Antonio E. Colás

Binding of 5α-dihydroprogesterone and other progestins to female rat anterior pituitary nuclear extracts

The specific binding of 5 alpha-dihydroprogesterone (5 alpha-DHP), progesterone and R5020 to anterior pituitary nuclear extracts was studied using ovariectomized rats treated with estradiol benzoate and progesterone. The binding equilibrium association constant for 5 alpha-dihydroprogesterone with different preparations of nuclear extract ranged from 4.0 +/- 0.54 microM-1 to 59 +/- 10 microM-1. The association constants for progesterone and R5020 were 0.39 +/- 0.81 nM-1 and 1.5 +/- 0.15 nM-1, respectively. The binding of 5 alpha-DHP was specific in that it could be competed only by R5020, progesterone and 5 alpha-DHP and not by other progesterone metabolites and other hormonal steroids tested. With [3H]-progesterone and [3H]R5020 as ligands the most efficient competitors also were R5020, progesterone and 5 alpha-DHP. Estrogen priming of ovariectomized rats consistently and significantly increased the number of binding sites for all three progestins and subsequent progesterone treatment enabled their detection at higher levels in the nuclei.

The Aromatizing Activity of Placental Mitochondrial and Microsomal Fractions

Examination with the electron microscope showed that mitochondrial and microsomal fractions from py human placenta were not homogeneous, despite their careful preparation by several methods. Repeated washing of the mitochondrial fraction, which substantially reduced its glucose-6-phosphatase activity, caused only a small decrease in the aromatizing activity of this fraction. In incubations with 4-androstene-3,17-dione, testosterone and 3Β-hydroxy-5-androstene-17-one, the ratio of 17Β-estradiol to estrone produced was greater with the microsomal ( > 2.00) than with the mitochondrial fraction (0.59–1.82) at each substrate level.

Circadian rhythm of rat hepatic cytochrome P-450 reductase

Abstract The activity of cytochrome P-450 reductase was measured in liver microsomes prepared from adult male rats which had been surgically adrenalectomized, pinealectomized, pinealectomized-adrenalectomized, or sham adrenalectomized-pinealectomized and from intact controls. Rats of each class were killed at 1, 4, 6, 10 hours after the beginning of the light period and 1, 4, 6, 10 hours after the lights were turned off (dark period). The activity of cytochrome P-450 reductase shows a significant diurnal variation in the control group with minimum and maximum at 1 and 10 hours after dark, respectively. The rhythm was altered in the animals surgically treated and the average reductase activity was decreased.

Binding Interactions of Progesterone and other C21 Steroids with Rat Uterine Cytosol

The binding of progesterone and other C21 steroids by rat uterine cytosol preparations was studied by equilibrium dialysis. Scatchard plots and preliminary computer-assisted analysis indicated at least two types of binding sites for progestrone. Individual Scatchard plots for cortisol and corticosterone binding were linear with Kas 0.4 and 1.4 times 10(9) M-1, respectively. Inhibition analyses and modeling of competition experiments suggest, however, that the situation is more complex. There was no binding of labeled dexamethasone.

Placental and luteal steroidogenesis in the pregnant rhesus monkey

Progesterone, 20alpha-dihydroprogesterone, estrone and estradiol-17beta concentrations were estimated by radioimmunoassay in blood plasma from uterine, uteroovarian and femoral veins of rhesus monkeys (Macaca mulatta) on days 22, 49, 128 and 160 of gestation. Steroids were consistently more concentrated in uterine and uteroovarian that in femoral venous plasma and in many cases levels in the uteroovarian vein were also higher than those in the uterine vein indicating luteal secretion of both progestins and estrogens thoughout gestation. In some animals, however, the corpus luteum appeared quiescent. As reflected in the decline in the uterine venous progesterone/estradiol-17beta concentration ratio, a shift in steroid contribution from the uterus and its contents occurred between days 22 and 49 of gestation with progesterone declining more rapidly than estradiol-17beta. Progesterone/20alpha-dihydroprogesterone was higher in both uterine and uteroovarian than in femoral venous plasma suggesting peripheral metabolism of progesterone to 20alpha-dihydroprogesterone.

Cholesterol side chain cleavage and aromatase activities in the corpus luteum of the pregnant rhesus monkey.

Cholesterol side-chain cleavage (CSCC) and aromatase activities were measured in luteal mitochondria and tissue pieces, respectively, from rhesus monkeys on days 22, 49, 128 and 160 of gestation. CSCC activity did not vary significantly during gestation and thus probably does not respond to chorionic gonadotropin which is elevated on day 22 of pregnancy. It is not known, however, whether CSCC can be stimulated prior to day 22 when the corpus luteum is steroidogenically more active. Both 3H-pregnenolone and 3H-progesterone were synthesized from [1,2-3/]cholesterol. Aromatase activity declined from high levels on days 22 and 49 to a nadir on day 128 of pregnancy. Utilizing either [1beta-3H]androstenedione or [1beta-3H]testosterone as substrate yielded comparable results throughout gestation.

The light cycle as a modulator of aminopyrine demethylation by liver from normal, orchiectomized or adrenalectomized rats

Abstract As part of studies on the interactive effects of light and endocrine factors on hepatic mixed‐function oxidases, the aminopyrine demethylase activity of liver microsomes and the serum concentrations of corticosterone and testosterone were measured in adult male rats (sham‐operated, castrated or adrenalectomized) exposed for 21 days to one of the following types of environmental lighting: (1) Normal light cyle (L from 09.00 to 21.00 h); (2) Reverse light cycle; (3) Constant light (LL) or (4) Constant darkness (DD). One half of the animals in each of the 12 groups was killed at 06.00 h and the other at 18.00 h on the last day of treatment. A 3‐way analysis of variance, multiple comparisons and correlations allowed the following conclusions: (1) The type of lighting had the most significant effect on demethylase activity, its highest values corresponding to periods of darkness (particularly in the rats exposed to a reverse light cycle) and its lowest to LL. (2) Castrated rats as a group had the lowe...

Endocrine control of daily variations in hepatic mixed function oxidase activity in the adult male rat

Abstract The hormonal control of rhythmic fluctuations in hepatic microsomal mixed function oxidase activity was examined by incubating microsomes prepared from intact control, sham‐operated, pinealectomized (PX), adrenalectomized (AX), and pinealectomized‐adrenalectomized (PXAX) adult male rats with aminopyrine (AP) and [4‐C14]3s‐hydroxy‐5‐androsten‐17‐one (DHA). The animals were kept on a 12:12 light‐dark cycle for 15 days prior to killingatl, 4, 6 and 10 hafter light and at 1, 4, 6 and 10 h after dark. The rate of both the N‐demethylation of AP and the 7α‐hydroxy lation of DHA varies diurnally with maximal and minimal activities in the dark and light periods, respectively. The fluctuation in both reactions fits a symmetric sine curve for all endocrine treatment groups except for the 7α‐hydroxylation in PX animals. PXAX was the only treatment resulting in a significant increase in 7α‐hydroxylase activity. The data suggest that the pineal may be involved in mediating the effects of environmental light on...