Regulation of aldosterone production from zona glomerulosa cells by ANG II and cAMP: evidence for PKA-independent activation of CaMK by cAMP

2006 ◽  
Vol 290 (3) ◽  
pp. E423-E433 ◽  
Author(s):  
Stepan Gambaryan ◽  
Elke Butt ◽  
Piet Tas ◽  
Albert Smolenski ◽  
Bruno Allolio ◽  
...  
Keyword(s):  
Intracellular Signaling ◽  
Mapk Signaling ◽  
Zona Glomerulosa ◽  
Intracellular Camp ◽  
Ang Ii ◽  
Intact Cells ◽  
Molecular Events ◽  
Aldosterone Production ◽  
Dependent Protein

Aldosterone production in zona glomerulosa (ZG) cells of adrenal glands is regulated by various extracellular stimuli (K+, ANG II, ACTH) that all converge on two major intracellular signaling pathways: an increase in cAMP production and calcium (Ca2+) mobilization. However, molecular events downstream of the increase in intracellular cAMP and Ca2+ content are controversial and far from being completely resolved. Here, we found that Ca2+/calmodulin-dependent protein kinases (CaMKs) play a predominant role in the regulation of aldosterone production stimulated by ANG II, ACTH, and cAMP. The specific CaMK inhibitor KN93 strongly reduced ANG II-, ACTH-, and cAMP-stimulated aldosterone production. In in vitro kinase assays and intact cells, we could show that cAMP-induced activation of CaMK, using the adenylate cyclase activator forskolin or the cAMP-analog Sp-5,6-DCI-cBIMPS (cBIMPS), was not mediated by PKA. Activation of the recently identified cAMP target protein Epac (exchange protein directly activated by cAMP) by 8-pCPT-2′- O-Me-cAMP had no effect on CaMK activity and aldosterone production. Furthermore, we provide evidence that cAMP effects in ZG cells do not involve Ca2+ or MAPK signaling. Our results suggest that ZG cells, in addition to PKA and Epac/Rap proteins, contain other as yet unidentified cAMP mediator(s) involved in regulating CaMK activity and aldosterone secretion.

Angiotensin II early regulated genes in H295R human adrenocortical cells

2004 ◽  
Vol 19 (1) ◽  
pp. 106-116 ◽  
Author(s):  
Damian G. Romero ◽  
Maria Plonczynski ◽  
Gaston R. Vergara ◽  
Elise P. Gomez-Sanchez ◽  
Celso E. Gomez-Sanchez
Keyword(s):  
Gene Expression ◽  
Angiotensin Ii ◽  
Intracellular Signaling ◽  
Zona Glomerulosa ◽  
Ang Ii ◽  
Cellular Functions ◽  
Cellular Mechanisms ◽  
Mediated Effects ◽  
Aldosterone Production ◽  
H295r Cells

Evidence for the dysregulation of aldosterone synthesis in cardiovascular pathophysiology has renewed interest in the control of its production. Cellular mechanisms by which angiotensin II (ANG II) stimulates aldosterone synthesis in the adrenal zona glomerulosa are incompletely understood. To elucidate the mechanism of intracellular signaling by ANG II stimulation in the adrenal, we have studied immediate-early regulated genes in human adrenal H295R cells using cDNA microarrays. H295R cells were stimulated with ANG II for 3 h. Gene expression was analyzed by microarray technology and validated by real-time RT-PCR. Eleven genes were found to be upregulated by ANG II. These encode the proteins for ferredoxin, Nor1, Nurr1, c6orf37, CAT-1, A20, MBLL, M-Ras, RhoB, GADD45α, and a novel protein designated FLJ45273 . Maximum expression levels for all genes occurred 3–6 h after ANG II stimulation. This increase was dose dependent and preceded maximal aldosterone production. Other aldosterone secretagogues, K+and endothelin-1 (ET-1), also induced the expression of these genes with variable efficiency depending on the gene and with lower potency than ANG II. ACTH had negligible effect on gene expression except for the CAT-1 and Nurr1 genes. These ANG II-stimulated genes are involved in several cellular functions and are good candidate effectors and regulators of ANG II-mediated effects in adrenal zona glomerulosa.

