In vitro steroidogenesis by the nonzoned adrenocortical tissue of the skink, Tiliqua rugosa.

Steroid formation by adrenocortical tissue from the shink, Tiliqua rugosa, has been studied using established in vitro techniques. Both in conventional incubations, with timed sampling, and in incubations with dialysis, aldosterone, and corticosterone were major products. From endogenous precursors, and from [¹⁴C]acetate, yields of the two products were of the same order, whereas from [³H]pregnenolone maximal yields of corticosterone were at least tenfold greater than aldosterone. Maximal rates of steroid formation from the radioactive precursors occurred within the first few minutes of incubation, but maximal rates of steroid formation from endogenous precursors occurred significantly later, between 1–2 hr.In incubations with dialysis [¹⁴C]aldosterone was significantly less dialysable than [³H]aldosterone under all conditions, whereas [¹⁴C] and [³H]corticosterone were homogeneous. In contrast, neither aldosterone nor corticosterone formed from endogenous precursors were bound under control conditions, although binding was increased following dexamethasone pretreatment, and decreased following stimulation with Tiliqua pituitary extract (but not Synacthen), with concomitant changes in yields and specific activities.Inter alia the results suggest that products formed from [¹⁴C]acetate and from [³H]pregnenolone may be maintained in separate pools within the tissue, and this accounts for their different metabolic fates. The bound pool, penetrated only by [³H]acetate, yields more aldosterone than the free, and may be termed a “biosynthetic pool.” In addition there exists a “secretory reserve pool.” This is suggested by the difference between rates of steroid secretion from endogenous and added precursors, and also from the changes in dialysibility seen in steroids formed from endogenous precursors under different conditions of stimulation.In both the compartmental arrangement of steroids, and the production of large yields of aldosterone the adrenocortical tissue of Tiliqua shows similarities to the zona glomerulosa, but not the inner zones of the rat adrenal cortex.     

Sex differences in steroidogenesis by adrenal homogenates of adult possum (Trichosurus vulpecula) attributable to the steroids formed by the adrenocortical special zone of the female

A marked sex difference was found in the nature of steroid conversion products formed from [¹⁴C]progesterone by whole adrenal homogenates of adult possums. The major conversion products from adrenals of males were: cortisol in yields of 46 ± 2% and corticosterone, 25 ± 2% (mean ± SEM, n = 35). The total 5β-reduced products were <10%. With glands from females 5β-reduced products comprised >80% of the yield, the predominant steroids being 5β-pregnane-3α,17α-diol-20-one, 25 ± 4% and 5β-pregnane-3α-o1-20-one, 21 ± 3% (mean ± SEM, n = 39). The yields of cortisol was <8%. No significant difference was found in the yields of products from animals used on the day of capture from the wild, or taken from the animal house 1–12 months after capture. Studies were carried out on the separated adrenocortical special zone and cortex proper of females. Homogenized cortex proper incubated with [¹⁴C]progesterone formed mainly: cortisol, 28 ± 2%; corticosterone, 18 ± 3%; and 11β-hydroxyprogesterone, 24 ± 4% (mean ± SEM). From the same substrate the conversion products by the homogenized special zone were 5β-reduced pregnane derivates in yields >70% and reduced 5α- and 5β-androstan derivates, <15%. The products of major yield were: 5β-pregnan-3α-o1-20-one, 5β-pregnane-3α,17α-diol-20-one, and 5β-pregnane-3α,17α,20α-triol. Similarly from [17α-¹⁴C]hydroxyprogesterone and [11-¹⁴C]deoxycortisol substrates the conversion products by the special zone were mainly 5β-reduced pregnane derivates, the C19 steroids being <10%. Dissected medullary tissue had no steroidogenic activity. The addition of medullary tissue to cortex proper had no effect on the synthetic activity of the cortex. With mixtures of cortex proper and special zone the conversion products were similar to those obtained by whole adrenal homogenates. It is concluded that the dissimilarity in the adrenal steroid formation was due to the activity of the 5β-reductase, which prevailed in the female adrenal and was very low in the male adrenal. It was found that the location of the reducing enzymes was specific to the unique adrenocortical special zone which is present only in the female possum.     

