High concentrations of pregnenolone and its sulfate have been found in several areas of rat and human brain and seem to be controlled by local mechanisms. In the present experiments we have demonstrated pregnenolone binding sites in the cytosolic fraction of the rat olfactory bulb. The pregnenolone binding component showed a Kd = 2.34 ± 0.66 × 10−7 M and Nmax = 7.25 ± 1.20 pmol/mg protein. Pregnenolone, pregnenolone sulfate and 17OH-pregnenolone competed equally for the binding sites while other steroids were less competitive. Protease and trypsin inhibited binding by 48 and 60% respectively. Sucrose density gradient analysis showed a minor peak at 4.6s and a major one at 3.6s. After gel filtration chromatography the pregnenolone binding component appeared as 2 peaks corresponding to molecular weights of approximately 150 and 220 kDa. Heating at 60°C increased binding by 150%. These results indicate that the olfactory bulb pregnenolone binding component is complex in nature. Rat plasma also bound pregnenolone. Plasma binding sites could be partially differentiated from those in the olfactory bulb on the basis of susceptibility to lipoprotein lipase, effect of heating and mobility during polyacrylamide gel electrophoresis.

We have previously detected progesterone, dehydroepiandrosterone, androstenedione, testosterone and estrone in rat retina. The present experiments show that the presence of these steroids in the retina may not be due to local biosynthesis. On the other hand, they demonstrate considerable 5 alpha-reductase and 3 beta-hydroxysteroid dehydrogenase activities in the rat retina. The significance of these activities in the retina is discussed.

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.

Sex steroids and 5-en-3β-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 5α-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-3β-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.

We have previously reported that the LH-induced decrease in the concentration of ovarian cyclic GMP (cGMP) in the rabbit was accompanied by a drop in ovarian guanylate cyclase activity. The present experiments were carried out to see if the increase in cGMP concentration that occurs in immature rat ovaries after stimulation with pregnant mare serum gonadotrophin (PMSG) is also accompanied by changes in guanylate cyclase activity. Total ovarian cGMP, along with ovarian weight, was found to be increased at 16 h after PMSG treatment. Ovarian concentrations of cGMP, however, increased only after that period (at 20, 24 and 48 h) and the increase was progressive. Guanylate cyclase activity was found in both the cytosol and 100 000 g particulate fractions of the immature rat ovaries. Forty-three hours after PMSG treatment, activity in the particulate fraction was found to be significantly increased. This increase in guanylate cyclase activity was also found at 20 h but not at 16 h. Thus, the increase in ovarian cGMP concentration in immature rats after PMSG treatment was accompanied by increased guanylate cyclase activity.

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.

Ovarian adenosine 3'-5'-monophosphate (cAMP) concentrations were determined in untreated female rabbits and in those sacrificed at various times (0.5-6.0 h) after administration of luteinizing hormone (LH, 50 micrograms) or human chorionic gonadotrophin (hCG, 100 IU). Both LH and hCG caused a rapid increase in ovarian cAMP concentrations within 0.5 h. After LH stimulation the elevated levels declined rapidly and by 2 h they were almost back to control levels. In the hCG-treated animals the decline in ovarian cAMP from the initial peak concentrations was much more gradual and the levels were significantly higher than control even at 6 h. The different response to the two gonadotrophins is discussed in relation to the physiologic importance earlier attributed to the prolonged elevation of cAMP in rabbit ovaries after stimulation with hCG.

The activities of guanylate cyclase and cyclic GMP (cGMP) phosphodiesterase, enzymes that are responsible for maintaining tissue levels of cGMP, were determined in the ovaries of rabbits killed without treatment or 4 h after administration of LH. Ovarian activities of the two enzymes were determined in the 100 000 g supernatant fraction (cytosol) and the resulting pellet (particulate fraction). Significant phosphodiesterase and cyclase activities were detected in both the cytosol and particulate fractions. Administration of LH had no significant effect on phosphodiesterase activity in either of the tissue fractions. On the other hand, LH caused a significant drop in guanylate cyclase activity in the cytosol and particulate fractions. This drop in the cyclase activity may be the cause of the decreased rabbit ovarian concentrations of cGMP that we have previously observed after LH stimulation.

