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Circulating maternal prolactin (PRL) levels have been reported to be higher in term pregnancies yielding male infants. The mechanism for this gender difference is unknown, but we theorized that it was mediated through the fetal adrenal cortex. To test this theory we measured circulating PRL and estriol (E3) concentrations with radioimmunoassay in 37 pregnant women at 34 and 36 weeks' gestation. We then separated the groups by newborn gender. Maternal serum PRL levels were significantly higher in the women bearing male fetuses. There was no significant difference by gender in E3 concentrations, and there was no PRL surge corresponding to the E3 surge at 34-36 weeks' gestational age. There was no correlation between E3 and PRL levels. Transmission of the fetal gender effect on maternal PRL does not appear to be mediated through the fetal adrenal as measured by the fetoplacental production of E3. The effect probably is mediated by the fetal gonad.

On the 20th day of fetal life the cell proliferation is higher in the zona glomerulosa. The fate of marked cells in each cell compartment shows centripetal migration. Their displacement, from the 20th day of pregnancy up to five days post-partum, while at that time the adrenal growth is slowered down, suggests a real migration.

Plasma renin activity (PRA) and adrenal angiotensin II receptors have been studied simultaneously in the rabbit at various stages of development. In the fetus both parameters show very low values but increase rapidly during the last 4 days of gestation. PRA reaches a maximal level in the early post-natal period but the concentration of adrenal angiotensin II receptors continues to increase further up to the adult state. In adult females, pregnancy results in an initial rise in PRA and adrenocortical angiotensin II receptors. However, PRA remains at a high level throughout the gestation period whereas the number of adrenocortical angiotensin II receptors decreases progressively as pregnancy progresses. The role of circulating angiotensin in the regulation of the concentration and affinity of its receptor sites is discussed.

The influence of ACTH (100 pg/ml), ethanol (10(-3)/M) and morphine (10(-6)/M) on the rate of formation of cholesterol and various steroids in female fetal adrenal tissue was studied in vitro. ACTH and ethanol had no effect on corticosterone formation, while morphine increased it significantly (0.586 +/- 0.049 to 0.799 +/- 0.027 microM/mg protein) (P less than 0.02). Cholesterol formation was increased significantly by ethanol (from 0.103 +/- 0.079 to 0.248 +/- 0.035 microM/mg protein) (P less than 0.01) but not by morphine. Both ethanol and morphine significantly decreased testosterone synthesis (from 0.079 +/- 0.043 to 0.019 +/- 0.002 and 0.006 +/- 0.003 microM/mg protein, respectively) (P less than .01, less than .001). Adrenocortical formation of 17-beta estradiol was similarly attentuated by both ethanol (from 0.372 +/- 0.056 to 0.948 +/- 0.024 microM/mg protein) and morphine (to 0.600 +/- 0.020 microM/mg protein). Adrenocortical pregnanediol formation was significantly decreased by both ethanol (50%; P less than .01) and morphine (36%; P less than .02). Thus, the effect of ethanol and morphine on testosterone, estradiol and pregnanediol was similar and consisted of suppression. The effects of these agents upon corticosterone and cholesterol formation were different. Ethanol, like ACTH, did not stimulate corticosterone formation, but such stimulation did occur with morphine. Ethanol, but not morphine, stimulated cholesterol formation. The data suggest that agents capable of placental transport affect the formation of adrenocortical hormones in the fetal adrenal.

A study was made of the functional activity of the adrenal cortex in 15 1/2-, 18 1/2-, and 21 1/2-day rat fetuses and neonates. Adrenal tissue was incubated after its preliminary trypsinization at 37 degrees C for over 2 weeks with and without ACTH addition into the culture medium, in the amount of 0.1 U/ml. The corticosterone level was determined in the culture medium fluorimetrically. The activity of 3 beta-oxysteroid dehydrogenase was determined histochemically. Corticosterone was revealed in the culture medium. The activity of 3 beta-oxysteroid dehydrogenase was found in the cells of the cultivated adrenal glands. Addition of ACTH into the culture medium considerably increased the corticosterone level and elevated the activity of 3 beta-oxysteroid dehydrogenase. It can be considered on the basis of the aforesaid that cells of the primary culture of the adrenal cortex of rat fetuses aged 15 1/2-, 18 1/2-, and 21 1/2-days and of neonates could form, in the process of incubation, corticosterone from the endogenous substrates in the course of 11, 16, 8 and 13 cultivation days, respectively. The presence of ACTH in the culture medium for the adrenal glands of rat fetuses (of all the gestation periods under study) and of neonates intensified the steroidogenesis.

