Angiotensin II and cAMP regulate AT1-mRNA expression in rat cardiomyocytes by transcriptional mechanism

Mechanisms of angiotensin II and cAMP regulating the expression of angiotensin II type 1 (AT(1)) receptor mRNA were studied in neonatal rat cardiomyocytes. Angiotensin II induced a transient decrease of AT(1)-mRNA expression in time- and dose-dependent manner. Maximal decrease (49.2 +/- 9.5% of control) occurred at 6 h of angiotensin II (10 nmol/l) treatment. AT(1) receptor antagonists 4-ethyl-2-n-propyl-1-[2'-(1H-tetrazol-5-yl)biphenyl-4-yl)methyl]imidazole-5-carboxylic acid (DMP811) and losartan as well as 1-(5-isoquinolinesulfonyl)-2-methylpiperazine dihydrochloride (H-7) reversed the down-regulation of AT(1)-mRNA expression. 6 h of phorbol 12-myristate 13-acetate (PMA) stimulation caused a decrease of AT(1)-mRNA level. Treatment by angiotensin II plus actinomycin D for 6 h produced the same effect as actinomycin D alone. These results suggest that angiotensin II down-regulates AT(1)-mRNA level of rat cardiomyocytes by inhibiting the transcription of AT(1) gene, which is mediated by AT(1) receptor and related to the activation of protein kinase C. Stimulation by forskolin plus 3-isobutyl-1-methyl-xanthine (IBMX) decreased the expression of AT(1)-mRNA to 68.1 +/- 21.5% of control at 6 h treatment; while increased to 207.9 +/- 27.1% of control at 48 h treatment. A series of 5'-upstream deletion mutants of AT(1A) promoter were produced and then were recombined with pGL(3) basic vector utilizing luciferase as reporter gene. Among all the constructors, p(-201/+ 74)Luc was of the highest luciferase activity (5.9 times higher than control) after stimulation by forskolin for 48 h. Further deletion from -201 to -61 resulted in a large decrease of activity. These results indicate that cAMP induces a time-dependent bi-directional regulation of AT(1)-mRNA expression. The cAMP responsible element (CRE) cis-element located in the region -201/-61 of rat AT(1A) promoter is forskolin inducible, which may mediate the up-regulation of AT(1)-mRNA expression induced by cAMP long-lasting stimulation.