Activation of distinct cAMP-dependent and cGMP-dependent pathways by nitric oxide in cardiac myocytes

Nitric oxide (NO) donors were recently shown to produce biphasic contractil e effects in cardiac tissue, with augmentation at low NO levels and depress ion at high NO levels. We examined the subcellular mechanisms involved in t he opposing effects of NO on cardiac contraction and investigated whether N O-modulates contraction exclusively via guanylyl cyclase (GC) activation or whether some contribution occurs via cGMP/PKG-independent mechanisms, in i ndo 1-loaded adult cardiac myocytes. Whereas a high concentration of the NO donor S-nitroso-N-acetylpenicillamine (SNAP, 100 mu mol/L) significantly a ttenuated contraction amplitude by 24.4+/-4.5% (without changing the Ca2+ t ransient or total cAMP), a low concentration of SNAP (1 mu mol/L) significa ntly increased contraction amplitude (38 +/- 10%), Ca2+ transient (26+/-10% ), and cAMP levels (from 6.2 to 8.5 pmol/mg of protein). The negative contr actile response of 100 mu mol/L SNAP was completely abolished in the presen ce of the specific blocker of PKG KT 5823 (1 mu mol/L); the positive contra ctile response of 1 mu mol/L SNAP persisted, despite the presence of the se lective inhibitor of GC 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ, 1 0 mu mol/L) alone, but was completely abolished in the presence of ODQ plus the specific inhibitory cAMP analog Rp-8-CPT-cAMPS (100 mu mol/L), as well as by the NO scavenger oxyhemoglobin. Parallel experiments in cell suspens ions showed significant increases in adenylyl cyclase (AC) activity at low concentrations (0.1 to 1 mu mol/L) of SNAP (AC, 18% to 20% above basal acti vity). We conclude that NO can regulate both AC and GC in cardiac myocytes. High levels of NO induce large increases in cGMP and a negative inotropic effect mediated by a PKG-dependent reduction in myofilament responsiveness to Ca2+. Low levels of NO increase cAMP, at least in part, by a novel cGMP- independent activation of AC and induce a positive contractile response.