Effect of CO2/pH on the aldosterone response to hypoxia in bovine adrenal cells in vitro

1993 ◽  
Vol 265 (4) ◽  
pp. R820-R825
Author(s):  
H. Raff ◽  
B. Jankowski
Keyword(s):  
Respiratory Acidosis ◽  
Inhibitory Effect ◽  
Zona Glomerulosa ◽  
Ang Ii ◽  
Adrenal Cells ◽  
Aldosterone Production ◽  
Per Se ◽  
Air Control

Acidosis increases and hypoxia decreases aldosterone production from the adrenal zona glomulerosa in vivo, in situ, and in vitro. These effects appear to be located at different steps in the steroidogenic process. Because respiratory acidosis and hypoxemia are common sequelae of chronic lung disease, the present experiments evaluated the interaction of hypoxia and CO2 (with uncompensated or compensated extracellular pH) on aldosteronogenesis in vitro. Bovine adrenal zona glomerulosa cells were stimulated with angiotensin II (ANG II) or adenosine 3',5'-cyclic monophosphate under room air control (21% O2-0% CO2), CO2 per se (21% O2-10% CO2), hypoxia per se (10% O2-0% CO2), and the combination of CO2 and hypoxia (10% O2-10% CO2). Furthermore, under CO2, pH was either allowed to decrease from 7.2 to 6.8 (uncompensated) or its decrease was minimized (> 7.05) with NaOH (compensated). CO2 without pH compensation led to a significant increase in ANG II-stimulated aldosterone release; when the decrease in pH was minimized, CO2 inhibited ANG II-stimulated aldosterone release. Hypoxia inhibited aldosterone release; the inhibitory effect of hypoxia predominated when combined with CO2. In the presence of cyanoketone, pregnenolone production from endogenous precursors (early pathway) was unaffected. However, the conversion of corticosterone to aldosterone (late pathway) was inhibited by low O2 but unaffected by CO2. It is concluded that the inhibitory effect of low O2 on the late pathway predominates over the effects of uncompensated or compensated simulated respiratory acidosis on aldosteronogenesis.

scholarly journals Renin knockout rat: control of adrenal aldosterone and corticosterone synthesis in vitro and adrenal gene expression

2015 ◽  
Vol 308 (1) ◽  
pp. R73-R77 ◽  
Author(s):  
Hershel Raff ◽  
Ashley Gehrand ◽  
Eric D. Bruder ◽  
Matthew J. Hoffman ◽  
William C. Engeland ◽  
...  
Keyword(s):  
Renin Angiotensin System ◽  
Plasma Renin ◽  
Immunohistochemical Analysis ◽  
Zona Glomerulosa ◽  
Ang Ii ◽  
Angiotensin System ◽  
Renin Angiotensin ◽  
Aldosterone Production ◽  
Renin Mrna

The classic renin-angiotensin system is partly responsible for controlling aldosterone secretion from the adrenal cortex via the peptide angiotensin II (ANG II). In addition, there is a local adrenocortical renin-angiotensin system that may be involved in the control of aldosterone synthesis in the zona glomerulosa (ZG). To characterize the long-term control of adrenal steroidogenesis, we utilized adrenal glands from renin knockout (KO) rats and compared steroidogenesis in vitro and steroidogenic enzyme expression to wild-type (WT) controls (Dahl S rat). Adrenal capsules (ZG; aldosterone production) and subcapsules [zona reticularis/fasciculata (ZFR); corticosterone production] were separately dispersed and studied in vitro. Plasma renin activity and ANG II concentrations were extremely low in the KO rats. Basal and cAMP-stimulated aldosterone production was significantly reduced in renin KO ZG cells, whereas corticosterone production was not different between WT and KO ZFR cells. As expected, adrenal renin mRNA expression was lower in the renin KO compared with the WT rat. Real-time PCR and immunohistochemical analysis showed a significant decrease in P450aldo ( Cyp11b2) mRNA and protein expression in the ZG from the renin KO rat. The reduction in aldosterone synthesis in the ZG of the renin KO adrenal seems to be accounted for by a specific decrease in P450aldo and may be due to the absence of chronic stimulation of the ZG by circulating ANG II or to a reduction in locally released ANG II within the adrenal gland.