The adrenal cortex from 31 fetal lambs ranging from 60 to 146 days of gestation was examined by light and electron microscopy. The adrenals of two newborn lambs and three adult ewes were also examined. The adrenal cortex of the adult gland is divided into three regions. The cortex of the fetal lamb consists of two regions only for no zona reticularis is ever clearly seen. The adrenal cortex grows steadily throughout gestation until a rapid phase of growth in the final few days. Most of this terminal growth takes place in the zona fasciculata; it coincides with the known sharp rise in fetal plasma corticosteroids. The cells of the zona glomerulosa appeared capable of secreting steroids in the earliest fetus examined. The cells changed little throughout gestation but from 116 days to term there was an obvious alteration in the proportion of tubular to lamellar cristae within the mitochondria. Cells within the zona fasciculata appeared to undergo functional differentiation as gestation progressed. At 60 days only a few cells appeared capable of secreting steroids. These were situated at the cortico-medullary border and contained mitochondria with mainly vesicular but some tubulolamellar cristae, smooth endoplasmic reticulum and large Golgi apparatus. The remainder of the cells within the region contained mitochondria with tubulolamellar cristae and smaller amounts of predominantly rough endoplasmic reticulum. As gestation progressed the number of differentiated cells increased slowly and in a radial direction towards the zona glomerulosa. As the cells differentiated the amount of endoplasmic reticulum increased and changed from mainly rough to predominantly smooth profiles. The mitochondrial cristae became mainly vesicular. These changes appear to relate to an increase in steroidogenic capacity of the cells. There is a rapid development of functionally differentiated zona fasciculata cells in the last few days of gestation.

Cortical cells of fetal rat adrenals in tissue culture were treated with 5-bromodeoxyuridine (BrdU) during their proliferative phase and during ACTH stimulation when nuclear DNA synthesis has almost ceased. Pretreatment with 0.5 mug/ml/day of BrdU inhibited the ACTH-induced differentiation of cortical cells as well as the secretion of corticosterone and 18-OH-deoxycorticosterone (18-OHDOC). When nuclear DNA synthesis was suppressed and mitochondrial DNA synthesis was stimulated by ACTH BrdU addition (30 mug/ml/day) permitted normal untrastructural differentiation of cortical cells, except that the development of mitochondrial inner membranes was inhibited. Simultaneously mitochondrial inner membranes was inhibited. Simultaneously mitochondrial 11beta- and 18-hydroxylations were strongly inhibited while cytoplasmic 21-hydroxylation was not affected.

This, the first linear morphometric analysis of the epigenesis of the fetal mammalian adrenal cortex, has shown that in the fetal sheep during the latter two thirds of gestation and in the newborn lamb, there are two periods of rapid growth separated by a period of much reduced growth. The fetal ages studies were 53 days (0.36 gestation), a period when the fetal adrenal cortex is actively steroidogenic; 100 days (0.68 gestation), a period of adrenocortical quiescence; 130 days (0.88 gestation), the period of increasing responsiveness to ACTH and cortisol production; 144 days (0.98 gestation), the period of maximal adrenocortical steroidogenesis; and 2 days postpartum, when cortisol production is normally maintained. The first adrenocortical growth period extends to mid-gestation, then growth slows to 0.85 gestation when the second growth period begins. The changes between the first growth period (0.36 gestation) and the period of quiescence (0.68 gestation) are characterised by the attainment of normal adrenocortical zonation and the separation of the medulla. The rate of adrenocortical cell division slows and the zona fasciculata cells become smaller in size. The volume density of the adrenocortical blood sinusoids decreases significantly. The onset of the second growth phase is associated with the previously reported increased levels of fetal plasma ACTH at 0.85 gestation and is expressed initially as a hypertrophic response. Cellular hypertrophy increases from 0.88 gestation to 0.98 gestation and then declines over the birth period. The rate of adrenocortical cell division increases from 0.88 gestation and maintains a maximal rate from 0.98 gestation to 2 days postpartum. These interactions of cellular hypertrophy and hyperplasia, which result in adrenocortical growth, may be explained as a response to fetal ACTH, which has the ability to stimulate the production of peptide growth and differentiation factors, e.g. IGF-II, and cortisol, which then control adrenocortical development in an autocrine and paracrine fashion.