The effect of prostaglandin PGF2 alpha 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 PGF2 alpha (10 micrograms/ml) in corpora lutea of mid and late luteal phases. PGF2 alpha alone did not show a consistent effect on basal progesterone production. The inhibition of hCG stimulated progesterone production by PGF2 alpha at times corresponding to luteolysis indicates a role for that prostaglandin in the process of luteolysis in the human corpus luteum.

Previously we had observed that indomethacin prevented LH-induced cholesterol depletion in rabbit ovarian follicles in vivo, indicating a role for prostaglandins in that LH action. In present experiments we have found that 1) pretreatment of the animals with either meclofenamic acid or acetylsalycylic acid, inhibitors of prostaglandin biosynthesis, prevented the LH-induced cholesterol depletion in the follicles; 2) administration of PGE2, but not PGF2α, caused cholesterol depletion in the follicles and the effect was not blocked by indomethacin; 3) simultaneous administration of LH and PGE2 resulted in the same magnitude of cholesterol depletion as was observed after their individual administration; and 4) cholesterol depletion caused by PGE2 was in the ester moiety only, and this effect was similar to the earlier reported effect of LH administration. Thus, it seems that PGE formation may be necessary for expression of the cholesterol depleting action of LH in the rabbit follicle.

The possible role of prostaglandins of the PGE and PGF series in the follicular compartment of human ovaries or the capacity of that tissue of form them is not well defined. In the present experiment we have examined 1) the endogenous concentrations of PGE and PGF in follicles of human ovaries at various stages of the menstrual cycle, 2) the capacity of separated theca and granulosa to form PGE and PGF in vitro in the presence of substrate arachidonic acid and 3) the possible modulation of that capacity by previous in vitro exposure to hCG (10 IU) alone or in combination with hMG (5 IU). PGE and PGF were determined by radioimmunoassay. Follicles from all stages of the cycle were found to contain measurable amounts of PGE (0.10 - 3.75 ng/follicle) and PGF (0.13 - 1.11 ng/follicle). These compounds were localized more in the theca than the granulosa cells. On a per follicle basis theca showed more capacity to form PGE and PGF in vitro than the corresponding granulosa cells. However, exposure to gonadotrophins stimulated PGE and PGF formation in the granulosa cells and not in the theca. The presence of PGE and PGF in the follicles indicates a physiological role for these compounds in that tissue. Although thecal tissue showed a greater intrinsic capacity to form PGE and PGF, the contribution of granulosa cells may be more important under acute gonadotrophin stimulation.

Over 15 yr, 24 patients underwent transsphenoidal pituitary surgery for Cushing's disease with a median follow-up of 12 months. Cures included 7 patients with normal sella turcicas (6 microadenomas), 6 patients with focal depressions (6 microadenomas) of the sella (grade I), and 3 patients (3 adenomas) with enlarged sellas (grade II). Three patients with sella destruction (grades III and IV), 2 with normal sellas, and 1 with focal sella depression (grade I) were not cured. Two apparent cures (microadenomas) recurred. Tumor histology revealed 19 basophilic adenomas; electron microscopy (14 tumors) and immunochemical studies (10 tumors) revealed only ACTH cells. Circadian rhythm returned in 6 cured patients. Impotence (in 2), amenorrhea (in 7), and galactorrhea (in 3) resolved in affected cured patients. The major surgical complication was hemorrhage at the operative site (3 patients). Transsphenoidal pituitary surgery is a valuable method for managing Cushing's disease in many patients.

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.

Rabbit ovarian follicles were incubated without stimulation, with LH and with LH + an inhibitor of steroid biosynthesis. Formation of prostaglandins PGE and PGF and of progesterone and estradiol was measured in these incubates. It was found that aminoglutethimide phosphate (AGP) inhibited the LH stimulated biosynthesis of both prostaglandins and steroids. However U 30870 and Metyrapone, while completely inhibiting the LH stimulated biosynthesis of progesterone and estradiol respectively, had no effect on the formation of prostaglandins. Further, the inhibition of prostaglandin formation by AGP could not be reversed by exogenous steroids. It, therefore, appears that the effect of AGP on prostaglandin biosynthesis may not be related to its effect on steroid biosynthesis. However, the response of rabbit follicles to AGP is contrary to that reported for rat follicles and indicates different control mechanisms for prostaglandin formation in the follicles of the two species.

