G protein receptors 7 and 8 are expressed in human adrenocortical cells, and their endogenous ligands neuropeptides B and w enhance cortisol secretion by activating adenylate cyclase- and phospholipase C-dependent signaling cascades.

Neuropeptides B and W (NPB and NPW) are regulatory peptides that act via two subtypes of G protein-coupled receptors, named GPR7 and GPR8. RT-PCR demonstrated the expression of these receptors in both zona glomerulosa and zona fasciculata-reticularis (ZF/R) cells of the human adrenal cortex. NPB and NPW did not affect aldosterone secretion from dispersed zona glomerulosa cells but enhanced cortisol production from ZF/R cells, NPB being more effective than NPW. NPB evoked sizable cAMP and inositol triphosphate responses from ZF/R cells, which were abrogated by the adenylate cyclase inhibitor SQ-22536 and the phospholipase C inhibitor U-73122, respectively. Cortisol response to NPB was lowered by either SQ-22536 and the protein kinase (PK) A inhibitor H-89 or U-73122 and the PKC inhibitor calphostin-C and abolished by the simultaneous exposure to H-89 and calphostin-C. NPW elicited only a rise in cAMP production from dispersed ZF/R cells, and its cortisol response was suppressed by both SQ-22536 and H-89. PreproNPB and preproNPW mRNAs were detected in human adrenal cortexes. We conclude that: 1) NPB and NPW exert a secretagogue action on human ZF/R cells, probably acting in an autocrine-paracrine manner; and 2) the effect of NPB is mediated by both the adenylate cyclase/PKA and the phospholipase C/PKC cascades, whereas that of NPW involves only the activation of the former signaling pathway.

Effects of prolonged cyclosporine-A treatment on the morphology and function of rat adrenal cortex.

The effects of prolonged (30 day) treatment with daily therapeutical doses of cyclosporine A (CSA) (20 mg/kg) on the function and morphology of adrenal cortex were studied in adult male rats. CSA-treated animals developed a notable hypertension, along with a striking rise in PRA, which was not coupled with significant changes in the plasma concentrations of aldosterone and corticosterone (hyperreninemic hypoaldosteronism). Morphometry showed that zona glomerulosa (ZG) and zona fasciculata, and their parenchymal cells were atrophic. Isolated capsular (ZG) and inner (zona fasciculata/reticularis) cells displayed reduced basal and stimulated secretory responses. However, while the response of ZG cells to angiotensin II was almost completely suppressed (96%), basal steroid secretion of isolated cells, as well as the aldosterone and corticosterone response of ZG cells to potassium and ACTH, and corticosterone production of inner cells in response to ACTH were decreased by only about 30-40%. The hypothesis is advanced that CSA exerts a dual effect on rat adrenal cortex: 1) a general inhibitory effect on the growth and steroidogenic capacity of adrenocortical cells, which manifests itself only after very prolonged treatment and may be caused by an impairment of protein synthesis; and 2) an acute effect involving the specific blockade of the angiotensin-II-induced stimulation of the secretory activity of ZG cells.

Endothelins stimulate deoxyribonucleic acid synthesis and cell proliferation in rat adrenal zona glomerulosa, acting through an endothelin A receptor coupled with protein kinase C- and tyrosine kinase-dependent signaling pathways.

The effects of endothelins (ET) on the proliferative activity of the rat adrenal cortex have been investigated in vivo, using an in situ perfusion technique of the intact left gland. The chemicals were dissolved in the perfusion medium, and the perfusion continued for 120 min. ET-1 concentration dependently increased the mitotic index and [3H]thymidine incorporation into DNA in the zona glomerulosa (ZG; 6- and 3-fold increases, respectively, at a 10(-8) M concentration), but not in the inner adrenocortical layers, where the basal proliferative activity was negligible. The effect of 10(-8) M ET-1 was blocked by the ETA receptor antagonist BQ-123, whereas the ETB receptor antagonist BQ-788 was ineffective. ET-2 and ET-3 (10(-8) M) enhanced DNA synthesis in the ZG, but their effects were less intense than that of 10(-8) M ET-1 and were directly related to their binding potency for the ETA receptor subtype (ET-1 > ET-2 >> ET-3). The selective ETB receptor agonists BQ-3020, IRL-1620, and sarafotoxin-6B were ineffective. The ZG proliferogenic action of 10(-8) M ET-1 was reversed by both the protein kinase C inhibitor Ro31-8220 and the tyrosine kinase inhibitor tyrphostin-23; a complete blockade was obtained at a 10(-6)-M concentration of each inhibitor. In contrast, neither the protein kinase A inhibitor H-89 (10(-5) M) nor the cyclooxygenase and lipoxygenase inhibitors indomethacin and phenidone (10(-5) M) affected ET-1 action. Collectively, our findings indicate that ETs stimulate the proliferation of rat adrenal ZG cells, acting through ETA receptors coupled with protein kinase C- and tyrosine kinase-dependent signaling pathways. The results of the present study are in keeping with the view that in mammals, ZG is the proliferative layer involved in the maintenance of growth of the entire adrenal cortex and with the previous autoradiographic demonstration that ZG is the only adrenocortical layer provided with ETA receptors.