Fibroblasts of the adult adrenal cortex are considered to be nonsteroidogenic connective-tissue cells. However, it has been reported that in response to regenerative stimuli, adrenocorticotropic hormone (ACTH), and transformation to malignancy, these cells acquire characteristics of parenchymal cells, which includes delta 5, 3 beta-hydroxysteroid-dehydrogenase (delta 5, 3 beta-HSD) activity. To determine whether such delta 5, 3 beta-HSD activity in adult adrenocortical fibroblasts was due to the activation or augmentation of gene expression normally occurring during embryogenesis, a histochemical study of adrenocortical development, with particular attention to the connective-tissue capsule, was undertaken. Cryostat sections of rat embryos, from 14-days postconception (PC) to birth, and of adrenal glands 1-8, 44 and 90 days after birth were tested histochemically for delta 5, 3 beta-HSD. The same or adjacent sections were stained for PAS-positive material and reticulin, and with hematoxylin and eosin. delta 5, 3 beta-HSD activity overlapped with fibroblast-like cells and with extracellular connective-tissue components in the periphery of the glands from day-17 PC onward. delta 5, 3 beta-HSD activity over the capsule diminished shortly after birth and was absent in the adult. Appropriate controls showed that the staining within the capsule was specific and not an artifact. 3 beta-HSD activity in the capsule was more intensive when dehydroepiandrosterone (DHEA) was replaced by etiocholan-3 beta-ol-17-one (ETIO) as the steroid substrate. Furthermore, the spatial distribution of 3 beta-HSD activity in the cortex differed depending on the substrate used, and the distribution patterns changed with developmental age. (ABSTRACT TRUNCATED AT 250 WORDS)

The roles of human low density lipoprotein (LDL)- cholesterol and high density lipoprotein (HDL)- cholesterol on adrenal steroidogenesis were investigated using cultured human adult and fetal adrenocortical cells and the findings were then compared to those obtained with bovine adrenocortical cells. The secretion of cortisol in both human and bovine adrenocortical cells was dose-dependently increased by the administration of LDL- or HDL-cholesterol in the presence of adrenocorticotropin (ACTH). LDL-cholesterol was utilized to a greater extent than HDL-cholesterol in both human and bovine adrenal steroidogenesis in the presence of ACTH. Exogenous lipoprotein-derived cholesterol was less utilized in human adrenal steroidogenesis than in bovine adrenal steroidogenesis, compared to the endogenous cholesterol. An increase in the secretion of cortisol and dehydroepi androsterone sulfate (DHEA-S) continued for the 5-day culture period, in the presence of lipoprotein cholesterol and ACTH in both human adult and fetal adrenocortical cells. The secretion of aldosterone increased on the first day of the culture period, then gradually decreased for the 5-day culture period in human adult adrenocortical cells, but not in human fetal adrenocortical cells in the presence of lipoprotein cholesterol and ACTH. These findings demonstrate that exogenous cholesterol utilized in the biosynthesis of steroids is mainly from LDL-cholesterol in both human adult and fetal adrenals and bovine adrenal and the proportion of cholesterol synthesized de novo is significantly larger in the human adult adrenal than in the bovine adrenal.

The fetal adrenal cortex is the central steroid modulator in the fetal placental complex. Its anatomic structure and physiology clearly identify it as a uniquely fetal organ that undergoes atrophy to a fraction of its intrauterine size after birth and assumes markedly different steroidogenic functions on the assumption of extrauterine life. Its importance as a regulator of maturation and parturition in normal gestation is just being understood. The fetal adrenal cortex no doubt plays major roles in the pathogenesis of some of the most important clinical problems currently faced in contemporary obstetrics. Continued research that provides a more complete understanding of this organ will serve as the major base for innovative diagnostic and therapeutic advancements of the future.

An aberrant adrenal, which is a supplementary to the normal adrenal, was found in the apex of hernial sacs in ten out of 350 operated children. This tissue contains adrenal cortex, which tends to atrophy and disappear during childhood, but uncommonly may persist into adult life. Rarely it may secrete abnormally or undergo neoplastic changes. The embryological development is discussed, and the literature briefly reviewed.

The presence of crystal-shaped bodies was demonstrated in the adrenocortical cells of perinatal rats. On the basis of former histochemical evidence on similar structures, they are supposed to have contained cholesterol. The total cholesterol concentration in the adrenal was not elevated during this period. The presence of a great number of crystal-shaped bodies is supposed to be the sign of a transient change in cholesterol compartmentalisation.