Sexually mature female rabbits were sacrified at 30 min, 1, 4, 5, 8 and 9 h after LH (50 µg) administration. From each animal, 8–10 follicles were isolated from both the ovaries, pooled and the total and free cholesterol contents were determined by the Schoenheimer-Sperry method. The total cholesterol content of the follicles was found to be significantly depleted with 30 min after LH administration as compared to saline-treated controls. The magnitude of this depletion was the same at all the time periods. The drop in cholesterol content was exclusively due to a drop in cholesterol esters and free cholesterol levels remained essentially unchanged. Pretreatment of the animals with indomethacin at a dose known to inhibit follicular prostaglandin biosynthesis prevented the LH-induced cholesterol depletion. These results are discussed in relation to the patterns of changes in steroid and prostaglandin concentrations that are known to occur in the LH-stimulated rabbit follicles.

Laboratoire d'Endocrinologie, Hôpital Notre-Dame et Département de Medicine, Université de Montréal, Montréal, Canada (Received 28 June 1978) Cyclic AMP has been implicated as an intermediate in some of the actions of luteinizing hormone (LH) on ovarian tissues, such as stimulation of steroidogenesis (LeMaire & Marsh, 1975). Both in vitro (Marsh, Butcher, Savard & Sutherland, 1966) and in vivo (Armstrong, Dorrington & Robinson, 1976), stimulation with LH results in a rapid increase in the amount of cyclic AMP in ovarian tissues, which precedes the LH-induced increase in steroidogenesis. Recently, studies on rat ovaries (Grinwich, Ham, Hichens & Behrman, 1976; Ratner, 1976; Ratner & Sanborn, 1976) have indicated that the ovarian tissue content of cyclic GMP may also be regulated by LH, but in a direction opposite to that of cyclic AMP. In the rabbit, Goff & Major (1975) have shown that administration of human chorionic gonadotrophin (HCG) causes a biphasic increase

Owing to the electron-deficient nature of boron atoms, the structures and properties of boron clusters can be enriched by doping various metal atoms, including lanthanide metal atoms. Nevertheless, the viability of actinide analogues has not been fully elucidated up to now. Here we demonstrate a series of highly stable low-valent actinide(III) boron clusters AnB7 (An = Pa, U, Np, and Pu) using first-principles calculations. The predicted global minimum structures of all the AnB7 complexes possess half-sandwich geometries with C6v symmetry and belong to MIII[B7]3--type species. In each AnB7 species, the B73- ligand possesses double aromaticity features with six delocalized π electrons and six delocalized σ electrons. Bonding analysis shows that although there is a substantial contribution of electrostatic interaction in each cluster, covalent interaction is responsible for the stability of AnB7. All the AnB7 species show significantly high formation energies, especially for NpB7, which is in line with the stronger Np-B covalent bonds. In addition, the simulated photoelectron spectroscopy analysis confirms the high electronic stability of neutral AnB7. These results imply that these ultrastable actinide(III) complexes are accessible in the gas phase at room temperature. This work may provide a theoretical basis for the design of highly stable boron-based nanomaterials as well as preparation of low-valent actinide complexes.

Laboratoire d'Endocrinologie, Hópital-Notre-Dame et Département de Médecine, Université de Montréal, Montréal, Canada (Received 8 June 1977) In ovarian tissues, the synthesis of oestrogen from pregnenolone can proceed via progesterone and the 3-oxo-4-unsaturated steroid pathway or by way of dehydroisoandrosterone and the 5-unsaturated steroid pathway. In human ovaries, the follicle seems to prefer the 5-unsaturated pathway for formation of oestrogen (Ryan & Petro, 1966), whereas in the corpus luteum, the 4-unsaturated steroid pathway may be the only one that is operative (Ryan, 1963). The situation is not so clear in other species. In the rabbit, a reflex ovulator, ovarian follicles are known to secrete oestradiol (YoungLai, 1972). Furthermore, isolated follicles can form oestradiol, androgens and progesterone from endogenous (Mills, 1975) or exogenous (Mills & Savard, 1972) precursors in vitro. Mills & Savard (1972) incubated rabbit follicles with [1-14C]acetate and isolated labelled oestradiol and intermediates involved in the 4- and 5-unsaturated steroid