Mechanism for muscarinic inhibition of ICa(L) is determined by the path for elevating cyclic AMP in cardiac myocytes

Objective: Does carbachol (CCh) require NO/cGMP for inhibition of L-type calcium current (I-Ca(L)) when either adenylyl cyclase activation or phosphodiesterase suppression is used to raise cAMP? Methods: The effects of the NO donor SIN-1 (3-morpholinosydnonimine), CCh and atrial natriuretic peptide (ANP) were evaluated when I-Ca(L) had been stimulated by isoproterenol (ISO) or 3-isobutyl-1-methylxanthine (IBMX) in guinea pig isolated ventricular myocytes (35 degreesC). Results: Carbachol, SIN-I or ANP did not affect basal I-Ca(L): each inhibited IBMX-stimulated I-Ca(L). Dialyzed (30-100 muM) ODQ (1H-(1,2,4)oxadiazolo(4,3-a)quinoxalin-1-one), a soluble guanylyl cyclase (sGC) inactivator, blocked inhibition of IBMX-stimulated I-Ca(L) by SIN-1 (10 muM) but not by CCh (1-100 muM) or ANP (100 nM). Dialysis with 3 muM LY83583 (6-anilino-5,8-quinolinedione), a particulate (pGC) and sGC inactivator, opposed muscarinic-, ANP- and SIN-1-induced inhibition of IBMX-stimulated I-Ca(L). Thus CCh can increase cGMP synthesis via pGC. Even with 100 muM [LY83583](pip), CCh inhibited ISO-stimulated I-Ca(L) an effect referable to suppression of adenylyl cyclase activity. However, 3 muM [LY83583](pip) prevented inhibition of ISO-stimulated I-Ca(L) by ANP. [LY83583](pip) did not affect inhibition by 8 bromo-cGMP (100 muM) of ISO- or IBMX-stimulated I-Ca(L). The observations indicate that: (1) myocytes have ODQ-sensitive sGC activated by NO and LY8353-sensitive pGC activated by ANP, (2) CCh does not inhibit I-Ca(L) via NO, (3) the mechanism for muscarinic inhibition depends upon the cAMP-elevating agent and (4) LY83583 distinguishes between two pathways for muscarinic inhibition. Conclusion: The nature of the stimulant pathway that increases cAMP determines intracellular transduction of muscarinic inhibition. This hypothesis accords with distinct cyclic nucleotide compartments for the differential expression of muscarinic inhibition of I-Ca(L). (C) 2001 Elsevier Science BY All rights reserved.