This report describes the development of a culture system for long-term growth and cloning of human fetal adrenocortical cells. Optimal conditions for stimulating clonal growth were determined by testing the efficacy of horse serum (HS), fetal bovine serum (FBS), fibroblast growth factor (FGF), epidermal growth factor (EGF), fibronectin, and a combination of growth factors, UltroSer G, in stimulating growth from low density. Optimal conditions for clonal growth were achieved using fibronectin-coated dishes and DME/F12 medium with 10% FBS, 10% HS, 2% UltroSer G, and 100 ng/ml FGF or 100 pM EGF. Conditions for growth at clonal density were found to be optimal for growth of early passage, nonclonal cultures at higher densities. The improved growth conditions used for cloning were shown to allow continued long-term growth of nonclonal human adrenocortical cells without fibroblast overgrowth. All cells in cultures grown in HS, FBS, and UltroSer G had morphologic characteristics of adrenocortical cells, whereas cells grown in FBS only rapidly became overgrown with fibroblasts. Clonal and nonclonal early passage human adrenocortical cells had similar mitogenic responses to FGF and EGF. Whereas FGF, EGF, and UltroSer G showed similar stimulation of DNA synthesis and clonal growth in human adrenocortical cells and human adrenal gland fibroblasts, the tumor promoter 12-O-tetradecanoylphorbol-13-acetate stimulated growth only in adrenocortical cells and was strongly inhibitory to growth in fibroblasts. In both cell types, forskolin inhibited DNA synthesis. Human adrenocortical cell cultures were functional and synthesized cortisol, dehydroepiandrosterone, and dehydroepiandrosterone sulfate. The improved growth conditions for clonal growth of human adrenocortical cells also provided optimal conditions for long-term growth of cultured rat adrenocortical cells and increased the cloning efficiency of cultured bovine adrenocortical cells.

The insulin-like growth factors are broadly distributed in the human conceptus and are thought to play a role in the growth and differentiation of tissues during development. Using in situ hybridization we have shown that a wide variety of specific cell types within tissues express the gene for insulin-like growth factor II at times of development from 18 days to 14 weeks of gestation. Examination of blastocysts produced by in vitro fertilization showed no expression, thus bracketing the time of first accumulation of IGF-II mRNA to between 5 and 18 days postfertilization. The pattern of IGF-II expression shows specific age-related differences in different tissues. In the kidney, for example, expression is found in the cells of the metanephric blastema which is dramatically reduced as the blastema differentiates. The reverse is also seen, and we have noted an increase in expression of IGF-II in the cytotrophoblast layer of the placenta with gestational age. The sites of expression do not correlate with areas of either high mitotic activity or specific types of differentiation, but the observed pattern of expression in the kidney, adrenal glands and liver suggests an explanation for the abnormally high IGF-II mRNA expression in developmental tumours such as Wilms' tumour.

ACTH was administered subcutaneously to rat fetus directly at the late stage of fetal development and acute reaction on the fetal adrenal cortex was observed histochemically and electron microscopically. By administration of ACTH the adrenal cortex became remarkably hyperemic and there were swollen cells in all layers, particularly in the middle and inner layers (corresponding to the zona fasciculata and reticularis in adult rat). Marked reduction of lipid, enlarged mitochondria with increased vesicular cristae and increased smooth-surfaced endoplasmic reticulum (SER) were characteristic. The alterations of mitochondria preceded the change of SER, and thereafter mitochondria showed rapid degeneration. The outer layer (corresponding to the zona glomerulosa in adult rat) also showed similar changes by ACTH to those of the other two layers. These results indicated that the fetal adrenal cortex of rats was exogenous ACTH-reactive and its reaction which was different from that of adult cortical cells, seemed to be specifically related to the development and differentiation of the cells.

