André Lanthier

Sex steroids and 5-en-3β-hydroxysteroids in specific regions of the human brain and cranial nerves

Sex steroids and 5-en-3 beta-hydroxysteroids were determined by radioimmunoassay in specific regions of the human brain, in the anterior and posterior pituitary, in one sensory organ, the retina and in the cranial nerves. Progesterone, androstenedione, testosterone and estrone were found in all areas of the brain and in all the cranial nerves but not in all cases. There was no sex difference except in the case of androstenedione where values were higher in women in some brain areas. Estrone values were always higher than those of estradiol in both men and women. No 5 alpha-dihydrotestosterone was detected in any of the samples studied. The values for pregnenolone, dehydroepiandrosterone and their sulfates were much higher than those of the sex steroids in all areas of the brain and in all the cranial nerves. Values for pregnenolone were greater than those of its sulfate while those of dehydroepiandrosterone were in general equal to or higher than those of its sulfate. The values for pregnenolone were greater than those of dehydroepiandrosterone. There were no obvious regional differences in the concentrations of the 5-en-3 beta-hydroxysteroids either in specific areas of the brain or in the cranial nerves. But there was a definite trend for the free dehydroepiandrosterone values to be higher in women. The possible significance of these observations is discussed.

Adrenal cortical function in reptiles. The in vitro biosynthesis of adrenal cortical steroids by adrenal slices of two common North American turtles, the slider turtle (Pseudemys scripta elegans) and the painted turtle (Chrysemys picta picta)

Abstract Incubation of surviving adrenal tissue slices from the slider turtle (Pseudemys scripta elegans) with progesterone-4- 14 C yielded 14 C labelled corticosterone, aldosterone (11β,21-dihydroxy-3,20-dioxo-pregn-4-ene-18-al-18-11-hemiacetal), 18-hydroxycorticosterone and 11-desoxycorticosterone, while adrenal slices from the painted turtle (Chrysemys picta picta) incubated simultaneously with pregnenolone-7- 3 H (3β-hydroxy-pregn-5-ene-20-one-7- 3 H) and progesterone-4- 14 C gave rise to the four above mentioned corticosteroids, all of them containing both 14 C and 3 H. It was concluded that both progesterone and pregnenolone may serve as precursors for the 17-desoxycorticosteroids isolated. Comparison of the 3 H/ 14 C ratio of the transformation products seemed to indicate a direct biosynthetic pathway with the transformation of pregnenolone to progesterone as the initial reaction. However, calculation of the relative specific activities of the transformation products (specific activity of product/specific activity of substrate) revealed an important dissociation between the endogenous steroid production and the transformation of the exogenous substrates.

Studies on the biosynthesis of 18-oxygenated steroids from exogenous corticosterone by domestic duck (Anas platyrhynchos) adrenal gland mitochondria

