oral contraceptive agents. However, following intravenous administration of glucose, plasma glucose levels were slightly but significantly elevated when subjects were using oral contraceptives. Plasma insulin concentrations were slightly but significantly higher than control values in response to oral and intravenous administration of glucose while subjects were using oral contraceptives. Plasma glucagon concentrations in response to oral and intravenous glucose were similar whether the subjects were using oral contraceptive agents or not. No significant differences from control values were observed after oral and intravenous amino acid challenges when subjects were using oral contraceptive agents. Mild elevations of glucose and insulin without any significant change in glucagon concentrations suggest that glucagon levels do not play a major role in the development of insulin resistance seen in some patients using oral contraceptive agents.

Dissociation constant of the norepinephrine-receptor complex in normotensive and hypertensive rats.

Previous reports have suggested that smooth muscle obtained from the thoracic aorta of spontaneously hypertensive rats is less responsive to vasoconstrictive agents than that obtained from normotensive rats. The present study was undertaken to determine whether the responsiveness of aortic muscles from normotensive and spontaneously hypertensive rats correlates with a difference in the affinity of the adrenergic receptors for norepineph-rine and whether antihypertensive therapy alters the affinity of the adrenergic receptors for norepinephrine. The affinity of the adrenergic receptors for norepinephrine was determined by computing the dissociation constant of the norepinephrine-receptor complex (KnR). The values computed for KDR in aortic muscles from normotensive and spontaneously hypertensive rats that had received no antihypertensive therapy were 1.07 × 10−7m and 1.17 × 10−7m, respectively. The values computed for KoR in aortic muscles from normotensive and spontaneously hypertensive rats that had received antihypertensive therapy were 1.38 × 10−7m and 1.29 × 10−7m, respectively. The differences in these values for KnR are not significant. These results indicate that the difference in the contractility of aortic muscles from normotensive and spontaneously hypertensive rats is not related to an alteration in the affinity of the adrenergic receptors for norepinephrine and that the affinity of the adrenergic receptors for norepinephrine is not altered by antihypertensive therapy. Thus, it appears that the etiology of hypertension cannot be directly correlated with a difference in the affinity of the adrenergic receptors for norepinephrine.

Studies on the Characterization of Pure Submaxillary Gland Renin

Summary The effect of the recently obtained pure submaxillary gland renin on blood pressure and its action on rat plasma renin substrate to generate angiotensin were studied. Administration of subnanogram quantities of this renin to nephrectomized rats caused a sustained rise of blood pressure and decrease in sensitivity of the animal to angiotensin. The optimum pH of its reaction with substrate was between 8 and 8.5 and under appropriate conditions the rate of angiotensin generation was proportional to the amount of enzyme, substrate concentration, and time of incubation. The enzyme liberated angiotensin from plasma renin substrate of a number of species.

A comparative study of the effects of cyclocytidine and norepinephrine on renin and esterase release from mouse submandibular gland

SummaryThe effects of cyclocytidine and norepinephrine on the release of renin and esterase from mouse submandibular gland were compared in in vivo and in vitro experiments. Cyclocytidine (150 mg/kg i.p.) produced the depletion of renin and esterase in in vivo experiments as did the α-adrenoceptor agonist norepinephrine (1 mg/kg i.v.). Cyclocytidine was more effective in depleting renin and esterase than norepinephrine. The α-adrenoceptor antagonist phenoxybenzamine (10 mg/kg i.p.), but not the β-adrenoceptor antagonist propranolol (10 mg/kg i.p.), attenuated the depletion of renin and esterase by both cyclocytidine and norepinephrine. These results suggest that the mechanism of action of cyclocytidine on the release of renin and esterase in mouse submandibular gland could be similar to that of norepinephrine, and the secretory process for both cyclocytidine and norepinephrine could be initiated by activation of α-adrenoceptors. In in vitro experiments using dispersed cells, cyclocytidine (10−7 M–10−2 M) had no effect on renin and esterase release, but dose-dependent release of both was clearly observed in response to norepinephrine (10−7 M–10−4 M). Immunocytochemical studies of renin in both in vivo and in vitro preparations showed similar enzymatic activity as measured by radioimmunoassay. The results of in vitro experiments did not correspond to those of in vivo experiments, which may suggest that the site of action of cyclocytidine in mouse submandibular gland may be different than that of norepinephrine. Pretreatment with bretylium (20 mg/kg, i.p.) prior to i.p. injection of cyclocytidine (150 mg/kg) completely blocked the esterase depletion induced by cyclocytidine. From these results, it seems possible to conclude that the in vivo effect of cyclocytidine on mouse submandibular gland is an indirect one and is mediated via reflex activation of sympathetic nerves.

Prostaglandins and renin release from submaxillary glands in vitro.