The development of the adrenal cortex in pig fetuses between the 50 and 112 day of pregnancy were investigated with histological and histochemical methods and in electron microscope. Three consecutive generations of the cortical cells were observed. They had features of steroidogenic cells and differed with mature of mitochondria and in amount of SER and in enzyme activity. The earliest differentiated fetal cortical cells demonstrated alkaline phosphatase activity and giant mitochondria. They were translocated from the periphery to the center of the adrenal, where they degenerated and disappeared in the late pregnancy. The second generation--transitional cortical cells appeared about 70 day of development. They have pleomorphic mitochondria and abundant SER and low alkaline phosphatase activity. The last generation--the definitive cortical cells differentiated from 100 day of development and their number increased till the parturition. They had active 3-beta-hydroxysteroid dehydrogenase, spherical mitochondria and characteristic concentric whorls of SER. A probable role of the three generations of cortical cells is was discussed. In the subcapsular region of fetal adrenal undifferentiated and differentiating cells were observed, forming glomerularlike groups. Their role in the formation of cortical blastema and future zona glomerulosa was discussed. In the first part of the studies the formation of the mesenchymal primordium of the adrenal cortex was examined in early fetuses of domestic pig, and then differentiation of mesenchymal cells into steroidogenic cortical cells. The changes encountered then at the ultrastructural level consisted of an increased area of SER and greater number of mitochondria. At the same time, the shape of mitochondria altered and the cristae from lamellar became tubular. In the period around 50 day of development fetal adrenal was already a separate organ, with a capsule, composed of cords of fetal cortical cells divided by the blood vessels and strands of chromaffinoblasts penetrating the adrenal. Fetal cortical cells were characterized by alkaline phosphatase activity. The following study presents farther development of the adrenal cortex till the time of birth.

Aspartate aminotransferase was detected in the adrenals of 6-7-week human embryos, the activity being the highest at the 7-9th and 12-14th weeks. Alanine aminotransferase was found in 15-week foetuses. During all the period investigated (6-28 weeks), the activity of aspartate aminotransferase was found to be higher than that of the second enzyme. The activity of both enzymes is higher in female foetuses than in male ones. However, age changes in the activity of both enzymes in male embryos and foetuses were of the same sign as in female ones.

The ability of human adult adrenal to utilize progesterone (P4) for cortisol (F) synthesis in vitro has been compared with that of fetal adrenal tissue. Explant cultures were studied for 6 days using Ham's F10 medium supplemented with 10% fetal bovine serum (FBS), with or without added P4 as substrate. Short-term (4h) incubations of fresh tissue minces were carried out in Ham's F10, with or without P4, in the absence of FBS. In contrast with the fetal gland, F production by cultured adult tissue was unaffected by addition of P4. During short-term incubations, P4 increased F production 150-fold in the fetal tissue as compared to 2- to 4-fold in the adult. ACTH had no acute effect on the P4 to F conversion in either tissue. These results demonstrate that the fetal adrenal exhibits a greater ability to utilize P4 for F production than the adult adrenal.

Fetal adrenal glands were obtained from legal abortions in the 14--22 gestational weeks. The adrenal cortex was separated from the medulla using a micromanipulator. The cortex was homogenized in a glass-teflon homogenizer and the microsomal fraction was isolated by centrifugation. Tissue slices were also prepared by the method of Deutsch. The microsomal fraction or the slices were incubated with (4-14C) pregnenolone or (4-14C) cholesterol in the presence of an NADPH regenerating system and oxygen. ACTH or HCG was added. After extraction and paperchromatography radioactivity was determined in a scanner. The conversion of pregnenolone to 17 alpha-hydroxypregnenolone and dehydroepiandrosterone was increased by ACTH. In the presence of HCG dehydroepiandrosterone production only was increased,--and this occurred in slices only. In the microsomal fraction HCG was without effect on steroid biogenesis from dehydroepiandrosterone.

Fetal adrenocortical responsiveness to ACTH declines during 90-120 days gestation and fetal pituitary peptides have been implicated in this refractoriness. In these studies the ACTH-induced cortisol responses were measured in 11 ovine fetuses of 114 days gestation. Five animals were hypophysectomized as evidenced by prolonged gestation, pituitary histology, TRH-testing, delayed maturation and decreasing fetal plasma prolactin concentrations (less than 1 ng.ml-1) (P less than 0.005). Resting cortisol concentrations decreased from 22.4 to 8.1 ng.ml-1 in the hypophysectomy group and were not different from the control group (19.6-14.9 ng.ml-1) over the 5 days of study. Responses measured as increments in plasma cortisol concentrations increased equally and successively in both groups. Since pituitary ablation fails to enhance fetal adrenal responsiveness to ACTH we conclude that refractoriness is unlikely to be caused by an inhibitor of pituitary origin.