Abstract The transformation of exogenous, isotopically labelled corticosterone to 18-hydroxycorti-costerone and aldosterone by domestic duck ( Anas platyrhynchos ) adrenal gland mitochondria was studied. The mitochondrial 18-oxygenating system was NADPH dependent. K + , Na + , Ca 2+ and Mg 2+ were necessary for maximal enzymatic activity. The enzyme present in 1 mg of mitochondrial protein became saturated with 13.2 βM of substrate (13.8 μg). Under saturation conditions the maximal production of 18-hydroxycorticosterone was found as 1.43 nmole/min/ mg protein and that of aldosterone 0.33 nmole/min/mg protein. The Michaelis constant for both 18-hydroxycorticosterone and aldosterone was 6.6 × 10 −6 M. Q 10 (average between 20°C and 40°C) was 2.16 for the reaction corticosterone → 18-hydroxycorticosterone and 1.34 for the reaction corticosterone → aldosterone. The mitochondrial enzyme system did not 18-oxygenate exogenous 11-deoxycorticosterone, 11-dehydrocorticosterone, 20β-dihydro-corticosterone. Exogenous 18-hydroxycorticosterone and aldosterone were not metabolized. The kinetics of the transformation of corticosterone to 18-oxygenated metabolites suggested a parallel pseudo-first order reaction rather than a series pseudo-first order reaction. 18-Oxygenation of corticosterone was strongly inhibited by d,1-18-hydroxycorticosterone, p-chloromercuribenzoate, carbon monoxide, metopirone (competitive inhibition; K i = 3.0 × 10 −6 M for 18-hydroxycorticosterone and 9.0 × 10 −6 M for aldosterone) and by aminopterin (noncompetitive inhibition; K i = for both metabolites 2.0 × 10 −5 M). Protein synthesis inhibitors did not have any effect. Cytochrome-P450 was shown to be present in mitochondria by spectrophotometric measurements. Addition of corticosterone. 11-deoxycorticosterone and metopirone produced type II difference spectra. The mitochondrial P450 became saturated with either corticosterone or metopirone at a concentration of 21.8 nmoles/mg protein. From these studies it was concluded that the duck adrenal mitochondrial 18-oxygenating system differed from the mammalian adrenal system previously described, especially in regard of substrate specificity. 18-Hydroxycorticosterone, either endogenous or exogenous, could not serve as substrate for aldosterone production with duck adrenal mitochondria. However, under ordinary circumstances. 18-hydroxycorticosterone synthesis was always associated with aldosterone synthesis in rather fixed proportions. Circumstantial evidence suggested the role of cytochrome-P450 in 18-oxygenation. As to the mechanism of the corticosterone → 18-hydroxycorticosterone → aldosterone reaction, this sequence could not be proven. The possibility should not be discarded that atmospheric oxygen is introduced into a hitherto unknown intermediary substance, which by enzymatic action and/or chemical rearrangement gives rise simultaneously to 18-hydroxy-corticosterone and aldosterone.

THE BIOSYNTHESIS IN VITRO OF RADIOACTIVE CORTICOSTEROIDS FROM (4-14C)PROGESTERONE BY ADRENAL SLICES OF THE DOMESTIC DUCK (ANAS PLATYRHYNCHOS).

Abstract Adrenal tissue slices originating from two female domestic ducks were incubated in a Krebs-Ringer medium using [4- 14 C]progresterone as precursor. After extracting the medium with organic solvents, the crude extract was subjected to extensive fractionation in paper partition systems. The transformation products isolated were identified by the carrier technique. The following is a list of the major [ 14 C]corticosteroids isolated and identified: aldosterone, 18-hydroxycorticosterone-20 → 18 cyclic hemiketal, corticosterone, cortisol and 11-deoxycorticosterone. 18-Hydroxy-11-deoxycorticosterone, though looked for, was not found in the mixture. The transformation rates (expressed as per cent of the precursor added) were the most important for 18-hydroxycorticosterone (15.33%); corticosterone (11.43%) and aldosterone (6.85%).

Luteal phase variations in endogenous concentrations of prosta-glandins PGE and PGF and in the capacity for their in vitro formation in the human corpus luteum

One evidence for a luteolytic role for prostaglandin F2 alpha in the human is the increase in luteal PGF at times corresponding to luteolysis as reported earlier by us and other groups. There have been other contradictory reports on this point. In the present experiments we have measured the concentrations of PGE and PGF in 16 more human corpora lutea and have determined the capacity of those tissues to form PGE and PGF in vitro. PGF concentrations were highest in the mid luteal phase but were accompanied by high PGE concentrations. On the other hand, in the late luteal phase PGF concentrations, lower than in mid luteal but generally higher than in early luteal phase, were significantly higher than PGE concentrations. This pattern in PGE and PGF concentrations was also evident in the capacity of these tissues to form these compounds in vitro. In view of the known capacity of PGE2 to counteract the luteolytic effect of PGF2 alpha, these variations in the relative concentrations of PGE and PGF during the luteal phase may be of significance in the process of luteolysis in the human.

Metabolism of androgens by isolated human beard hair follicles

Abstract Human beard hair follicles were incubated in vitro with testosterone-7- 3 H, androstenedione-1,2- 3 H and dehydroepiandrosterone-4- 14 C.The principal metabolites of testosterone were androstenedione, 5α-androstanedione, 5α-dihydrotestosterone and androsterone. The conversion of androstenedione into testosterone and 5α-dihydrotestosterone was very limited. Dehydroepiandrosterone was metabolized to 5-androstene-3β,17β-diol, 7α-hydroxy-dehydroepiandrosterone and androstenedione, with rates of formation far exceeding those reported for human skin. The significance of these metabolic transformations is discussed with special reference to the mechanism of action of androgens.