Abstract 423: Angiotensin II Type 1 Receptor (AT1R) Activating Autoantibodies Contribute to the Pathophysiology of Primary Hyperaldosteronism

Hypertension ◽  
2013 ◽  
Vol 62 (suppl_1) ◽  
Author(s):  
David C Kem ◽  
Hongliang Li ◽  
Xichun Yu ◽  
Carolina Velarde-Miranda ◽  
Celso Gomez-Sanchez
Keyword(s):  
Cho Cells ◽  
Rabbit Serum ◽  
Ang Ii ◽  
Therapeutic Implications ◽  
Aldosterone Production ◽  
Target Epitope ◽  
Activating Autoantibodies ◽  
Normal Controls

Objectives: Activating autoantibodies (AA) to the AT1R second extracellular loop (ECL2) are present in patients with primary aldosteronism (PA). We investigated whether AA from PA and from AT1R-ECL2 immunized rabbits activate AT1R and thereby contribute to hypertension by a direct contractile effect on the vasculature and stimulate adrenal aldosterone production. Methods: We studied 5 normal controls (NC) and 15 subjects with biochemically confirmed PA [4 APA, 4 IAH non- and 7 indeterminate (IPA)]. AT1R-AA activity in serum or IgG was analyzed in AT1R transfected CHO cells. Contractile effects were assayed in isolated perfused rat cremaster arteries. We determined the effects of an ARB and the L-aa AT1R-ECL2 target epitope on in vitro AT1R cremaster artery responsiveness to patient sera and to MAP-L-aa (AFHYESQ) peptide immunized rabbit serum. We examined the effect of IgG from 7 PA subjects on aldosterone production in HAC15 cells in vitro w/wo 1.0 and 10 nM Ang II; and w/wo ARB. Results: Serum from PA significantly (p<0.05) increased Ang II-equivalent activity in AT1R-CHO cells in IAH 199± x pM, n=4; APA 177±y pM, n=3; IPA 187±z pM, n=7; vs NC 83±a pM, n=5 and was blocked by 10 μM losartan. IgG from PA decreased perfused rat arteriole diameter in IAH by 22±8(SD)% of a 100% (normalized) baseline, n=4; APA 14±0.8%, n=3; IPA 21±6%, n=7; and NC 7±3.5%, n=5 and was blocked by 10 μM losartan. IgG from 7 PA subjects increased HAC15 aldosterone (aldo) 1.8 fold over baseline (p<0.01) and markedly increased 1nM and 10 nM Ang II-induced aldo by 3.5 fold (p<0.01); and was blocked by ARB. BP in hi Na rabbits rose from 80/60 to 140/90 6 weeks after immunization with AT1R MAP-L-aa AFHYESQ peptide. Arteriole contraction rose from 5% (preimmune) to 30% (post-immunization). Preincubation with the L-aa 2 nd ECL target reduced contractility to 8%; not different from baseline. Conclusions: AA from PA and AT1R-immunized rabbits activates AT1R transfected CHO cells, increases in vitro arterial contractility and stimulates basal and Ang II induced HAC15 aldosterone secretion. ARBs and decoy peptides were effective in blocking AA effects in vitro . These AA have important etiological and therapeutic implications.

Abstract 128: Leptin: A Regulator of Aldosterone Synthase Expression & Aldosterone Secretion in Visceral Adipocytes, in Mice

Hypertension ◽  
2016 ◽  
Vol 68 (suppl_1) ◽  
Author(s):  
Eric J Belin de Chantemele ◽  
Anne-Cecile Huby ◽  
P. T Menk ◽  
Weiqin Chen ◽  
Brian Lane ◽  
...  
Keyword(s):  
Adipose Tissue ◽  
Cross Sections ◽  
Leptin Receptor ◽  
Zona Glomerulosa ◽  
Aldosterone Synthase ◽  
Leptin Receptors ◽  
Fat Depots ◽  
Physiological Relevance ◽  
Aldosterone Production

