Sex differences in adrenocortical structure and function. VI. Long-term effect of gonadectomy and testosterone or estradiol replacement on rat adrenal cortex

Gonadectomy or sham operations were performed on rats of Wistar strain weighing 150--170 g. Animals were sacrificed in groups, males 397 days and females 395 days after surgery. 14 days before autopsy some of the gonadectomized rats were injected with a single dose of depo-testosterone or depo-estradiol respectively. Absolute as well as relative adrenal weights were larger in female than in male rats. Long-term orchiectomy did not change these weights while testosterone, if compared with control, lowers the weights of the adrenals. Long-term ovariectomy lowers the weights of the glands and estradiol reversed them to the control level. Adrenal cortex of rats studied show irregular zonation. Within the zona fasciculata two types of cells were observed--with strongly eosinophilic and pale cytoplasm. In some cases groups of hypertrophied cells surrounded by spindle-shaped cells were observed. Neither long-term gonadectomy nor gonadal hormone replacement has a marked effect on the structure of the adrenal cortex. In male and female rats there was no difference in the volume of single zona fasciculata and reticularis cells. Long-term orchiectomy has no effect on this volume while testosterone resulted in a lowering of volume of zona fasiculata cells. Neither long-term ovariectomy nor estradiol has an effect on morphometric parameters studied in the fasciculata and reticularis zones. Distinct sex differences and dependence on sex hormones were found in reactions for alpha-glycerophosphate dehydrogenase and nonspecific esterases activity.     

Sex differences in adrenocortical structure and function

Summary The zonation and cellular composition of the adrenal cortex of intact mature male and female rats of different strains (Chbb Thomm, CFY) and three strains of Wistar rats (W1, W2 and W3) at the age of 84–90 days were compared using morphometry.Both absolute and relative adrenal gland weights were higher in female than male rats. Rats of W3 and Chbb Thomm strains had the largest adrenals. These differences depended upon the higher volume of the zona fasciculata (ZF) and zona reticularis (ZR) in the W3 and Chbb Thomm strains than in the remaining animals. Females had larger adrenocortical zones than corresponding males. The average volume of the ZF cell ranged in males from 1589 m³ (W1 strain) to 2111 m³ (Chbb Thomm) and in females from 2249 m³ (W1 strain) to 2894 m³ (W3 strain). The average nuclear volume of the ZF cells was higher in female rats whereas there were no strain-and sex-differences in the size of zona glomerulosa (ZG) and ZR cells.The total number of parenchymal cells per pair of adrenals varied markedly among the studied strains, with the highest number in W3 and Chbb Thomm females. W3, Chbb Thomm and CFY females had larger number of parenchymal cells than males; the reverse was true for W1 strain whereas no differences were found among W2 male and female rats.     

Sex differences in adrenocortical structure and function

Summary Adrenal glands of adult male hamsters are larger and secrete more cortisol than those of females. Stereology was therefore used to study zonal and cellular aspects of development of the adrenal cortex of male and female hamsters. Adrenal glands were studied at weekly intervals from day 21 to day 77 of postnatal ontogenesis. Within this period, body weight did not differ significantly between the sexes. During development, absolute and relative adrenal weights were higher in males; their zona glomerulosa (ZG), zona fasciculata (ZF) and zona reticularis (ZR) become markedly larger than those in females. No marked changes in the volume of individual ZG cells occurred although ZF cells and ZR cells become larger in male than female animals. The total number of adrenocortical cells increased within the period studied, a greater increase being observed in ZG and ZF in males. No distinct sex difference was observed in the number of ZR cells throughout development. From day 56 of postnatal life the adrenal cortex of male hamster contained more parenchymal cells than the female gland. These results thus indicate that sex differences in hamster adrenal cortex depend upon changes in number and size of parenchymal cells.Supported in part by a grant from the Committee of Zoology, Polish Academy of Sciences     

Sex differences in adrenocortical structure and function

Summary Adrenal glands of adult female rats are heavier than the glands of corresponding male rats. Postpubertal orchiectomy increases the adrenal weight, an effect restored by testosterone replacement. Under the same conditions ovariectomy of 8 weeks duration does not change the adrenal weight while estradiol replacement enhances the relative adrenal weight.Karyometric studies have shown that nuclei in the female zona fasciculata cells are larger (app. 18%) than those of the male. Similar but only slight differences (2%) were observed in the zona reticularis. Orchiectomy results in enlargement of cell nuclei within all zones of the adrenal cortex; testosterone replacement has the opposite effect. Ovariectomy of 8 weeks duration slightly enhances the volume of nuclei of the zona glomerulosa cells, has no effect on the nuclei of the zona fasciculata and reduces the volume of nuclei in the zona reticularis. Estradiol replacement reduces the volume of nuclei of the zona glomerulosa cells but increases nuclear volume in the zona fasciculata and in the zona reticularis.Thus testosterone has an inhibitory effect on the adrenal cortex of the rat while the physiologic effect of estradiol on the morphology of this gland, particularly on the zona fasciculata cells is rather dubious.The author wishes to thank Mrs. B. Westerska and Miss K. Siejak for excellent technical assistance.This research was supported in part by a grant from the Zoological Committee, 2nd Department, Polish Academy of Sciences.This paper is dedicated to the memory of late Kazimierz Mietkiewski, M.D., Ph.D. whose encouragements and suggestions were most stimulating for my work.     

