René Vaillant

Immunoreactive melanocyte-stimulating hormone (α-MSH) in the brain of the frog (Rana esculenta L.)☆

Abstract MSH and ACTH levels in the hypophysis, diencephalon, telencephalon, and posterior encephalon of Rana esculenta were measured by use of sensitive and specific radioimmunoassays. Significant amounts of α-MSH-like peptide were observed in the diencephalon, telencephalon, and rhombencephalon. This component seems to be of extrahypophysial origin, since it was found in animals 8 days after hypophysectomy. Sephadex G-50 gel chromatography of tissue extracts showed that frog brain MSH behaved similarly to rat MSH and to synthetic α-MSH. Frog brain MSH was degraded by rat plasma and by liver or brain homogenates at the same velocity as rat hypophysial MSH or synthetic α-MSH. Enzymatic systems able to degrade endogenous MSH were also found in the frog telencephalon. These results provide evidence that frog brain extracts contain an α-MSH-like peptide and that a number of tissues, including rat plasma, brain, and liver, as well as frog brain, contain enzymatic mechanisms capable of degrading the α-MSH.

Regulation of the aldosterone secretion in the frog Rana esculenta L

Abstract The concentration of aldosterone in plasma of the frog Rana esculenta L. has been measured by radioimmunoassay without chromatography. In the normal frogs, the concentration of this steroid is 0.82 ± 0.16 μg/100 ml in the plasma of blood removed by heart puncture. Using daily administration of metopirone or aminoglutethimide, the steroid is no more detectable after six injections. Hypophysectomy produces, 15 days after the operation, a small but significant diminution of the hormone concentration. On the contrary, one administration of synthetic 1–24 ACTH, or extract of frog kidney, produces an elevation of the aldosterone level, not only in the normal, but also in the hypophysectomized frogs. These results suggest that, in the amphibians, the aldosterone level depends on the renin-angiotensin system and pituitary ACTH. On the other hand, the injection of corticosterone into normal frogs produces a significant decrease of aldosterone concentration, which suggests feedback mechanisms in the control of aldosterone secretion.

A radioimmunoassay for plasma corticotropin in frogs (Rana esculenta L.)

A radioimmunoassay technique has been developed for measuring frog plasma corticotropin (ACTH) without prior extraction. Using synthetic porcine ACTH as a reference standard, 131I-labeled synthetic human ACTH (sp act > 500 mCi/mg) as tracer and rabbit anti-porcine ACTH serum, the lower measurable value was estimated at about 4 pg ACTH. Only human and porcine ACTH, 1–24ACTH, and frog pituitary ACTH reacted with the rabbit anti-porcine ACTH serum. No cross-reactivity has been found with synthetic 1–16,17–39ACTH, αMSH, and bovine βMSH. Appearance of damaged 131I-h ACTH components after storage in plasma solutions was followed for 7 days. The conditions making it possible to reduce ACTH damage have been ascertained. The average plasma corticotropin level (±CI) was found to be 38.8 ± 7.8 pg/ml without any significant difference between males and females. These results suggest that frog ACTH secretion has much in common with mammalian secretions.

Changes in corticotropin producing cells in the pituitary of Rana esculenta L. following interrenalectomy and metopirone treatment. An immunohistochemical study

The presence and the distribution of corticotropin producing cells in the adenohypophysis of Rana esculenta has been demonstrated by immunohistological means. In untreated and control animals alike, fluorescent cells were localized (a) in the rostroventral part of the pars distalis around portal capillaries; (b) in almost all of the pars intermedia. Daily Metopirone injections brought about a decrease in the number of pars distalis fluorescent cells (Day 3) and eventually disappearance (Day 7). Metopirone had no marked influence on intermediate lobe fluorescence. Twenty-four hours after adrenalectomy, the number of anti-ACTH binding cells decreased about 30 or 40%. These cells totally disappeared 4 days after adrenalectomy. In the intermediate lobe, the number of anti-ACTH binding cells was almost identical to that in the control animals. These results suggest that a polypeptide, immunologically similar to mammalian ACTH, is simultaneously secreted in both pars distalis and pars intermedia cells in the frog pituitary. They also point out the existence of a pars distalis-adrenal feedback mechanism.

Comparative effects of canrenoate-K and prorenoate-K upon aldosterone biosynthesis in perifused frog interrenal glands.

To investigate the possible direct effect of two aldosterone antagonists (Canrenoate-K and Prorenoate-K) upon mineralocorticoid biosynthesis a perifusion system technique has been developed. Frog interrenal tissue was selected for its ability to secrete huge amounts of aldosterone (twice as much as corticosterone in resting conditions). Throughout the experiment, secretion of aldosterone was measured every ten minutes by means of a sensitive and highly specific radioimmunoassay method. Increasing concentrations of both Canrenoate-K and Prorenoate-K (ranging from 10(-4)M to 10(-3)M) caused a dose-related inhibition of aldosterone output. At a dose of 3.16 x 10(-4)M, Prorenoate-K appeared to be somewhat more potent (57.8% inhibition) than Canrenoate-K (47.8% inhibition). Infusion of both Canrenoate-K and Prorenoate-K at a dose of 5 x 10(-4)M during 1 or 2 hours induced a similar sharp decrease in mineralocorticoid secretion. Thus, it appears that Canrenoate-K and Prorenoate-K beside their well known effects at renal tubular receptor sites do also inhibit aldosterone biosynthesis. These results indicate that in vivo administration of aldosterone antagonists may first involve a transient decrease in aldosterone secretion. Furthermore, they suggest that mineralocorticoid biosynthesis might be regulated by a short loop feedback mechanism.

Adsorption of polypeptide hormones to silicate powders: Development of reproducible methods for extracting ACTH and MSH from plasma

Abstract Several rapid and easy methods have been compared to extract human ACTH. α-MSH, and human β-MSH from small plasma samples (0.5 ml). The extraction methods were based on the adsorption of these polypeptide hormones to various silicate powders and subsequent elution by different eluants. The best extraction procedures were achieved under the following conditions: The ACTH was adsorbed to 40 mg of silicic acid and eluted with 2 ml of an acetic acid:acetone:water mixture (1:40:59); α-MSH was adsorbed to 50 mg of talcum powder and eluted with 2 ml of 75% aqueous ethanol: β-MSH was adsorbed to 40 mg of silicic acid and eluted with 2 ml of 0.1 n HCl. These extraction techniques did not affect the immunoactivity of the hormones and will prove useful for radioimmunoassays.

Contrôle de la synthèse du glycogène dans le foie fœtal de rat

Summary We have studied through the late foetal period of the rat, the evolution of two enzymes involved in glycogen metabolism of the liver : UDPG glycogen synthetase (a and b forms) and branching enzyme. The activities were measured during the development of normal foetuses and of foetuses experimentally deprived of corticosteroids. In normal foetus the activity of glycogen synthetase and the branching enzyme increased progressively between days 18 and 21 ; but the percentage of glycogen synthetase a increased promptly between days 18 and 19. This variation coincides with the beginning of glycogen accumulation in the liver. In foetuses submitted to corticosteroid shortage, the activity of each enzyme, and the amount of glycogen in the liver, were reduced at term. Cortisol given to the decapited foetus restores subnormal glycogen storage and normal activity of branching enzyme. The activity of synthetase a was slightly increased but remained very low ; only synthetase b was restored to normal. It seems that there is no relationship between the synthesis of glycogen, in the foetal rat liver, and the activity of synthetase a.