Steroid synthesis in the adrenal gland of the sea snake Hydrophis cyanocinctus: The metabolism of exogenous precursors

Sea snake (Hydrophis cyanocinctus) adrenal glands (whole homogenates, preincubated minces, or mitochondrial preparations) were incubated in vitro with exogenous radioactive precursors. Hydrophis adrenal tissue was capable of synthesizing 17-deoxycorticosteroids from exogenous cholesterol, pregnenolone, progesterone, and DOC, but not from sodium [¹⁴C]-acetate. Products identified after incubation were pregnenolone, progesterone, 11β-hydroxyprogesterone, DOC, corticosterone, 18-hydroxycorticosterone, and aldosterone. The major product was corticosterone with lesser quantities also of 18-hydroxycorticosterone and aldosterone. In the case of the mitochondrial preparation 11β-hydroxyprogesterone predominated. No evidence for the biosynthesis of cortisol from cholesterol was found. Two types of kinetic incubation were employed: One sampled the incubation medium alone, while the other sampled both medium plus tissue. It was concluded that sampling the medium only did not allow the identification of the biosynthetic pathways operating in vitro. However, from sampling both the medium and the tissue it was concluded that both the C21-C11 and C11-C21 sequences of hydroxylation operated in the conversion of progesterone to corticosterone. The data contrast with those obtained from previous studies on cobra adrenal tissue, particularly with regard to the ability of sea snake adrenals in vitro to 18-oxygenate exogenous precursors.     

Steroid biosynthesis by adrenal tissue of snowshoe hares (Lepus americanus) collected in a year of peak population density.

Homogenates of adrenal glands from individual snowshoe hares (Lepus americanus) collected during a year of peak population density were incubated for 3-hr periods with [4-¹⁴C] pregnenolone substrate. At substrate concentrations of 82.5 μM, the primary product formed was cortisol, accounting for 32–37% of the added substrate. No corticosterone formation was detected. At 250 μM substrate concentrations, the formation of cortisol, 17α-hydroxypregnenolone, and corticosterone accounted for 13–21, 23–38, and 1.6–4.0% of the added substrate, respectively. Unmetabolized pregnenolone accounted for 4–13% of the exogenous substrate in the incubations with the lower substrate concentrations, and 32–50% in the ones with the higher concentrations. The data suggest that snowshoe hare adrenal tissue primarily forms cortisol during years of peak population density, and that 17α-hydroxypregnenolone is an intermediate in the biosynthetic pathway.     

Seasonal variation in adrenal 11beta-hydroxysteroid dehydrogenase activity in the meadow vole (Microtus pennsylvanicus).

[4-¹⁴C]Progesterone is converted to corticosterone by adrenal homogenates of Microtus pennsylvanicus. The 11-keto derivative, 11-dehydrocorticosterone, is a secondary conversion product formed by the action of an adrenal 11β-hydroxysteroid (11β-OHD) dehydrogenase on corticosterone. There is a marked seasonal variation in the 11β-OHD enzyme with peak activity in late fall-winter and very low activity in early spring and summer. Adrenal 11β-OHD enzyme activity is greatest during the period when adrenal responsiveness is low and the voles are reproductively inactive.     

Formation of corticosteroids in vitro by interrenal tissue from the teleost fish, Coregonus clupeoides.

Adrenocortical tissue from the freshwater teleost, Coregonus clupeoides, was incubated with [¹⁴C] acetate and [³H] pregnenolone. Evidence was obtained for the formation of doubly labeled cortisol, cortisone, and deoxycortisol, and the 17-deoxycorticoids, corticosterone, deoxycorticosterone, and aldosterone. This corresponds very closely to the range of steroids isolated from circulating plasma in this species.Under conditions of incubation with dialysis, it was found that the tritiated products tended to be freely dialysable, whereas the [¹⁴C] products were significantly less so. The addition of Coregonus pituitary extract to the incubation medium caused both a release of [¹⁴C] steroid from the bound condition and a significant decrease in the yield of [¹⁴C] aldosterone; neither effect was seen with Synacthen. In all cases tritiated products were largely unaffected both in yield and dialysability.The results were consistent with the view that aldosterone is preferentially formed through a pathway involving bound intermediates. The native ACTH-sensitive bound pool of steroids and intermediates is penetrable by exogenous precursors occurring early in the biosynthetic pathway, such as [¹⁴C] acetate, but not by late precursors such as [³H] pregnenolone. The results are comparable with those obtained with other species, including the skink, Tiliqua rugosa and the zona glomerulosa (but not the inner zones) of the rat adrenal cortex.     