Sex differences in adrenocortical structure and function

Summary The cytological aspects of sexual dimorphism in the rat adrenal cortex and its relationship to the gonads have been investigated. The adrenal glands of mature female rats were heavier than those of males, and morphometry showed that this was almost exclusively due to conspicuous differences in the volume of cells of the zona fasciculata (ZF) and zona reticularis (ZR). Stereology demonstrated that the volume of the mitochondrial and lipid droplet compartments, as well as the surface area per cell of mitochondrial cristae and smooth endoplasmic reticulum tubules, were markedly higher in the ZF and ZR cells of female animals. Orchiectomy increased and ovariectomy decreased the adrenal weight, by eliciting hypertrophy and atrophy, respectively, of ZF and ZR cells; these effects of gonadectomy were reversed by the appropriate gonadal hormone replacement. It is suggested that the sexual dimorphism of the rat adrenal cortex may depend upon the inhibitory action of testosterone and the stimulatory effect of estradiol on the hypothalamo-hypophyseal-adrenal axis.     

Sex differences in adrenocortical structure and function. XI

Summary Adrenal glands from orchectomized and ovariectomized rats, with and without replacement therapy, and also from intact controls of both sexes, were examined by autoradiography with ³H-thymidine. The labelling index after 1 or 2 nucleoside injections was higher in the zona glomerulosa of females than in male rats, while no differences were found in the fascicular and reticular zones. Orchiectomy increased the labelling index in the fascicular and reticular zones, an effect prevented by testosterone. Ovariectomy did not change the labelling index, while estradiol lowered it in the zona glomerulosa. Duration of the S phase was longer in the zona fasciculata cells of males than in females. Both orchiectomy and testosterone shortened this phase in cells of the zona fasciculata and zona reticularis. Ovariectomy prolonged the S phase in the zona fasciculata and shortened this time in the reticular zone, an effect reversed by estradiol.In the glomerular and fascicular zones, cell cycle time was longer in males than in females. Orchiectomy shortened this time in all adrenocortical zones, an effect reversed by testosterone. Ovariectomy shortened cell cycle time in the glomerular and reticular zones and prolonged it in the zona fasciculata; these effects were reversed by estradiol. Turnover rate in adrenocortical cells was markedly higher in females than in males, a difference due to testosterone which markedly decreased turnover rate.     

Sex differences in adrenocortical structure and function. III. The effects of postpubertal gonadectomy and gonadal hormone replacement on adrenal cholesterol sidechain cleavage activity and on steroids biosynthesis by rat adrenal homogenates.

In rats postpubertal orchiectomy results in an increase in the adrenal weight, testosterone replacement restores the adrenal weight to the normal level. Neither ovariectomy (8 weeks of duration) nor estradiol replacement has an effect on adrenal weight in female rats. Pregnenolone synthesis as well as corticosterone and blue tetrazolium-positive steroids secretion is significantly higher in homogenates of adrenals from female rats than from males. Orchiectomy results in a marked increase in pregnenolone biosynthesis, testosterone replacement restores the value to the normal levels. Neither ovariectomy nor estradiol replacement has an effect on pregnenolone synthesis in v i t r o. In both sexes gonadectomy causes a marked decrease in corticosterone output by adrenal homogenates, concomitantly the increase in the adrenal 5alpha-reductase activity is observed. The ratio of secreted corticosterone to pregnenolone is significantly lower in gonadectomized rats of both sexes than in control animals. Estradiol or testosterone replacement inhibits the adrenal 5alpha-reductase activity and restores the corticosterone output as well as corticosterone/pregnenolone ratio to the normal values. The above described findings show that the sex differences in steroids secretion by the rat adrenal are partially conditioned by a cholesterol sidechain cleavage activity. Testosterone inhibits this activity while estradiol under applied experimental conditions has no effect on the cholesterol sidechain cleavage activity.     

Heparin-releasable lipase activity of rat adrenals, ovaries and testes.

