Subsarcolemmal cAMP signals monitored by cyclic nucleotide gated channels in hypertrophied cardiac myocytes

Abstract

The second messenger cAMP is the most important modulator of sympathetic control over cardiac contractility. Recent studies have documented the existence of local pools of cAMP in cardiac cells (Jurevicius & Fischmeister 1996, PNAS 93:295-299; Zaccolo & Pozzan, 2002 Science 295:1711-15; Rochais et al. 2004 J Biol Chem. 279:52095- 105) thought to permit selective activation of cAMP targets. In cardiac hypertrophy and failure, alterations in some of the major components of the cAMP pathway occur, but whether these modifications affect cAMP compartmentation is unknown. In an attempt to address this question, 3 weeks-old rats were subjected to aortic coarctation (H) or sham operated (S). 5 weeks after surgery, the mass of the left ventricle to tibia length ratio was 59% increased in the H group compared to the S group while the right ventricle mass to tibia length ratio was unchanged, attesting compensated hypertrophy of the heart in H rats. Isolated left ventricular myocytes from both groups were infected with an adenovirus encoding a mutant of the olfactory cyclic nucleotide-gated (CNG) channels D subunit (E583M C460W CNGA2). This channel is gated by cAMP and thus allow to follow cAMP dynamics by recording the associated cationic current (ICNG) with the whole cell patch clamp technique 24 h after infection. Cumulative concentrationresponse curve to the non-selective E-adrenergic agonist isoprenaline (Iso) indicated a 33% decrease of the maximal cAMP accumulation produced by Iso while the effect of the hydrosoluble forskolin analog, L-858051 at a saturating concentration (100 µM) was unchanged between the two groups. These results show that CNG channels allow to measure cAMP variations at the membrane in living cardiomyocytes and to detect alterations of cAMP signals during early hypertrophy.