Evidence of the in vitro formation of cortisol by the adrenal gland of embryonic and young chickens (Gallus domesticus)

The aim of this study is to elucidate the ontogenetic aspect of corticosteroidogenesis in the chicken. The adrenal gland of embryonic and very young chicks contains an enzyme system which converts progesterone to 17α-hydroxyprogesterone. 4-¹⁴C-Labeled cholesterol, pregnenolone, progesterone, and 17α-hydroxyprogesterone were incubated with the homogenates of adrenal gland from 17- and 21-day-old chick embryos and chickens. The metabolic products were identified by their mobilities on a thin-layer chromatogram and recrystallization to constant ³H:¹⁴C ratio after adding the corresponding ³H-labeled steroid. Cholesterol was metabolized to pregnenolone in the tissue homogenates from chick embryos and chickens at all ages. Pregnenolone was metabolized to progesterone, 11-deoxycorticosterone, and other minor metabolites, but not to 17α-hydroxyprogesterone. The major products from progesterone were 11-deoxycorticosterone and 17α-hydroxyprogesterone. The yield of 17α-hydroxyprogesterone decreased with advancing age and became zero at 7 days posthatching. 11-Deoxycortisol and cortisol were produced from progesterone by the homogenates from 17- and 21-day-old embryos and 3-day-old chicks, but neither was produced by those from 7-day-old chicks or those from 150-day-old hens. Radioactive 17α-hydroxyprogesterone was converted to 11-deoxycortisol and cortisol in large amounts and to cortisone in small amounts. Androstenedione and testosterone were detected in the adrenal homogenate from 17 days of incubation to 7 days posthatching, but not in the tissue from 14 days posthatching. The activity of 17α-hydroxylase was high at 17 days of incubation, decreasing with advancing age, and disappeared between 7 and 14 days posthatching. These results represent definite evidence of cortisol and testosterone formation in vitro by embryonic and very young chick adrenals.     

In vitro corticosteroidogenesis by the adrenal gland of the chicken (Gallus domesticus)

[4-¹⁴C]Pregnenolone, [4-¹⁴C]progesterone, and [4-¹⁴C]11-deoxycorticosterone were indubated with chicken adrenal tissue slices, whole homogenates, and subcellular fractions, with and without the addition of ACTH to the incubation medium. Progesterone, 11-deoxycorticosterone, corticosterone, 11β-hydroxyprogesterone, and aldosterone were identified as metabolites of these radioactive precursors. The rate of conversion of pregnenolone to progesterone by the slices and progesterone to corticosterone by the mitochondrial fraction significantly increased by the addition of ACTH to the medium. The activity of Δ⁵-3β-hydroxysteroid dehydrogenase associated with Δ⁵-Δ⁴ isomerase upon pregnenolone and the activity of 21-hydroxylase upon progesterone were concentrated in the microsomal fraction, while the activity of 11β-hydroxylase upon 11-deoxycorticosterone was in the mitochondrial fraction. No 17α-hydroxylase activity was observed.The main pathway for steroidogenesis in the chicken adrenal gland is proposed to be: pregnenolone → progesterone → 11-deoxycorticosterone → corticosterone.     

123I-metaiodobenzylguanidine (MIBG) scintigraphy for the detection of adrenal and extra-adrenal phaeochromocytomas: CT and MRI correlation

Context Evidence regarding the accuracy of [¹²³I] metaiodobenzylguanidine (MIBG) imaging for phaeochromocytoma localization is currently limited to small series. Objective We present the largest series of primary phaeochromocytomas in which the performance of [¹²³I]MIBG has been evaluated and correlated with cross‐sectional imaging. Design We identified 76 patients with both preoperative [¹²³I]MIBG and cross‐sectional imaging for confirmed primary phaeochromocytoma between 1995 and 2005 at our institution. This comprised 60 adrenal tumours in 55 patients and 33 extra‐adrenal tumours in 23 patients (2 patients had both adrenal and extra‐adrenal tumours). Phaeochromocytoma metastases were not evaluated. Main outcome measure(s) [¹²³I]MIBG studies were independently reviewed and correlated with CT and MRI examinations, as well as tumour functional status, to identify features that may predict a false negative [¹²³I]MIBG result. Results The overall sensitivity of [¹²³I]MIBG was 75%. Tumour detection was lower for extra‐adrenal (58%) vs. adrenal (85%) phaeochromocytomas (P = 0·005). For extra‐adrenal tumours, [¹²³I]MIBG demonstrated 8 of 14 carotid body, 2 of 2 intrathoracic, 8 of 14 retroperitoneal and 2 of 3 pelvic phaeochromocytomas. Overall, MRI and CT demonstrated 68 of 68 and 72 of 74 primary phaeochromocytomas, respectively. Tumour size correlated with [¹²³I]MIBG uptake for adrenal (P = 0·009) but not extra‐adrenal tumours. When tumours were adjusted for size, no other imaging feature or functional status correlated with [¹²³I]MIBG negativity, although two large [¹²³I]MIBG negative adrenal tumours contained large areas of necrosis or haemorrhage. Conclusions Extra‐adrenal and small adrenal phaeochromocytomas are more likely to result in false negatives on [¹²³I]MIBG. Tumoural necrosis or haemorrhage do not consistently relate to [¹²³I]MIBG uptake, although adrenal phaeochromocytomas containing minimal solid tissue due to extensive necrosis may predict a negative [¹²³I]MIBG result.     