The presence of NaCl-resistant, neutral triacylglycerol hydrolase (lipase) activity in rat adrenal gland, ovary and testis was studied. Both adrenals and ovaries but not testes were found to contain such a lipase. The activity of the enzyme in the adrenal gland was lowered during cortisol treatment and hypothyroidism. An elevated adrenal lipase activity was found during hyperthyroidism. Pseudo-pregnant and lactating rats had higher ovarian lipase activities than cyclic rats. Ovarian lipase activity in lactating rats was positively correlated with the serum concentrations of progesterone and of 20 alpha-hydroxyprogesterone and negatively correlated with the high-density-lipoprotein non-esterified cholesterol concentration. The lipase activity of adrenals and of ovaries was largely releasable from these organs by heparin and could be inhibited by an antibody against heparin-releasable liver lipase. This indicated that the lipase is extracellularly located and is similar to 'liver' lipase. A possible role of this lipase in adrenals and ovaries is discussed.     

Sex differences in adrenocortical structure and function. V. The effects of postpubertal gonadectomy and gonadal hormone replacement on nuclear-cytoplasmic ratio, morphology and histochemistry of rat adrenal cortex.

Studies were carried out on adult rats of Wistar strain. Six weeks after postpubertal gonadectomy some of the orchiectomized rats were injected with a single dose of testosterone cypionate and ovari-ctomized rats with estradiol cypionate (17 beta-cyclopentyloproprionate esters of testosterone or estradiol). The control rats were sham operated. Investigations were carried out 8 weeks after surgery. Absolute and relative adrenal weight is lower in the male than in the female rat. Orchiectomy increases these weights while testosterone restores them to their normal level. Ovariectomy has no effect on the weight of adrenal, estradiol, however, increases the relative weight of the gland. The adrenal cortex of the adult female is wider than in the adult male rat and in female gland sudanophobic zone is lacking. Orchiectomy leads to the broadening of the adrenal cortex and testosterone replacement has an opposite effect. Ovariectomy has no effect on the structure of adrenal cortex and estradiol replacement resulted in the narrowing of zona glomerulosa. There were no differences in the nuclear-cytoplasmic ratio of zona glomerulosa cells in male and Female rats and neither gonadectomy nor gonadal hormone replacement has no effect on this parameter. The nuclear-cytoplasmic ratio of zona fasciculata cells in the male is markedly higher than in female. Orchiectomy lowers this ratio and testosterone restores it to the normal level. Neither ovariectomy nor estradiol replacement has effect on the nuclear-cytoplasmic ratio of zona fasciculata cells. Similar changes as those in the zona fasciculata were observed in zona reticularis cells. Among the oxidoreductases studied, the most marked sex differences were found in alpha-glycerophosphate dehydrogenase activity. In control female rats an intense reaction for this enzyme is observed in broad band of cells of the zona fasciculata interna, while in male rats only individual cells showed this reaction. In orchiectomized rats the reaction for this enzyme is similar as in control female rats and testosterone restores it to normal. Ovariectomy has no effect on localisation of reaction while after estradiol replacement reaction was more intense. Regarding the remaining oxidoreductases studied, in the adrenal cortex of rats of both sexes no marked differences were observed in localization, however, intensity of reaction depended upon applied experimental conditions. More distinct sex differences were observed in topochemistry of some hydrolases and intensity of reaction depended upon the applied experimental conditions.     

Sex differences in adrenocortical structure and function: 29. Morphometric and functional studies on the effects of gonadectomy and gonadal-hormone replacement on the hamster adrenal cortex.

The morphologic changes evoked in the adrenal glands of male and female hamsters by gonadectomy (15 weeks) and gonadal-hormone replacement (single s.c. injection of 5 mg/100 g b.w. testosterone or 0.5 mg/100 g b.w. estradiol, 2 weeks before the sacrifice) were investigated by morphometric techniques and correlated with the functional alterations of the hypothalamopituitary-adrenal axis. Our findings evidenced a stimulatory role of androgens and a moderate inhibitory one of estrogens on the growth and steroidogenic capacity of the adrenal glands in male and female hamsters, respectively. Moreover, they suggest that these adrenal effects of gonadal hormones are, at least partly, independent of alterations in the pituitary ACTH release.     

Sex differences in adrenocortical structure and function. XXVIII. ACTH and corticosterone in intact, gonadectomised and gonadal hormone replaced rats

Plasma ACTH and corticosterone (B) concentration, ACTH content in the anterior pituitary gland and B content in the adrenals were measured in intact, gonadectomised and testosterone or estradiol replaced rats. Plasma ACTH and B levels and adrenal B content were higher in female than male rats. Neither orchiectomy nor testosterone replacement had an effect on plasma ACTH and B concentration. Orchiectomy did not affect adrenal B content and decreased pituitary ACTH while testosterone significantly lowered ACTH and B content in studied glands. On the other hand ovariectomy did not change pituitary ACTH and adrenal B content and notably lowered concentrations of these hormones in the blood. Estradiol replacement resulted in an increase in plasma ACTH and B concentrations, an effect accompanied by a marked drop in pituitary ACTH and an increase in adrenal B. These findings indicate the distinct sex differences in basal plasma ACTH and B concentrations with higher values in female rats, an effect dependent on the stimulatory action of estradiol on pituitary-adrenocortical axis.     