Abstract The present studies were undertaken to investigate the effect of prostaglandins (PGs) on renin release from the submaxillary glands of mice. Pooled mouse submaxillary gland slices were incubated in Krebs-Henseleit buffer solution following a preincubation period, and renin release was measured by a radioimmunoassay for the direct measurement of submaxillary gland renin. Arachidonic acid (AA) significantly stimulated renin release at 10, 20, and 30 min of incubation. These increases of renin release were abolished by the presence of indomethacin. The synthetic prostaglandin endoperoxide analogue (EPA) strongly stimulated renin release at 10, 20, and 30 min of incubation. However, at a higher concentration the stimulating effect of EPA virtually disappeared. PGI2 caused the highest increase of renin release at 10 and 20 min of incubation. At higher concentrations the effect of PGI2 on renin release was drastically reduced, although it was still statistically significant. PGE2 and PGF2α also exerted a significant increase in renin release; however, the extent of this effect was much less than that of EPA and PGI2. Other prostaglandins such as PGE1, PGA2, PGD2, PGF1α, and 6-keto-PGF1α were found to have no significant effect on renin release. These results suggest that the prostaglandin system directly affects renin release from submaxillary gland independent of systemic hemodynamic and neurogenic influences.

The Renin-Angiotensin System and Renal Prostaglandin E2 Release in Dogs

Abstract The relationship between renin and renal prostaglandin E2 (PGE2) release was investigated in anesthetized dogs using a highly specific radioimmunoassay for PGE2 measurement. There was a dissociation between the acute inhibition of renin release with intra-renal infusion of angiotension II (AII) or angiotensin III (AIII) and renal PGE, release. Intrarenal infusion of norepinephrine resulted in a significant increase both in renin and renal PGE2 release. Intrarenal infusion of bradykinin increased renal PGE2 but not renin release, and infusion of isoproterenol increased renin release but not renal PGE2 release. Renal pressure reduction from 134 to 71 mm Hg increased renin release while the renal PGE2 release was unaltered. In addition, renal vasoconstriction was observed with an infusion of norepinephrine, AII, and AIII, while renal vasodilation was observed with an infusion of isoproterenol and bradykinin, and renal pressure reduction. Neither renal vasoconstriction nor vasodilation, however, correlated with renal PGE2 release. These results suggest that the renin-angiotensin system probably does not play a regulatory role in the control of renal PGE2 release, and that the change of renal vascular tone does not accompany renal PGE2 release.

Different effects of compound 48/80 and histamine on plasma renin activity

The effects of subcutaneous injection of compound 48/80 and histamine on the water intake, plasma renin activity (PRA) and plasma histamine levels were investigated in the rat. The results suggest that compound 48/80 and histamine stimulate water intake by different mechanisms. The compound 48/80-induced water intake seems to be mainly mediated by stimulation of the renin-angiotensin system. On the other hand, the histamine-induced water intake seems to be directly mediated by its action in the brain.

The purification and characterization of multiple forms of mouse submaxillary gland renin

Five forms of renin, A0, A, C, D and E, from mouse submaxillary gland were purified by a two-step procedure including chromatography on the immunoaffinity column and CM-cellulose column. Four renin fractions, A0, A, C and E were purified to homogeneity by the criteria of polyacrylamide gel electrophoresis, analytical isoelectric focusing and Ouchterlony double immunodiffusion. All these forms of renin have molecular weights of 40 000 as determined by gel filtration on Sephadex G-100 column. No high molecular weight renin could be demonstrated. Individual renin fractions showed similar angiotensin I formation activity, 52-158 ng angiotensin I/ng protein per h. No other protease activity could be detected with hemoglobin or casein as substrate. These purified proteins showed a discrete pattern of migration under polyacrylamide gel electrophoresis. Under denaturing condition in SDS-gel electrophoresis, all but fraction D showed a protein band with a molecular weight of 30 000. Fraction D showed a major component with molecular weight of 33 000. The isoelectric points of these renin forms varied from 5.46 to 5.76. They all reacted with antibody raised against renin A and showed similar pressor response activity with 20 ng quantities of the purified proteins. The closely related characteristics of these five forms of renin were further demonstrated by their similarity in peptide mapping patterns after limited digestion with Staphylococcus aureus V8 protease. The data suggest that these proteins are homologous proteins.

Distribution and Disappearance Rate of Submaxillary Renin

Summary Highly purified submaxillary renin (SR) labeled with 125I was injected intravascularly into adult male mice following removal of submaxillary glands and kidneys, and the disappearance of this labeled SR from the circulating vascular volume was studied on the basis of a two compartment system. There was a fast and a slow component to the disappearance curves. Mean half-times of the fast and slow component were 12.4 ± 0.4 min and 86 ± 3 min in sialoadenectomized mice, while in mice whose submaxillary glands and kidneys were removed the half-times were 14.7 ± 0.4 min and 108 ± 7 min, respectively. The uptake of radioactivity by various organs of the mouse was also measured. Accumulation of radioactivity occurred in the kidneys and liver. Only trace amounts of radioactivity were found in the other organs. The findings suggest that the fast component of the disappearance curve was probably due to equilibration of the injected labeled SR in the circulation. However, the fast component may be related to some extent to the rapid uptake of labeled SR by the kidneys. The half-time of the slow component may represent the true halflife of SR in mice, since a significant reciprocal relationship between the half-times of the slow component and metabolic rate constant k 10 was observed both in sialoadenectomized mice and in nephrectomized-sialoadenectomized mice.