Zona glomerulosa morphology and function in streptozotocin-induced diabetic rats.

Streptozotocin-induced diabetic rats showed a significant lowering in both PRA (-31%) and basal plasma aldosterone concentration (-59%), coupled with a notable atrophy of the zona glomerulosa (-30%) and its parenchymal cells (-36%). Kalaemia and the blood level of ACTH were not affected. Insulin infusion reversed all the streptozotocin-evoked effects. Analogous, though less conspicuous, changes were induced by experimental diabetes also in rats whose hypothalamo-hypophyseal-adrenal axis and renin-angiotensin system had been pharmacologically interrupted by the simultaneous administration of dexamethasone-captopril and maintenance doses of ACTH-angiotensin II: the drops in the basal plasma aldosterone concentration and in the volume of zona glomerulosa and its cells ranged from -20% to -22%. In these animals, experimental diabetes significantly depressed the aldosterone response to the acute stimulation with angiotensin II (-55%), potassium (-50%), and ACTH (-43%). These findings indicate that the well known impairment of renin release may only partially account for the antiadrenoglomerulotrophic effect of experimental diabetes in rats. The hypothesis is advanced that the chronic lack of insulin may directly depress both the growth of the zona glomerulosa and the newly synthesis of some enzymes of aldosterone synthesis.

Investigations on the morphology and function of adrenocortical tissue regenerated from gland capsular fragments autotransplanted in the musculus gracilis of the rat.

This paper describes the function and morphology of regenerated adrenocortical nodules obtained by implanting, in the musculus gracilis of rats, several (n = 6-7) fragments of the capsular tissue of their excised adrenal glands. Four months after the operation, each bilaterally adrenalectomized rat developed six or seven well encapsulated adrenocortical nodules about 2-3 mm in diameter and always lacking chromaffin cells, and displayed almost complete normalization of basal and stimulated blood levels of corticosterone, but not of aldosterone. In vitro study showed that regenerated nodules were well functioning as far as glucocorticoid production was concerned. Accordingly, electron microscopy and stereology indicated that the majority of the parenchymal cells (independently of their location in the outer subcapsular, middle, or inner portions) closely resembled those of the zonae fasciculata/reticularis of the adrenal gland of age-matched sham-operated rats. By contrast, regenerated nodules evidenced a relative impairment in aldosterone secretion, and this was coupled with the presence of only a few zona glomerulosa-like cells. Such cells were grouped in small islets located near the few connective trabeculae detaching from the capsule, and autoradiography showed that they were the only parenchymal elements of the nodule able to bind [125I]angiotensin-II. The possibility is suggested that the paucity of zona glomerulosa-like cells in regenerated nodules could be ascribed to the absence of zona medullaris, which is currently thought to exert a paracrine control on the growth and secretion of zona glomerulosa in the rat adrenal glands.

Human pheochromocytomas express orexin receptor type 2 gene and display an in vitro secretory response to orexins A and B.

Orexins A and B are hypothalamic peptides, that act through two receptor subtypes, called OX1-R and OX2-R. OX1-R selectively binds orexin A, whereas OX2-R is nonselective for both orexins. High levels of OX1-R mRNA and low levels of OX2-R mRNA have been previously detected in the human adrenal cortex and medulla. Here we demonstrated by RT-PCR the expression of the OX2-R, but not the OX1-R, gene in 10 benign secreting pheochromocytomas. Both orexins A and B stimulated catecholamine secretion from pheochromocytoma slices; the maximal effective concentration was 10(-8) mol/liter. Orexins A and B (10(-8) mol/liter) increased IP3, but not cAMP production, by tumor slices, and the effect was blocked by the PLC inhibitor U-73122. The catecholamine response to 10(-8) mol/liter orexins A and B was abolished by either U-73122 or the PKC antagonist calphostin C and was unaffected by the adenylate cyclase inhibitor SQ-22536 and the PKA inhibitor H-89. Collectively, these findings suggest that orexins stimulate catecholamine secretion from human pheochromocytomas, acting through OX2-R coupled to the PLC-PKC signaling pathway.