21-Hydroxylase deficiency results in congenital adrenal hyperplasia and leads to masculinization of the external genitalia of affected females. This complication could be avoided if fetal adrenal gland function were suppressed. A woman with mild 21-hydroxylase deficiency whose previous female child had classic congenital adrenal hyperplasia with masculinization was given dexamethasone beginning at the tenth week of gestation. Maternal estriol and cortisol values indicated rapid and sustained fetal and maternal adrenal gland suppression. At 39 weeks' gestation, the patient was spontaneously delivered of a female neonate with normal external genitalia. Postnatal tests indicated the infant was a single heterozygote for 21-hydroxylase deficiency. This study demonstrates prolonged suppression of the fetal adrenal gland with dexamethasone and suggests it might prevent abnormal masculinization in fetuses with severe congenital adrenal hyperplasia.

Enkephalin immunoreactive material from adrenal glands was characterized both in maternal and fetal sheep at various gestational ages. Whole gland extracts from both maternal and fetal sheep contained three major peaks of Enk immunoreactivity corresponding to apparent molecular weights of 10,000, 2800, and less than 1200 daltons. The majority of maternal adrenal Enk immunoreactivity was found in medullary tissue, although cortex also contained low but detectable amounts. This was also the case in newborn lambs and 139 day fetuses, where adrenal cortex was sufficiently developed to allow extraction and quantitation of opioid material. In fetuses at mid-gestation (70-80 days), adrenal medullary Enk immunoreactivity was approximately 75% of maternal values. Met-Enk and Leu-Enk content in 139 day fetal medulla were 70 and 76% of maternal values respectively, while newborn Met- and Leu-Enk medullary content were similar to maternal values. The molar ratio of Met-Enk to Leu-Enk was approximately 4:1 in both maternal and fetal adrenal medulla, and 2:1 in adrenal cortex, suggesting different synthetic processing of opioid peptides in the two tissues. The early appearance of significant levels of adrenal medullary Enk immunoreactivity and subsequent development paralleling that of catecholamines suggest a predominant role for adrenal enkephalins in regulation of fetal cardiovascular function early in gestation.

A study was made of lipid, cholesterol, and ascorbic acid content in the adrenal cortex of rabbit fetuses developing under conditions of normal pregnancy and under the action of hypoxic and hyperoxic exposures during its third trimester. It appeared that in normal pregnancy the adrenal cortex was activated, this being associated with the adaptive motor reactions of the developing fetus. The action of moderate hypoxic exposures led to a greater activation of moderate hypoxic exposures led to a greater activation of the adrenal cortex, this being expressed in a marked fall of lipid, cholesterol and ascorbic acid content. The weight of the total muscle mass and of the fetuses increased as a whole. The adrenal cortex became disactivated under conditions of the hyperoxic exposures; this is expressed in a marked increase in the content of the mentioned formations. The weight of the total mass and of the fetuses decreases.

Thirteen chronic fetal lamb preparations between 95 and 142 days of gestation (term 145--150 days), and 10 newborn lambs were studied before and after the acute (1--2 min) infusion of furosemide (2 mg/kg). The baseline to peak plasma renin activity (PRA) response to furosemide increased from delta 3.0 +/- 1.3 ng/ml/hr (M and SEM) and 95--106 days of gestation to delta 18.4 +/- 4.0 (P less than 0.01) at 123--142 days and delta 33.6 +/- 6.5 (P less than 0.001) in the newborn. Baseline plasma aldosterone concentrations were similar in the fetus and pregnant ewe; aldosterone levels were higher in the newborn lamb than in the nonpregnant ewe. The newborn plasma aldosterone response to furosemide via the endogenous renin-angiotensin was delta 17.1 +/- 4.2 ng/dl (P less than 0.01); the fetal lamb plasma aldosterone level did not increase. The results indicate that the renin-angiotensin system cannot be stimulated by furosemide under 106 days of gestation; the response after 110 days increases with gestational age. Aldosterone concentrations in the fetal lamb are probably maintained primarily by the pregnant ewe and do not increase in response to endogenous renin stimulation as in the newborn.