Concentration of prostaglandins PGE and PGF, estrone, estradiol, and progesterone in human corpora lutea

The concentrations of prostaglandins PGE and PGF, estrone, estradiol and progesterone in human corpora lutea were measured by radioimmunoassay at various stages of the luteal phase of the menstrual cycle. The concentrations of PGF were found to be significantly higher in both the mid and late luteal phases than in the early luteal phase. In the mid luteal phase there was a concomittant increase in PGE levels, but these levels had declined in the late luteal phase. Steroid concentrations were generally lower in the late luteal phase. It has been postulated that in the human corpus luteum locally produced prostaglandins may be responsible for luteolysis. Our data on the concentrations of PGF and PGE in corpora lutea at various stages of the luteal phase support such a possibility.

Effect of prostaglandin F2α on the hCG-stimulated progesterone production by human corpora lutea

Abstract The effect of prostaglandin PGF2α on the hCG stimulated and basal progesterone production by human corpora lutea was examined in vitro . hCG (40 i.u./ml) stimulated progesterone formation in corpora lutea of early (days 16–19 of a normal 28 day cycle), mid (days 20–22) and late (days 23–27) luteal phases. This stimulation was inhibited by PGF2α (10 μg/ml) in corpora lutea of mid and late luteal phases. PGF2α alone did not show a consistent effect on basal progesterone production. The inhibition of hCG stimulated progesterone production by PGF2α at times corresponding to luteolysis indicates a role for that prostaglandin in the process of luteolysis in the human corpus luteum.

Adrenal cortical function in birds. The in vitro transformation of sodium acetate-1-14C and cholesterol-4-14C by adrenal gland preparations of the domestic duck (Anas platyrhynchos) and the goose (Anser anser)☆

Abstract The in vitro incorporation of sodium acetate-l- 14 C and cholesterol-4 14 C into corticosteroids by duck adrenal preparations and the incorporation of sodium acetate-l- 14 C into corticosteroids by goose adrenal slices was investigated. Labelled sodium acetate gave rise to 14 C-labelled cholesterol, corticosterone (11β,21-dihydroxy-pregn-4-ene-3,20-dione), aldosterone (11β,21-dinydroxy-3,20-dioxo-pregn-4-ene-18-al-18-11-hemiacetal), and 18-hydroxycorticosterone (11β,18, 21-trihydroxy-3,20-dioxo-pregn-4-ene-20-18-cyclic hemiketal). In both duck and goose adrenals. Duck adrenal slices did not incorporate cholesterol-4- 14 C or cholesteryl-4- 14 C-linoleate or cholesteryl-4- 14 C-oleate. However, a fortified homogenate of duck adrenals transformed cholesterol-4- 14 C partially to corticosterone.

Effect of heterosexual olfactory and visual stimulation on 5-en-3β-hydroxysteroids and progesterone in the male rat brain

Pregnenolone, dehydroepiandrosterone and progesterone were measured in specific areas of the brain and in the retina of young adult male rats exposed to the scent or to the scent and view of young cycling females and compared to the levels obtained in males similarly exposed to other males (control). All the steroid values observed were much higher than those found in plasma, varied considerably from one area to another and responded sometimes in opposite directions to stimulation of young adult males by the scent or by the scent and view of cycling females. In male rats exposed to the scent of young cycling females pregnenolone decreased significantly only in the olfactory bulb in comparison with the controls. There were no significant changes in dehydroepiandrosterone or progesterone in this case. When male rats were exposed to the scent and view of female rats, pregnenolone again decreased significantly in the olfactory bulb while dehydroepiandrosterone increased significantly in the olfactory bulb and the retina and progesterone decreased significantly in the hypothalamus, amygdala and parietal cortex. These results are discussed in relation with a possible new role for certain steroids in neuroregulatory mechanisms related to behaviour.