Expression and function of vasoactive intestinal peptide, pituitary adenylate cyclase-activating polypeptide, and their receptors in the human adrenal gland.

VIP and pituitary adenylate cyclase-activating polypeptide (PACAP) are two regulatory peptides that possess remarkable amino acid sequence homology and act through common receptors, named PAC(1), VPAC(1), and VPAC(2). PAC(1) receptor is selective for PACAP, whereas VPAC(1) and VPAC(2) receptors bind both VIP and PACAP. We have investigated the expression and function of VIP, PACAP, and their receptors in the zona glomerulosa (ZG), zonae fasciculata and reticularis, and adrenal medulla (AM) of the human adrenal cortex. RT-PCR and RIA detected VIP and PACAP expression exclusively in AM cells. RT-PCR demonstrated the presence of PAC(1) mRNA only in AM and of VPAC(1) and VPAC(2) mRNAs in both ZG and AM cells. VIP and PACAP concentration-dependently increased aldosterone and catecholamine secretion from cultured ZG and AM cells. The catecholamine response to both peptides was higher than the aldosterone response, and the secretagogue action of PACAP was more intense than that of VIP. The aldosterone response of cultured ZG cells to VIP or PACAP was unaffected by the PAC(1) receptor antagonist PACAP-(6-38) (PAC(1)-A), but was significantly decreased by the VPAC(1) receptor antagonist [Ac-His(1),D-Phe(2),Lys(15),Arg(16)]VIP-(3-7),GH-releasing factor-(8-27)-NH(2) (VPAC(1)-A). The catecholamine response of cultured AM cells to VIP was lowered by VPAC(1)-A and unaffected by PAC(1)-A; conversely, the catecholamine response to PACAP was reduced by both PAC(1)-A and VPAC(1)-A. Simultaneous exposure to both antagonists did not abolish the catecholamine response to PACAP. Collectively, our findings allow us to conclude that in human adrenals 1) VIP and PACAP biosynthesis exclusively occurs in AM cells; 2) ZG cells are provided with functional VPAC(1) and VPAC(2) receptors, whose activation by VIP or PACAP elicits a moderate aldosterone response; 3) AM cells possess PAC(1), VPAC(1), and VPAC(2) receptors, whose activation evokes a marked catecholamine response; and 4) the catecholamine response to PACAP is more intense than that to VIP, because it is mediated by all subtypes of VIP/PACAP receptors.

Age-dependent changes in the function and morphology of mitochondria of rat adrenal zona fasciculata.

The function and morphology of adrenal zona-fasciculata (ZF) mitochondria were studied in 4-, 10- and 16-month-old rats, since in this species ageing causes a marked decline in glucocorticoid secretion coupled with high levels of circulating ACTH. Dispersed intact ZF cells displayed a significant age-dependent impairment of their basal pregnenolone (PREG) secretion, but isolated ZF mitochondria showed an increased capacity to convert cholesterol to PREG (the first rate-limiting step of steroid synthesis). These data are in keeping with the contention that the age-related deficit of rat ZF secretion is located prior to the activity of intramitochondrial cholesterol side-chain cleaving enzymes (cytochrome-P450scc). Stereology showed a notable age-dependent increase in the number of mitochondria per unit cell-volume, coupled with a marked decrease in their average volume. The width of the mitochondrial intermembrane space remained unchanged, but its average volume strikingly decreased. This last finding fits well with the enhanced capacity of mitochondria to produce PREG, since intermembrane space is an aqueous barrier to the translocation of free cholesterol from the outer membrane to the cristae, where cytochrome-P450scc is located. In conclusion, the hypothesis is advanced that all these age-related functional and morphological mitochondrial changes are an ACTH-dependent compensatory response enabling ZF cells to partially counteract their decreased glucocorticoid secretory capacity, which in turn is due to the impaired utilization of intracytoplasmic stores of cholesterol esters.