The differentiation and development of the adrenal gland in pig embryos from 18 to 48 days of pregnancy were investigated with histological and histochemical methods and in electron microscope. Primordia of the adrenal cortex appear at 21st day as two symmetrical streaks of the condensed mesenchymal cells between the mesonephros, aorta dorsalis and mesenterium. The capillaries and chromaffinoblasts penetrate the primordium. The mesenchymal cells differentiate into adrenal cortical cells. In the differentiating cells both, rough and smooth endoplasmic reticulum as well as the number of mitochondria increase. The shape of the cells changes and their projections transform into microvilli. The fetal cortical cells have features of the steroidogenic cells with abundant SER in whorls and numerous mitochondria with tubular cristae. The giant mitochondria and dense lamellar bodies were observed in these cells. The microvilli on cell surfaces form a microlabirynth in intercellular and perivascular spaces.

The aim of the present study was to assess whether glucocorticoids could be directly involved in the maturation of adrenocortical cells from 120-138 days old ovine fetuses. The cAMP response to ACTH1-24 of cells cultured for 24 hours in the presence of ACTH1-24 was 2 fold higher than that of control cells. However, the response of cells cultured in the presence of ACTH1-24 plus metyrapone or aminoglutethimide was lower than that of cells cultured in the presence of ACTH1-24 alone. Cells cultured for 48 hours in the presence of dexamethasone or cortisol released more cAMP than control cells when stimulated by ACTH1-24, but not in response to forskolin. However corticosteroid production stimulated by ACTH1-24, forskolin or dibutyryl cAMP was enhanced by dexamethasone treatment. These results suggest that glucocorticoids can affect the maturation of ovine fetal adrenocortical cells by an auto and/or a paracrine process, and that this effect is exerted, at least, at two different levels in the cell.

The adrenal glands from a series of 300 fetuses and infants which had died in the Hospital Saint-Vincent-de-Paul, Paris, were dissected, weighed and processed for routine histology. The development of the fetal cortex and its regression have been compared to the general development of the gland. Our results show that there is an important development of the fetal cortex up to birth, when the adrenal glands represent almost one third of the size of the kidneys and possess only very reduced permanent cortex and practically no medulla. The regression of the fetal cortex in the postnatal period is marked by a decrease in weight of the adrenal glands up to the 2nd or 3rd month, by the persistence of a few dispersed cells up to the fifth month and by the persistence of a few fibrous elements up to 2 years of age.

The effects of corticosterone and cortisol in concentrations attainable in the adrenal gland were studied on ACTH-induced steroidogenesis in cultured cortical cells of foetal human and rat adrenals. Corticosterone at a concentration of 5.8 x 10(-5) mol/l clearly inhibited cortisol production (65.5%; P less than 0.005) and simultaneously increased androgen production in tissue culture of foetal human adrenals. Cortisol at a concentration of 2.8 x 10(-4) mol/l clearly inhibited 18-OH-DOC (74.0%, P less than 0.001) and aldosterone (83.7% P less than 0.005) production in tissue culture of foetal rat adrenals. In primary culture of foetal human adrenals cortisol did not decrease aldosterone production absolutely, but it significantly decreased the relative amount of aldosterone with respect to corticosterone. Cortisol did not inhibit corticosterone production in either culture. The results demonstrate that cortisol and corticosterone have qualitatively different effects on adrenal steroidogenesis and that these steroids may play a basic role in the functional zonation of the adrenal gland.

By means of transmissive electron microscopy the adrenals have been studied in 25 human embryos and fetuses at the age of 6-36 weeks. Certain stages have been revealed in formation of the adrenal cortex microcirculatory bed. In 6-7-week-old embryos (period of diffuse protocapillary bed) endothelial structure and mesenchymal cells, surrounding the adrenal anlage, resemble one another. A distinguished feature of the endothelium is regularly revealed desmosomes and large vacuoles, often found in cytoplasm of endotheliocytes. In 8-12-week-old fetuses (period when the organospecific microcirculatory bed is forming) sinusoid capillaries differentiate in the internal zone of the adrenal cortex; in endothelium fenestrae, "hatches", "locks" are revealed, the capillary basal membrane is formed. During subsequent time of the intrauterine development perfection of the microcirculatory pathways in the adrenals takes place, the arteriolar link of the subcapsular layer including. By the time of birth morphofunctional maturity of the microcirculatory bed in the adrenal cortex is noted.