Steroid biosynthesis by the ovary of the European eel, Anguilla anguilla L., at the silver stage.

The ovary of the European eel, Anguilla anguilla, at the silver stage, was incubated either as an intact tissue preparation or as a homogenate with and without cofactors in the presence of [4-¹⁴C] pregnenolone and [4-¹⁴C]progesterone. Intact tissue incubates displayed a more complex metabolite profile than reinforced homogenates, and deprivation of exogenous cofactors reduced the profile even further. Among the metabolites derived from pregnenolone, the following steroids were identified by their isopolarity and isomorphicity with standard compounds: 17α-hydroxypregnenolone; dehydroepiandrosterone; progesterone; 17α-hydroxyprogesterone; and androstenedione. The last three steroids plus testosterone, 17β-hydroxyandrostenedione, and adrenosterone were identified using progesterone as a precursor. Metopirone inhibited the formation of 11-oxygenated androgens. 11-Deoxycorticosteroids were not found, indicating the absence of steroid 21-hydroxylase activity in the eel ovary. Integration of the product yield-time curves demonstrates that in vitro the activities of the enzymes 3β-, 17β-, and 11β-hydroxysteroid dehydrogenases were less apparent than those of steroid 17α,20-C₂₁-desmolase, and 17α-, and to a lesser extent 11β-hydroxylase. Irrespective of the incubation conditions, pregnenolone produced more Δ⁵-3β-hydroxy-thanΔ⁴-3-ketosteroids, suggesting a predominance of the former biosynthetic pathway. Among the unidentified metabolites, water-soluble compounds were formed from both precursors in intact tissue incubates.     

Bomb-curve radiocarbon measurement of recent biologic tissues and applications to wildlife forensics and stable isotope (paleo)ecology

Above-ground thermonuclear weapons testing from 1952 through 1962 nearly doubled the concentration of radiocarbon (¹⁴C) in the atmosphere. As a result, organic material formed during or after this period may be radiocarbon-dated using the abrupt rise and steady fall of the atmospheric ¹⁴C concentration known as the bomb-curve. We test the accuracy of accelerator mass spectrometry radiocarbon dating of 29 herbivore and plant tissues collected on known dates between 1905 and 2008 in East Africa. Herbivore samples include teeth, tusks, soft tissue, hair, and horn. Tissues formed after 1955 are dated to within 0.3–1.3 y of formation, depending on the tissue type, whereas tissues older than ca. 1955 have high age uncertainties (>17 y) due to the Suess effect. ¹⁴C dating of tissues has applications to stable isotope (paleo)ecology and wildlife forensics. We use data from 41 additional samples to determine growth rates of tusks, molars, and hair, which improve interpretations of serial stable isotope data for (paleo)ecological studies. ¹⁴C dating can also be used to calculate the time interval represented in periodic histological structures in dental tissues (i.e., perikymata), which in turn may be used as chronometers in fossil teeth. Bomb-curve ¹⁴C dating of confiscated animal tissues (e.g., ivory statues) can be used to determine whether trade of the item is legal, because many Convention of International Trade of Endangered Species restrictions are based on the age of the tissue, and thus can serve as a powerful forensic tool to combat illegal trade in animal parts.     

Steroid formation in the larval and parasitic adult sea lamprey, Petromyzon marinus L.