Endothelin-1[1-31] acts as a selective ETA-receptor agonist in the rat adrenal cortex.

Endothelin-1 (ET-1) is a 21-amino acid residue (ET-1[1-21]) hypertensive peptide, which together with its receptor subtypes A and B (ETA and ETB) is expressed in the rat adrenal cortex, where it stimulates steroid-hormone (aldosterone and corticosterone) secretion through the ETB receptor and the growth (proliferative activity) of the zona glomerulosa (ZG) through the ETA receptor. ET-1[1-21] is generated from bigET-1 by the endothelin-converting enzyme (ECE-1). However, recent evidence indicates the existence of an alternative chymase-mediated biosynthetic pathway leading to the production of an ET-1[1-31] peptide, which was found to reproduce the ETA receptor-mediated vascular effects of ET-1[1-21]. We found that ET-1[1-21], but not ET-1[1-31], concentration-dependently raised steroid secretion from dispersed rat adrenocortical cells, its effect being blocked by the ETB-receptor selective antagonist BQ-788. Both ET-1s concentration-dependently increased the number of "S-phase" cells (as detected by the 5-bromo-2'-deoxyuridine immunocytochemical method) in capsule-ZG strips within a 240 min incubation. The ZG proliferogenic action of both ET-1s was blocked by the ETA-receptor antagonist BQ-123, and ET-1[1-31] was found to be significantly more potent than ET-1[1-21]. Autoradiography showed that in the rat adrenal ET-1[1-21] displaced the binding of selective ligands to both ETA ([125I]PD-151242) and ETB receptors ([125I]BQ-3020), while ET-1[1-31] eliminates only the binding to ETA receptors. Collectively, our findings provide strong evidence that ET-1[1-31] acts in the rat adrenal glands as a selective ETA-receptor agonist, mainly involved in the stimulation of ZG proliferative activity.

Effects of 4-aminopyrazolo (3,4-d) pyrimidine on rat hepatocytes: an ultrastructural morphometric study.

The prolonged administration of the hypolipidemic drug 4-aminopyrazolo (3,4-d) pyrimidine (4APP) induced conspicuous morphological changes in rat hepatocytes, which are clearly demonstrated by stereology. We observed a significant decrease in the rough endoplasmic reticulum, and an accumulation of lipid droplets, which was coupled with a comparable rise in the hepatic concentrations of total cholesterol and triglycerides. These changes were interpreted as the expression of the 4APP-provoked impairment of the synthesis of the polypeptide chains of lipoproteins and of the consequent suppression of the assembly of lipid molecules in exportable lipoproteins. We also noted a neto lowering in smooth endoplasmic reticulum and peroxisomes, a decrease which was hypothesized to be the morphological counterpart of a reduced de novo cholesterol synthesis, due to the 4APP-induced rise in the intracellular cholesterol concentration.

Prolonged kallikrein inhibition does not affect the basal growth and secretory capacity of rat adrenal cortex, but enhances mineralo- and glucocorticoid response to ACTH and handling stress.

The effects on the pituitary-adrenocortical functions of the prolonged (7-day) blockade of endogenous bradykinin (BK) synthesis, obtained by the administration of the kallikrein inhibitor (K-I) cyclohexylacetyl-Phe-Arg-Ser-Val-Gln amide, were investigated in the rat. K-I treatment did not cause significant changes in the (i) body and adrenal weights; (ii) basal plasma levels of ACTH, aldosterone and corticosterone; and (iii) average volume of adrenocortical cells and their basal secretory capacity. Conversely, K-I administration induced a significant magnification of the in vivo mineralo- and glucocorticoid responses to the intraperitoneal (i.p.) bolus injection of ACTH. Moreover, K-I-treated rats, but not control ones, displayed a moderate and short-term adrenal secretory response to the mild stress evoked by the placebo i.p. injection. Collectively, these findings rule out the possibility that endogenous BK plays a relevant role in the control of adrenocortical function under basal conditions. However, they suggest that endogenous BK may be involved in quenching exceedingly high adrenocortical responses to ACTH and stresses.

Morphology and functional responses of isolated inner adrenocortical cells of rats infused with interleukin-beta.

