Activation of coronary arterial guanylate cyclase by nitric oxide,nitroprusside, and nitrosoguanidine—Inhibition by calcium,lanthanum, and other cations,enhancement by thiols

Although reports that certain vasodilate s activate soluble guanylate cyclase, especially in the presence of thiols, and elevate cyclic GMP levels in smooth muscle suggest that cyclic GMP is involved in vascular smooth muscle relaxation, earlier reports that Ca²⁺ activates guanylate cyclase and that Ca²⁺ -dependent contractile agents elevate cyclic GMP levels are seemingly at odds with this hypothesis. The objective of this study was to examine the effects of Ca²⁺ related cations, and thiols on bovine coronary arterial soluble guanylate cyclase. Guanylate cyclase activity was detected in the presence of Mg²⁺ or Mn²⁺ but not of other cations. Basal activity was greater in the presence of Mn²⁺ than of Mg²⁺. Activity of guanylate cyclase stimulated by nitroprusside, nitric oxide, or nitrosoguanidine, however, was greater with Mg²⁺, although the requirement of activated enzyme for Mn²⁺ was reduced about 10-fold. Ca²⁺ markedly inhibited guanylate cyclase activation in the presence of Mg²⁺ but not of Mn²⁺. La²⁺ inhibited enzyme activation in the presence of Mg²⁺ or Mn²⁺. Neither Ca²⁺ nor La³⁺ altered basal enzymatic activity. Results that were qualitatively similar to those indicated above were observed with partially purified, heme-free, coronary arterial soluble guanylate cyclase. Nitric oxide and nitroso compounds activated partially purified enzyme, and thiols enhanced enzyme activation by nitroprusside and nitrosoguanidine without appreciably altering basal activity. Irreversible sulfhydryl binding agents such as ethacrynic acid and gold inhibited both basal and activated guanylate cyclase. These results suggest that changes in intracellular concentrations of free Ca²⁺ and sulfhydryl groups could influence the rate of formation of cyclic GMP by vasodilators and that this, in turn, could alter smooth muscle tone.     

Inhibitory effects of calmodulin antagonists on plasma membrane cyclases in Tetrahymena: calmodulin-dependent guanylate cyclase and calmodulin-independent adenylate cyclase

Trifluoperazine was shown previously to inhibit the activation of Tetrahymena guanylate cyclase activity by calmodulin [S. Nagao, S. Kudo and Y Nozawa, Biochem. Pharmac. 19, 2709 (1981)]. The present paper reports that N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide (W-7), another representative calmodulin inhibitor, inhibited the calmodulin-induced activation of the guanylate cyclase, and that trifluoperazine and W-7 also inhibited Tetrahymena adenylate cyclase. The adenylate cyclase activity was found to be present in a membrane-bound form and not to be influenced by calmodulin. The inhibitions of the adenylate cyclase activity by these agents were dose-dependent and not Ca2+-dependent. These findings suggest that the inhibitory actions of these drugs may not necessarily be specific for calmodulin-dependent enzymes in T. pyriformis.     

Inhibition of the cyclic AMP-adenylate cyclase system and of secretion by high concentrations of adenosine in the dog thyroid

Adenosine inhibits cyclic AMP accumulation in stimulated slices and adenylate cyclase in acellular preparations of dog thyroid. The onset of this inhibition is rapid, requires relatively high adenosine concentrations (greater than or equal to 10 microM) and occurs with all activators tested (TSH, PGE1, forskolin and cholera toxin). The manganous ion, which uncouples receptor and cyclase, enhances the inhibition by adenosine. The effect of 2',5'-dideoxyadenosine, the high concentration of adenosine needed, the Mn2- effect and the lack of reversal by methylxanthines all suggest that this effect bears on the "P"-site, i.e. on the cyclase itself. Adenosine also inhibits thyroid secretion, which shows that its effect bears on the follicular cells. However the fact that cyclic AMP and DB cyclic AMP induced secretion are also reduced by adenosine suggests that adenosine also inhibits cyclic AMP action.     

Calcium-dependent modulation of guanosine 3′,5′-monophosphate in renal cortex: Possible relationship to calcium-dependent release of fatty acid

The effects of Ca²⁺ on cGMP accumulation in rat renal cortical slices were correlated with the effects on ¹⁴C-fatty acid release in tissue prelabeled with ¹⁴C arachidonate. Ca²⁺ in the presence and absence of ionophore A23187 exerted parallel effects on the release of labeled arachidonate from slices and on slice cGMP content. Thus, Ca²⁺ stimulated both arachidonate release and tissue cGMP accumulation 2 to 3-fold when added to slices of renal cortex previously deprived of Ca²⁺ and Mg²⁺, whereas Mg²⁺ had no stimulatory effect on either arachidonate release or tissue cGMP content. In the presence of A23187, Ca²⁺ increased arachidonate release and tissue cGMP accumulation 4 to 6-fold. Tetracaine partially inhibited Ca²⁺-induced arachidonate release and completely blocked Ca²⁺-induced cGMP accumulation. Ca²⁺-induced arachidonate release was unaffected by the absence of O₂. Addition of exogenous arachidonate to slices of renal cortex increased tissue cGMP content 2-fold. Linoleate exerted a lesser effect on tissue cGMP, while palmitate and oleate had no effect. Ca²⁺- and arachidonateinduced cGMP contents in renal cortical slices were not additive, and both were abolished by exclusion of O₂. Since nitroprusside increased cGMP accumulation 10- to 15-fold in O₂-deprived slices, loss of the Ca²⁺ and arachidonate responses under these incubation conditions was selective. Ca²⁺-induced cGMP accumulation was unaffected by indomethacin (100 μg/ml), but was abolished by 200 μM 5,8,11,14-eicosatetraynoic acid (TYA). The results are consistent with the possibility that the Ca²⁺-dependent processes regulating cGMP in renal cortex include Ca²⁺-dependent acyl hydrolase activity, which limits the availability of free polyunsaturated fatty acids. A role for fatty acid oxygenation products in the stimulation of cGMP is suggested, but not established, by the O₂ dependence of the actions of both Ca²⁺ and exogenous fatty acids. The failure of exogenous arachidonate or linoleate to mimic quantitatively the actions of Ca²⁺ on cGMP may reflect the involvement of other Ca²⁺- and O₂-dependent processes in modulation of cGMP in this tissue or limited access of exogenous fatty acid to cGMP regulatory sites in the cell.