Presumptive adrenocortical tissue (PAT) from all known sites in parasitic adults or larval sea lamprey, Petromyzon marinus L., were incubated with [4-¹⁴C]progesterone in a buffered medium with an NADPH generating system. Testicular tissue from parasitic adults was similarly incubated. PAT from both parasitic adults and larvae formed 11-deoxycortisol (S), 17α-hydroxyprogesterone (17αOHP), and androstene dione (AD) but no cortisol (F) cortisone (E), corticosterone (B), 11-deoxycorticosterone (DOC), or testosterone (T) were formed. Therefore PAT may be lacking in 11β-hydroxylase activity but does contain 17α- and 21-hydroxylase and 20-desmolase activities. Testicular tissue failed to produce F, E, B, S, T, 17αOHP, or AD. However, testicular tissue did form DOC indicating the presence of 21-hydroxylase activity. After the 4-hr incubations, histological examination indicated that the PAT and testicular tissue were normal in appearance.     

Effects of gonadectomy and replacement therapy on adrenal function in the domestic duck, Anas platyrhynchos

Adrenal function in ducks of both sexes was studied in intact and in castrated animals and in castrated animals receiving appropriate replacement therapy. No significant changes in adrenal weights were observed in the various groups of animals. In the males, in vitro production of corticosteroids from progesterone-4-¹⁴C by both the intact and castrated animals receiving testosterone phenylpropionate were significantly less than by the castrates, indicating an inhibitory effect of androgens on adrenal activity. No differences were observed in the three groups of female ducks.     

Prostaglandin formation from exogenous precursor in homogenates of human cardiac tissue

Summary The ability of some different components of human cardiac tissue to synthesize prostaglandins (PGs) from exogenous precursor was investigated. Fractions of the cardiac tissue containing either parts of valve cusps and the papillary musculature or some selected tissue components such as myocytes, endocardial elements, endothelial cells and connective tissue were prepared and homogenized. Lowspeed supernatants of the various homogenates thus obtained were incubated with [¹⁴C]-labelled arachidonate ([¹⁴C]-AA). [¹⁴C]-PGs formed in the incubates were extracted, separated by thin-layer chromatography and quantified using liquid scintillation spectometry.In the incubates of all the fractions [¹⁴C]-AA was converted to [¹⁴C]-PGs with a time-dependent yield, most effectively at 3 minutes' incubation time. The endocardial and endothelial fractions were found to exhibit the highest cyclo-oxygenase activity, the [¹⁴C]-AA conversion rate in these incubates being twice as high as in the others. [¹⁴C]-labelled PGF_2a and 6-keto-PGF_1a were found to be the principal PG products and there was no evidence of TxB₂ formation in any of the incubates. [¹⁴C]-6-keto-PGF_1a was the main PG formed, constituting about 40% or more of the [¹⁴C]-PG activity in the incubates of all the fractions, whereas labelled PGE₂ and PGF_2a were observed in considerably smaller and nearly equal amounts.The results demonstrate a considerable ability of human cardiac tissue to synthesize prostacyclin (PGI₂) and, at the same time, the existence of local differences in tissue cyclo-oxygenase activity, which appears to be significantly higher in the endocardial layer than in the myocardium itself.     

Characterization of angiotensin II receptors in the anterior pituitary gland

Specific and high affinity binding sites for angiotensin II were demonstrated in the anterior pituitary gland by binding studies with [¹²⁵I]iodoangiotensin II. The binding properties of the pituitary receptors were similar to those of angiotensin II receptors present in the adrenal gland. The concentration of binding sites in rat anterior pituitary (293 ± 50 fmoles/mg protein) was less than in the adrenal gland, but was much greater than in smooth muscle. Angiotensin II receptors were identified in the anterior pituitary tissue of mature and immature animals of both sexes, and in species including rat, rabbit and dog. No binding of angiotensin II was detected in posterior pituitary homogenates, or in GH₃ pituitary tumor cells. Collagenase-dispersed anterior pituitary cells also contained specific binding sites for angiotensin II, with equilibrium binding constant (K_a) of 3.6 × 10⁹ M^?1. The presence of specific high-affinity angiotensin II receptor in the anterior pituitary gland provides a mechanism by which angiotensin-like peptides could modulate the process of pituitary hormone secretion.     