Norepinephrine turnover in the rat pineal gland. Acceleration by estradiol and testosterone.

The administration of 0.5 mg of testosterone propionate (TP) to orchiectomized rats or of 2 ug of estradiol to oophorectomized rats resulted in significantly less ³H-norepinephrine remaining in the pineal gland 60 and 120 min after a pulse injection of the radioactive compound. This effect was not observed in animals administered with ³H-norepinephrine 45 or 180 min after a single hormone injection. Half-lives for ³H-norepinephrine disappearance in estradiol- and TP-treated rats were 62 and 60 min respectively whereas those of female and male vehicle-injected controls were 109 and 123 min respectively. Hormone treatment did not affect pinela norepinephrine content in any of the schedules used. Norepinephrine efflux expressed as pg. mg tissue⁻¹. min⁻¹ was 33 in estradiol-treated females, 17 in vehicle-treated females, 28 in TP-treated males and 12 in vehicle-treated males. These data indicate that the norepinephrine turnover in neurons innervating the pineal gland is increased by chronic administration of female and male sex hormones.     

Sex differences in the brain: The relation between structure and function

In the fifty years since the organizational hypothesis was proposed, many sex differences have been found in behavior as well as structure of the brain that depend on the organizational effects of gonadal hormones early in development. Remarkably, in most cases we do not understand how the two are related. This paper makes the case that overstating the magnitude or constancy of sex differences in behavior and too narrowly interpreting the functional consequences of structural differences are significant roadblocks in resolving this issue.     

Sex differences in acute luteinizing hormone responses to gonadectomy remain after progesterone antagonist and dopamine agonist treatment.

In male rats, LH pulse frequency and amplitude increase dramatically by 24 h after gonadectomy; in females they increase only slightly by this time. Mean FSH levels increase significantly in both sexes by 24 h after gonadectomy. The objectives of the present studies were to compare pulsatile LH, FSH, and prolactin (PRL) secretion in intact versus gonadectomized and in male versus female rats, and to determine whether the acute postovariectomy lag in LH rise is due to a lingering effect of the higher PRL and/or progesterone (P) levels seen in intact females. LH pulse amplitude, frequency, and mean levels increased significantly by 24 h after gonadectomy in both sexes, but the increases were greater in the males. FSH mean levels, but not pulse amplitude or frequency, increased similarly in both sexes by 24 h after gonadectomy. PRL did not change with gonadectomy. Treatment with CB-154 (a dopamine agonist), with or without RU486 (a P antagonist), 1 h before gonadectomy significantly suppressed pulsatile PRL secretion 1 day later in both sexes. There was no effect of either treatment on LH secretion. We have demonstrated that there is a sex difference in LH, but not FSH or PRL, pulsatility at 24 h after gonadectomy, and that female rats' higher PRL and P levels do not account for their slow rate of LH rise after ovariectomy.     

Brain and plasma levels of testosterone, dihydrotestosterone and estradiol in the one-day-old rat.

Current evidence indicates that the secretion of testosterone during perinatal life is essential in organizing the male brain which subsequently directs the male pattern of gonadotrophin (GTH) secretion and adult male sexual behavior in the rat. It has been hypothesized that testosterone is converted into estradiol enzymatically in the brain prior to its action. In the absence of testosterone and with the resultant low levels of estradiol, female patterns of gonadotrophin secretion and behavior result. In order to investigate this hypothesis further, the endogenous levels of gonadal steroids in the plasmas and brains of 24–48 hr old male and female rats were determined. Pooled samples were analyzed for testosterone, dihydrotestosterone and estradiol by radioimmunoassay. Testosterone levels in male brain and plasma samples were significantly (10-fold) higher than those in the female brain and plasma samples. Brain levels of estradiol were significantly higher in the male than in the female neonate, while plasma levels were identical. Whether the higher level of estradiol in the male brain is due to enzymatic conversion from testosterone within the brain differences in permeability or some other mechanism cannot be stated at this point. The significantly higher brain levels of both testosterone and estradiol in male neonates do fit the pattern predicted by the present concept of sexual differentiation. Dihydrotestosterone levels in brain and plasma of male rats were about 25% of those of testosterone. However in females the brain levels of dihydrotestosterone were significantly higher than testosterone even though the plasma levels of these hormones were identical. This may reflect a protective mechanism through which permeability of testosterone is lowered in the neonatal female brain during the critical period or simply a functional conversion of testosterone to dihydrotestosterone in the female during this period.