The effects of KN62, a Ca2+/calmodulin-dependent protein kinase II inhibitor, on adrenocortical cell aldosterone production.
The effects of KN62 on aldosterone secretion have been studied using an angiotensin II (AII)- and K(+)-responsive human adrenocortical tumor cell line (H295R). Basal aldosterone secretion (measured by RIA) was 0.57 +/- 0.22 pmol/mg protein.h. The physiologicial agonists AII (10 nM) and K+ (14 mM) increased aldosterone secretion by 6.9- and 5.0-fold, respectively. Aldosterone secretion was also stimulated by dibutyryl cyclic AMP (dbcAMP, 1 mM, 10.3-fold over basal). Nifedipine dose-dependently inhibited K(+)- and AII-stimulated aldosterone secretion. In contrast, dbcAMP-stimulated secretion was relatively insensitive to this agent (26.8% inhibition at 1 microM nifedipine). K(+)- and AII-stimulated aldosterone production was also dose-dependently inhibited by KN62, which produced 93.9% and 82.3% inhibition at 10 microM KN62 (both p < 0.01). In order to test the specificity of KN62 in H295R cells, its effects on various other steroidogenic agonists were assessed. KN62 dose-dependently inhibited aldosterone secretion stimulated by dbcAMP, 22-hydroxycholesterol and pregnenolone. In addition, KNO4, a derivative of KN62 which is not a potent inhibitor of CaM Kinase II, exhibited a similar pattern of inhibition. These data confirm the requirement for extracellular Ca2+ in the stimulation of human adrenocortical cell aldosterone secretion by AII and K+. However, the non-specific inhibitory effects of KN62 in H295R cells limit the usefulness of this agent as a tool for investigations of the involvement of CaM kinase II in adrenocortical steroidogenesis.
Clyne
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CD
Dept of Obstetrics & Gynecology, University of Texas Southwestern Medical Center, Dallas 75235, USA.
Nguyen
A
A
Rainey
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WE
D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine / Q000031:analogs & derivatives*; D000302:Adrenal Cortex / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D000450:Aldosterone / Q000096:biosynthesis* / Q000378:metabolism; D000804:Angiotensin II / Q000037:antagonists & inhibitors; D003994:Bucladesine / Q000037:antagonists & inhibitors; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000037:antagonists & inhibitors*; D006801:Humans; D007546:Isoquinolines / Q000494:pharmacology*; D009543:Nifedipine / Q000494:pharmacology; D010879:Piperazines / Q000494:pharmacology*; D011188:Potassium / Q000037:antagonists & inhibitors; D014407:Tumor Cells, Cultured
D007546:Isoquinolines; D010879:Piperazines; D000804:Angiotensin II; D000450:Aldosterone; C063302:KN 62; D003994:Bucladesine; D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D009543:Nifedipine; D011188:Potassium