The effects of the prolonged infusion with interleukin-1 beta (IL-1 beta) (20 pM.kg-1.min-1) on the function and morphology of the isolated inner cells of the rat adrenal cortex were investigated. After 3 and 5 days of IL-1 beta infusion, the level of circulating ACTH was below the control level, while the plasma concentration of corticosterone was strikingly elevated. After 5 days of infusion, isolated inner adrenocortical cells showed an enhanced basal and ACTH-stimulated corticosterone secretion, and showed a conspicuous hypertrophy. The acute exposure to IL-1 beta 10(-6) M did not affect the secretory activity of dispersed cell from either control or IL-1 beta-infused rats. These findings indicate that the prolonged exposure to high levels of circulating IL-1 beta, like those occurring during chronic inflammatory diseases, is able to enhance the growth and steroidogenic (glucocorticoid) capacity of the rat inner adrenocortical zones. Moreover, they suggest that the mechanism underlying this adrenocorticotrophic effect of IL-1 beta does not involve either a stimulation of the hypophyseal ACTH release or a direct stimulatory effect of monokine on adrenocortical cells. It is suggested that IL-1 beta may activate an intra-adrenal paracrine regulatory mechanism.

Corticotropin-inhibiting peptide enhances aldosterone secretion by dispersed rat zona glomerulosa cells.

Corticotropin-inhibiting peptide (CIP), the 7-38 fragment of human ACTH(1-39), is known to act as an antagonist of ACTH receptors. Accordingly, CIP has been found to inhibit ACTH-stimulated glucocorticoid secretion of dispersed rat adrenocortical cells, without per se affecting the basal production. We confirmed these findings, but unexpectedly observed that CIP concentration-dependently raised basal aldosterone secretion from fresh suspensions of rat zona glomerulosa (ZG) cells, maximal effective concentration being 10(-6) M. CIP (10(-6) M) partially reversed the ZG-cell response to ACTH, but not to the Ca2+-dependent agonists angiotensin-II (ANG-II) and K+. The aspecific ANG-II-receptor antagonist saralasin (10(-6) M) blocked the aldosterone response of ZG cells to 10(-6) M CIP, and in the presence of the Ca2+-channel blocker verapamil CIP was ineffective. Collectively, these findings suggest that CIP enhances aldosterone secretion of rat ZG through a mechanism involving the activation of ANG-II receptors and the consequent rise in the cytosolic Ca2+ concentration. They also stress that this side-effect of CIP must be taken into account in interpreting the results of investigations on the adrenal cortex, where CIP has been employed as an ACTH-receptor antagonist.

Effects of prolonged cysteamine administration on the rat adrenal cortex: evidence that endogenous somatostatin is involved in the control of the growth and steroidogenic capacity of zona glomerulosa.

A week daily administration of cysteamine (CYS, 300 mg kg-1) lowered plasma aldosterone concentration in rats, without affecting PRA, kalaemia and the plasma levels of ACTH and corticosterone. Prolonged CYS treatment caused a notable hypertrophy of adrenal zona glomerulosa (ZG) and its parenchymal cells, without inducing any apparent change in zona fasciculata morphology. Isolated ZG cells from CYS-treated rats evidenced a notable enhancement in their basal and maximally-stimulated productions of aldosterone and corticosterone. All these effects of chronic CYS administration were completely reversed by the simultaneous infusion of rats with somatostatin (SRIF, 12 micrograms kg-1 h-1). CYS exposure was not found to directly affect the secretory activity of isolated ZG cells from normal rats. Since CYS is known to be a specific depletor of SRIF in different organs of rats, these findings suggest that endogenous SRIF may be involved in the modulation of ZG function.

Evidence that endogenous vasoactive intestinal peptide (VIP) plays a role in the maintenance of the growth and steroidogenic capacity of rat adrenal zona glomerulosa.