Obesity is associated with inappropriately high aldosterone levels, which contribute to the development of metabolic and cardiovascular disorders. The origin of these high aldosterone levels is incompletely understood. We recently demonstrated that the adipocyte-derived hormone leptin regulates aldosterone synthase (CYP11B2) expression and stimulates aldosterone release from adrenal zona glomerulosa cells. Recent studies demonstrate that adipocytes express CYP11B2 and secrete aldosterone. However, the mechanisms regulating aldosterone release from adipocytes remain unclear. Likewise, whether visceral (Visc) and subcutaneous (SubQ) adipose tissue contribute to a similar extent to aldosterone production is unknown. We tested the hypothesis that leptin increases adipocyte CYP11B2 expression and aldosterone production and investigated whether Visc and SubQ adipose tissues respond similarly to leptin. Immunostaining of mouse adipose tissue cross-sections and isolated mature adipocytes revealed that Visc and SubQ adipose tissue express leptin receptors. Treatment of mouse freshly isolated mature adipocytes, non-differenciated (stromal fraction) and differentiated adipocytes revealed that leptin dose-dependently increased CYP11B2 expression and aldosterone production in Visc adipose tissue only. Although leptin receptor and CYP11B2 levels were similar in SubQ and Visc adipocytes, SubQ adipocytes were unresponsive to leptin. The physiological relevance of these in vitro data was tested by measuring plasma aldosterone levels in mice deprived of adipose tissue (lipodystrophic mice) treated with leptin. Absence of adipose tissue in lipodystrophic mice blunted leptin-induced increases in aldosterone levels (WT-vehicle: 471±82 vs. WT-Leptin: 1699±396, p<0.05; KO-vehicle: 539±71 vs. KO+leptin: 787±156, NS). The human relevance of these data was determined by reporting that CYP11B2 expression gradually increased with body mass index in human mediastinal and omental fat depots. In summary these data strongly suggest that leptin regulates CYP11B2 levels and aldosterone release in visceral adipose tissue and that leptin-induced, adipocyte-derived aldosterone may contribute to obesity-associated hyperaldosteronism.

Inhibition of aldosterone release by hypoxia in vitro: interaction with carbon monoxide

1994 ◽  
Vol 76 (2) ◽  
pp. 689-693 ◽  
Author(s):  
H. Raff ◽  
B. Jankowski
Keyword(s):  
Carbon Monoxide ◽  
Angiotensin Ii ◽  
Zona Glomerulosa ◽  
Michaelis Constant ◽  
Aldosterone Synthase ◽  
Aldosterone Production ◽  
Glomerulosa Cells ◽  
In Vitro Interaction ◽  
Zona Glomerulosa Cells

We have demonstrated that the aldosteronogenic pathway of the zona glomerulosa is unusually sensitive to modest changes in PO2 (Michaelis constant for O2 approximately 95 Torr). The current study evaluated the interaction of CO (the classic ligand for P-450 enzymes) and the decreases in O2 on aldosteronogenesis in vitro. Bovine adrenocortical zona glomerulosa cells were incubated for 2 h and stimulated with either adenosine 3′,5′-cyclic monophosphate (cAMP) or angiotensin II. Ten and 20% CO led to significant decreases in cAMP- and angiotensin II-stimulated aldosteronogenesis. The combination of 20% CO and moderate decreases in PO2 (from approximately 140 to approximately 100 Torr) led to an interactive decrease in aldosterone production. The conversion of corticosterone to aldosterone catalyzed by aldosterone synthase, which is the site of O2 sensitivity, was not significantly inhibited by CO. We conclude that the aldosterone pathway is not exceptionally sensitive to CO compared with other steroidogenic pathways. This observation suggests that the unique O2-sensitive properties of the aldosterone pathway located primarily within aldosterone synthase may not reside in its CO binding site (i.e., heme).

scholarly journals Regulation of Microtubule Dynamics by Extracellular Signals: cAMP-dependent Protein Kinase Switches Off the Activity of Oncoprotein 18 in Intact Cells

1998 ◽  
Vol 140 (1) ◽  
pp. 131-141 ◽  
Author(s):  
Helena Melander Gradin ◽  
Niklas Larsson ◽  
Ulrica Marklund ◽  
Martin Gullberg
Keyword(s):  
Protein Kinase ◽  
Genetic System ◽  
Cell Surface Receptor ◽  
In Vitro Assays ◽  
Dependent Protein Kinase ◽  
Intact Cells ◽  
Camp Dependent Protein Kinase ◽  
Oncoprotein 18 ◽  
Dependent Protein