21-Hydroxylase is a member of the P-450 superfamily of genes involved in the biosynthesis of cortisol and aldosterone in the adrenal cortex. Congenital adrenal hyperplasia, a well-characterized disease, originates from a lack of this enzyme. We present in this report an in situ hybridization study aimed at detecting 21-hydroxylase activity during murine development, from mid gestation to adulthood. Our results demonstrate that even during the embryonic period the adrenal cortex is the only major site of transcription of this enzyme, which is detectable beginning at embryonic day 14. In addition, a peculiar topographical pattern of transcriptional activity, characteristic of the stage of differentiation of the gland, could be drawn. Using a computer-assisted method, we were able to quantitate the relative transcription level at each stage of development. A steady increase in the level of transcription was demonstrated throughout embryonic life to birth, with a drop during the prepubertal period and a final rise at adult age. The possible physiological significance of our findings is discussed.

The formation of several steroids was determined in vitro in adrenals removed from 18 female and six male fetuses of 113-115 days' gestation and in two female and two male fetuses at near term (137-143 days). Samples were incubated with 14C-acetate and the formation of labeled steroids was determined by two-dimensional paper chromatography. Protein and corticosterone concentrations were determined by chromophore absorption and acid hydrolysis fluorescence methods, respectively. Tissue corticosterone concentrations were significantly higher in female (0.145 +/- 0.010 microgram/mg protein) than in male (0.083 +/- 0.010 microgram/mg protein) adrenal tissue at both stages, whereas corticosterone formation was similar in both sexes. Cholesterol formation was significantly higher in female (0.103 +/- 0.079 muM/mg protein) than in male (0.044 +/- 0.011 muM/mg protein) adrenals at both stages. Both testosterone and estradiol were synthesized at higher rates in female than in male adrenals (52% and 33%, respectively), whereas pregnanediol formation was 21% higher in the male. These results indicate that significant sex differences exist in the formation of various adrenocortical hormones by fetal tissues. The relevance of these findings to better survival of female premature newborns from respiratory distress syndrome in contrast with male, is discussed.

In the hamster, the weight of the adrenal glands increases during the course of gestation, with the highest value at day 5. In comparison to non-pregnant control animals, there were no changes in the volume of the zona glomerulosa (ZG) and zona fasciculata (ZF), while the volume of the zona reticularis (ZR) increased notably. The average volume of ZG-cells rose at day 5 of pregnancy and thereafter gradually decreased to that of control hamsters. A marked drop in the volume of ZF-cells was seen at days 5 and 10 of pregnancy, whereas at day 15 the cells were larger than in controls. At day 5 of pregnancy, a conspicuous increase in the cell volume was found in ZR, followed by lower values at day 10 and again higher than in control hamsters at day 15. The total number of parenchymal cells in hamster adrenal cortex increased at day 10 of gestation, then underwent a marked decrease, reaching the control value at the final day of pregnancy; this drop was mainly due to a reduction in the number of ZF-cells. The changes in the cell volume were paralleled by rather proportional changes in the volume of the mitochondrial compartment and in the quantity of smooth endoplasmic reticulum. The volume of the lipid-droplet compartment significantly rose in the course of gestation in both ZF- and ZR-cells. The cortisol output by adrenal homogenates gradually decreased during pregnancy.

The steroidogenic capacity of cells cultured from the definitive zone and fetal zone of the human fetal adrenal cortex was compared, using a serum-free medium without lipoproteins. Comparison of [3H]pregnenolone and [3H]progesterone metabolism in cultures from each zone incubated without ACTH indicated that only 3 beta-hydroxysteroid dehydrogenase, delta 4,5-isomerase (3 beta-HSD) activity was deficient in fetal zone cultures. Basal 3 beta-HSD activity was 3- to 5-fold lower in fetal zone cultures than in definitive zone cultures assayed after 3 or 10 days in culture. Although basal hydroxysteroid sulfotransferase activity was 2- to 4-fold greater in 3-day-old fetal zone cultures than in definitive zone cultures, this difference was not found in 10-day-old cultures due to a 3-fold decrease in fetal zone basal hydroxysteroid sulfotransferase activity. However, older cultures of fetal zone cells did maintain the characteristic high delta 5-steroid sulfate and low delta 4,3-ketosteroid basal production from [3H] pregnenolone as compared to definitive zone cultures. ACTH treatment for 48 h in serum-free medium increased the steroidogenic capacity of cell cultures from both zones and stimulated delta 4,3-ketosteroid production from [3H]pregnenolone and 3 beta-HSD activity in fetal zone cultures to levels characteristic of the definitive zone. These studies show that in the absence of ACTH the difference in steroidogenic capacity between the fetal zone and the definitive zone (due to the lower 3 beta-HSD activity in fetal zone cells) was maintained in cell cultures for a period up to 10 days.