The effects of a 7-day intraperitoneal infusion with VIP (0.03 nmol.kg-1.min-1) and its antagonist [4-Cl-D-Phe6,Leu17]-VIP (VIP-A; 3 nmol.kg-1.min-1) were studied in sham and bilaterally adrenalectomized rats bearing ACTH and angiotensin II (ANG-II)-responsive adrenocortical autotransplants. VIP significantly increased plasma aldosterone (ALDO) concentration (PAC) and lowered plasma renin activity (PRA) in both groups of animals, without affecting plasma levels of ACTH and corticosterone. This treatment caused a marked hypertrophy of adrenal zona glomerulosa (ZG) and its parenchymal cells (without inducing any significant change in the zona-fasciculata morphology), as well as of ZG-like cells of autotransplants. Isolated ZG cells and autotransplant quarters obtained from VIP-infused rats evidenced a notable increase in both their basal and maximally ACTH- or ANG-II-stimulated ALDO secretion. The simultaneous infusion of rats with VIP-A completely reversed all these effects of VIP. The infusion with VIP-A alone caused, in sham-operated rats, a net decrease in PAC, coupled with a rise in PRA, and a marked atrophy of ZG and ZG cells; basal and maximally stimulated ALDO secretion of dispersed ZG cells was also significantly lowered. Conversely, VIP-A did not evoke any appreciable effect in autotransplanted rats. These findings suggest that endogenous VIP is specifically involved in the maintenance of the growth and secretory capacity of rat adrenal ZG. Since regenerated adrenocortical autotransplants, which are responsive to VIP but not to VIP-A infusion, are completely deprived of chromaffin cells, the hypothesis is advanced that adrenal medulla may be the source of endogenous VIP regulating ZG function.

Morphology and function of the adrenal zona glomerulosa of transgenic rats TGR [mREN2] 27: effects of prolonged sodium restriction.

Heterozygous female transgenic rats for the murine Ren-2 gene (TGR) display a high blood pressure, together with a low kidney and high adrenal renin content. The effects of prolonged sodium restriction on the morphology and secretory activity of adrenal zona glomerulosa (ZG) of TGR and their age- and sex-matched Sprague-Dawley control rats (SDR) were investigated. Under basal conditions, TGR had a moderately hypertrophic ZG, that showed a significantly higher secretion of 18-hydroxylated (18OH) steroids: 18-hydroxy-11-deoxycorticosterone (18OH-DOC), 18-hydroxycorticosterone (18OH-B) and aldosterone (ALDO); ZG cells of TGR showed angiotensin II (AII)-binding site concentrations and ALDO secretory responses to AII similar to those of SDR ZG cells. Prolonged sodium restriction increased plasma ALDO level in both SDR and TGR, and significantly raised the volume of ZG. ZG hypertrophy was due to the increase in both the number and average volume of its parenchymal cells. The secretion of 18OH-steroids was markedly enhanced in both groups of rats; however, in TGR this rise was exclusively due to increases of 18OH-DOC and 18OH-B, while in SDR also ALDO production was enhanced. The yield of non-18OH-steroids was not affected. 11-Dehydrocorticosterone production was not changed in SDR, but doubled in TGR. ZG cells of sodium-restricted SDR and TGR displayed similar increases in their AII-binding site concentration and ALDO secretory response to AII. In conclusion, our present findings confirm that TGR possess a hypertrophic ZG and an elevated secretory capacity o 18OH-steroids, but show only slight differences in ZG and ZG-cell responses to prolonged sodium deprivation.

Evidence that endogenous somatostatin (SRIF) exerts an inhibitory control on the function and growth of rat adrenal zona glomerulosa. The possible involvement of zona medullaris as a source of endogenous SRIF.

The effect of SRIF and its antagonist cyclo(7-aminoheptanonyl-Phe-D-Trp-Lys-Thr magnitude of Bzl)(SRIF-A) were studied in sham-operated and bilaterally adrenalectomized rats bearing ACTH- and angiotensin II (ANG-II)-responsive adrenocortical autotransplants. SRIF-A (10(-5) M) completely annulled SRIF (10(-6) M)-induced inhibition of ANG-II (10(-8) M)-evoked rise in aldosterone (ALDO) secretion by both dispersed zona glomerulosa (ZG) cells and autotransplant slices. A 7-day intraperitoneal infusion with SRIF (0.3 nmol.kg-1.min-1) significantly lowered plasma ALDO concentration (PAC) in both groups of animals, without affecting plasma renin activity and the plasma levels of ACTH and corticosterone. This treatment caused a marked atrophy of adrenal ZG and its parenchymal cells (without inducing any significant change in the zona fasciculata morphology), as well as of ZG-like cells of autotransplants. Isolated ZG cells and autotransplant slices from SRIF-infused rats evidenced a notable decrease in both their basal and maximally ACTH- or ANG-II-stimulated ALDO production. The simultaneous infusion of rats with SRIF-A (3 nmol.kg-1.min-1) completely reversed all these effects of SRIF. The prolonged infusion with SRIF-A alone caused, in sham-operated rats, a marked increase in PAC and a significant hypertrophy of ZG and ZG cells; basal and maximally-stimulated ALDO secretion of dispersed ZG cells was also notably raised. Conversely, SRIF-A infusion did not evoke any appreciable effect in autotransplanted rats. These findings suggest that endogenous SRIF is specifically involved in the negative control of the secretion and growth of the rat adrenal ZG. Since regenerated adrenocortical autotransplants, which are responsive to SRIF but not to SRIF-A infusion, are completely deprived of chromaffin cells, the hypothesis is advanced that adrenal zona medullaris may be the source of endogenous SRIF regulating ZG function.