Oncoprotein 18 (Op18, also termed p19, 19K, metablastin, stathmin, and prosolin) is a recently identified regulator of microtubule (MT) dynamics. Op18 is a target for both cell cycle and cell surface receptor-coupled kinase systems, and phosphorylation of Op18 on specific combinations of sites has been shown to switch off its MT-destabilizing activity. Here we show that induced expression of the catalytic subunit of cAMP-dependent protein kinase (PKA) results in a dramatic increase in cellular MT polymer content concomitant with phosphorylation and partial degradation of Op18. That PKA may regulate the MT system by downregulation of Op18 activity was evaluated by a genetic system allowing conditional co-expression of PKA and a series of kinase target site–deficient mutants of Op18. The results show that phosphorylation of Op18 on two specific sites, Ser-16 and Ser-63, is necessary and sufficient for PKA to switch off Op18 activity in intact cells. The regulatory importance of dual phosphorylation on Ser-16 and Ser-63 of Op18 was reproduced by in vitro assays. These results suggest a simple model where PKA phosphorylation downregulates the MT-destabilizing activity of Op18, which in turn promotes increased tubulin polymerization. Hence, the present study shows that Op18 has the potential to regulate the MT system in response to external signals such as cAMP-linked agonists.

scholarly journals Oxygen-dependence of ACTH-stimulated aldosterone and corticosterone synthesis in the rat adrenal cortex: developmental aspects

2002 ◽  
Vol 172 (3) ◽  
pp. 595-604 ◽  
Author(s):  
ED Bruder ◽  
AK Nagler ◽  
H Raff
Keyword(s):  
Enzyme Activity ◽  
Age Groups ◽  
Maximal Activity ◽  
Zona Glomerulosa ◽  
Zona Fasciculata ◽  
Steroid Synthesis ◽  
Camp Generation ◽  
Aldosterone Production ◽  
Health And Disease

The control of ACTH-stimulated steroidogenesis under decreasing levels of O(2) is not fully understood. The purpose of this study was to examine the effects of decreased O(2) in vitro on rat adrenocortical steroid synthesis at different stages of development. Of interest was the evaluation of the effect of low O(2) on steroidogenesis during the stress hyporesponsive period of the neonate. Rats were killed at 7, 14, or 42 days of age, adrenals collected and capsules (zona glomerulosa, ZG) separated from subcapsules (zona fasciculata/reticularis, ZFR). Cells were dispersed and placed into glass vials each gassed with a different level of O(2) (21, 5, 2, 1, or 0% O(2)). The entire steroidogenic pathway was analyzed by measuring ACTH-stimulated cAMP, corticosterone and aldosterone production during a 2 h incubation. In addition, the early (P450 scc) and late (P450c11 beta and P450 aldo) pathway activities were examined in the presence of cyanoketone. The PO(2) for half-maximal activity (P(50)) for aldosterone synthesis in ZG cells from 7- and 42-day-old rats was approximately 28 mmHg and 7 mmHg respectively, indicating that cells from older rats were more resistant to inhibition by low O(2). The P(50) for cAMP production from the ZG was approximately 14 mmHg for both age groups. The P(50) for corticosterone synthesis was approximately 28 mmHg and <7 mmHg in ZFR cells from 7- and 42-day-old cells respectively. The only enzyme activities affected by low O(2) (<35 mmHg) were P450 aldo and P450 scc. Moderate decreases in O(2) (from approximately 150 mmHg) decreased aldosteronogenesis, possibly due to observed decreases in cAMP generation, but not due to decreases in steroidogenic enzyme activity (7-day-old). Severe decreases in O(2) presumably inhibited P450 aldo through a direct effect on enzyme activity (both ages). P450 scc activity (including cholesterol transport) also seems to be decreased by very low O(2) (7-day-old). These findings illustrate a novel developmental alteration in O(2)-regulated steroid production, and may have implications for neonatal health and disease.

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