Physiology of mammary glands: Edited by Akira Yokoyama,Hideo Mizuno,and Hiroshi Nagasawa. University Park Press,Baltimore. $44.50. x + 387 pp.; Illustr.; Subject index. 1978

A perifusion system technique has been used to compare the effects of metyrapone (SU-4885; Ciba) and aminoglutethimide, two compounds which are known to inhibit corticosteroid biosynthesis in mammal adrenal glands, upon aldosterone output by frog interrenal tissue. Rana ridibunda adrenal fragments were continuously perifused for 13 hr with amphibian culture medium. Fractions were set apart every 5 min and aldosterone levels were measured by means of a specific and sensitive radioimmunoassay method. For doses ranging from 1 × 10^?6to 1 × 10^?3M, metyrapone induced a decrease in aldosterone secretion rate. The infusion of three equimolar doses (1 × 10^?5M)_of metyrapone during three consecutive periods of 1 hr or during various times (30, 60, 150 min) within the same experiment made it possible to study the kinetics of the response of the glands. The lag period (10 min), the amplitude of the inhibition (75%), and the duration of the inhibitory effect after withdrawal of metyrapone (50 min) were almost uniform, whatever the duration of the infusion. Similar patterns were obtained using aminoglutethimide, although much higher doses (up to 100-fold) were required to inhibit aldosterone production. These results demonstrate that both metyrapone and aminoglutethimide are potent inhibitors of mineralocorticoids output in amphibia. Conversely, they suggest that a single injection of each inhibitor, in vivo, would only induce a transient and light diminution in corticosteroid secretion.     

Synthesis and degradation of methylated proteins of mouse organs: Correlation with protein synthesis and degradation

L-(Methyl-¹⁴C)-methionine was administered i.p. to mice, and the incorporation of radioactive methionine into proteins and methyllysine and methylarginine residues formed by the transfer of the methyl-¹⁴C group of methionine were measured.Tissue protein was actively methylated in organs having a high activity of protein synthesis, and the in vivo methylating activity in organs was not correlated with the protein methylating activity of the organs determined in vitro. Puromycin inhibited both protein synthesis and protein methylation in mouse organs to a similar degree. Neither the formation of S-adenosyl-(methyl-¹⁴C)-methionine nor protein methylase was inhibited by puromycin. The data suggests that proteins are methylated immediately after protein synthesis, that is, newly synthesized proteins are the substrates of protein methylation.Radioactive methionine and the [C¹⁴] methyl groups of methyllysine and methylarginine residues of tissue proteins are degraded in parallel over a period of 3 wk, suggesting that protein methylation is an irreversible type of protein modification.     

Metabolism of pregnenolone-4-14C and pregnenolone-7-alpha-3H sulfate by the Macaca mulatta fetal adrenal in vitro

These studies were initiated to ascertain the feasibility of utilizing the fetal monkey adrenal as a model for further studies on comparative steroid metabolism at various periods of gestation. Homogenates of midtrimester fetal monkey (Macaca mulatta) adrenals were incubated simultaneosly with pregnenolone-4-¹⁴C and pregnenolone-7α-³H sulfate. Conversion of both substrates to 17α-hydroxypregnenolone, 17α-hydroxyprogesterone, dehydroepiandrosterone, androstenedione, 11β-hydroxyandrostenedione, and cortisol as well at 17α-hydroxypregnenolone sulfate and dehydroepiandrosterone sulfate was demonstrated. Conversion of pregnenolone to progesterone was also shown. Neither free nor conjugated 16α-hydroxypregnenolone or 16α-hydroxydehydroepiandrosterone were found.     

Effect of aging and adrenocorticotropin on adrenal Δ5-3β-hydroxysteroid dehydrogenase activity in male rats

Adrenal Δ⁵-3β-hydroxysteroid dehydrogenase (3β-HSD) activity was determined in male rats 4, 6, 12, 18 and 24 months of age. Mean (± SE) adrenal 3β-HSD concentration (μg Δ⁴-androstenedione formed/minute/mg tissue), specific activity (μg/minute/mg protein) and total content (μg/minute/pair of adrenals) were less (p<0.001 to p<0.025) in male rats 12 months of age (0.222 ± 0.010, 1.66 ± 0.09 and 8.6 ± 0.8, respectively) or older, than in males four months of age (0.372 ± 0.011, 2.69 ± 0.07 and 13.4 ± 1.1, respectively). Subcutaneous administration of 10 IU adrenocorticotropin daily for a period of five days to male rats 24 months of age elevated adrenal weight by 50 percent and restored dehydrogenase activity to that of the young untreated animal. Therefore, adrenal function in male rats as determined by 3β-HSD activity declines with advancing age, but remains responsive to adrenocorticotropic stimulation.