A comparative study of the effects of three guanylyl cyclase inhibitors on the L-type Ca2+ and muscarinic K+ currents in frog cardiac myocytes

1 To investigate the participation of guanylyl cyclase in the muscarinic regulation of the cardiac L-type calcium current (I-Ca), we examined the effects of three guanylyl cyclase inhibitors, 1H-[1,2,4]oxidiazolo[4,3-a]quinoxaline-1-one (ODQ), 6-anilino-5,8-quinolinedione (LY 83583), and methylene blue (MBlue), on the beta-adrenoceptor; muscarinic receptor and nitric oxide (NO) regulation of I-Ca and on the muscarinic activated potassium current I-K,I-ACh, in frog atrial and ventricular myocytes. 2 ODQ (10 mu M) and LY 83583 (30 mu M) antagonized the inhibitory effect of an NO-donor (S-nitroso-N-acetylpenicillamine, SNAP, 1 mu M) on the isoprenaline (Iso)-stimulated I-Ca which was consistent with their inhibitory action on guanylyl cyclase. However, MBlue (30 mu M) had no effect under similar conditions. 3 In the absence of SNAP, LY 83583 (30 mu M) potentiated the stimulations of I-Ca by either Iso (20 nM), forskolin (0.2 mu M) or intracellular cyclic AMP (5-10 mu M). ODQ (10 mu M) had no effect under these conditions, while MBlue (30 mu M) inhibited the Iso-stimulated I-Ca. 4 LY 83583 and MBlue, but not ODQ, reduced the inhibitory effect of up to 10 mu M acetylcholine (ACh) on I-Ca. 5 MBlue, but not LY 83583 and ODQ, antagonized the activation of I-K,I-ACh by ACh in the presence of intracellular GTP, and this inhibition was weakened when I-K,I-ACh was activated by intracellular GTP gamma S. 6 The potentiating effect of LY 83583 on Iso-stimulated I-Ca was absent in the presence of either DL-dithiothreitol (DTT, 100 mu M) or a combination of superoxide dismutase (150 u ml(-1)) and catalase (100 u ml(-1)). 7 All together, our data demonstrate that, among the three compounds tested, only ODQ acts in a manner which is consistent with its inhibitory action on the NO-sensitive guanylyl cyclase. The two other compounds produced severe side effects which may involve superoxide anion generation in the case of LY 83583 and alteration of beta-adrenoceptor and muscarinic receptor-coupling mechanisms in the case of MBlue.