Stimulation of endogenous nitric oxide production is involved in the inhibitory effect of adrenomedullin on aldosterone secretion in the rat.

Adrenomedullin (AM) (10(-8) M) partially suppressed aldosterone response of dispersed rat zona glomerulosa (ZG) cells to 10 mM K+, and the nitric oxide (NO) synthase inhibitors L-NAME (10(-3) M) and 1400W (10(-4) M) effectively counteracted this effect of AM. The NO donor L-Arginine (L-Arg) (10(-5) M) decreased both basal and K+ -stimulated aldosterone secretion. The guanylate-cyclase inhibitor Ly-83583, at a concentration (10(-4) M) abolishing either the guanylate-cyclase activator guanylin- or L-Arg-induced cGMP release from dispersed ZG cells, did not affect the aldosterone antisecretagogue action of AM and L-Arg. AM (10(-8) M) evoked a moderate increase in cGMP release by dispersed ZG cells, and the effect was blocked by both 10(-4) M Ly-83583 and 10(-3) M L-NAME. Collectively, these findings allow us (1) to confirm that NO inhibits aldosterone secretion through a cGMP-independent mechanism; and (2) to suggest that stimulation of endogenous NO synthesis plays a role in the mechanisms underlying the inhibitory effect of AM on K+ -stimulated aldosterone secretion from rat ZG cells.

Arginine-vasopressin stimulates CRH and ACTH release by rat adrenal medulla, acting via the V1 receptor subtype and a protein kinase C-dependent pathway.

Arginine-vasopressin (AVP) is a hypothalamic hormone that, like CRH, stimulates the pituitary release of ACTH, thereby activating adrenal glucocorticoid secretion. Evidence indicates that rat adrenal medulla contains a CRH-ACTH system duplicating that existing at the hypothalamo-pituitary level and involved in the paracrine stimulation of the cortex secretion. Therefore, we investigated by RIA the effect of AVP on the release of CRH and ACTH immunoreactivities (IR) by rat adrenal medulla in vitro. AVP concentration-dependently enhanced the release of both CRH-IR and ACTH-IR, and the effect was blocked by a selective antagonist of the V1 subtype of AVP receptors. The CRH receptor antagonist alpha-helical-CRH partially reversed AVP-evoked rise in ACTH-IR release, without altering either CRH response or basal secretions of CRH and ACTH. The specific inhibitors of protein kinase C Ro31-8220 and calphostin C abolished both CRH and ACTH responses to AVP. In conclusion, our present findings suggest that AVP stimulates intramedullary the CRH-ACTH system, acting via V1 receptors and activating protein kinase C.

Effects of prolonged infusion with endothelin-1 on the function and morphology of rat adrenal cortex.

A week of SC infusion with endothelin-1 (ET-1) (0.2 microgram.kg-1.hr-1) lowered PRA and raised plasma aldosterone (A) concentration in rats. Kalaemia and the plasma levels of ACTH and corticosterone (B) were not affected. Prolonged ET-1 administration caused a notable hypertrophy of zona glomerulosa (ZG) and its parenchymal cells, without inducing any apparent change in zona fasciculata. Stereology showed that ZG cell hypertrophy was mainly due to the increase in the volume of the mitochondrial compartment and to the proliferation of smooth endoplasmic reticulum (i.e., the two organelles in which the enzymes of steroid synthesis are contained). Isolated ZG cells from ET-1-infused animals evidenced a notable enhancement in their basal production of A and B. The secretory responses of ZG cells to the maximal effective concentrations of their three main stimulators (ACTH, angiotensin-II and K+) displayed comparable increases. These findings indicate that ET-1, when chronically administered, is able to specifically enhance the growth and steroidogenic capacity of rat ZG, and suggest that the mechanism underlying this ET-1 effect involves stimulation of the de novo synthesis of both the steroidogenic enzymes and the membrane framework in which they are located.