Enzyme cytochemical study of rat cardiac muscle I. Adenylate cyclase and guanylate cyclase.

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Abstract
The cytochemical localization of adenylate cyclase and guanylate cyclase in rat cardiac muscle was demonstrated with the lead nitate method using dimethyl sulfoxide (DMSO) (Fuimoto et al., 1981). Tissues were fixed in a mixture of 2% paraformaldehyde, 0.25% glutaraldehyde and 5% DMSO in 0.1 M cacodylate buffer, pH 7.4 for 30 min and then washed with 5% DMSO in 0.1 M cacodylate buffer. Vibratome or microslicer sections were incubated in the following medium for 30 min at 37.degree. C. The medium for adenylate cyclase (ACLase) was 80 mM Tris-maleate buffer, pH 7.4, 4 mM MgSO4, 10 mM NaF, 2 mM theophylline, 2 mM lead nitrate, 0.25 M sucrose, 5% DMSO, 2.5 mM levamisole and 0.5 mM AMP-PNP. The medium for guanylate cyclase (GCLase) was 80 mM Tris-maleate buffer, pH 7.4; 3 mM MnCl2; 2 mM theophylline, 2 mM lead nitrate, 0.25 M sucrose; 5% DMSO, 2.5 mM levamisole and 0.5 mM GMP-PNP. The most intense reaction of ACLase was localized in the sarcolemma, sarcoplasmic reticulum and gap junction. GCLase activity was also seen in the same regions. In the unfixed isolated gap junctions, the reaction product was seen to interrupt the interspace of the junction, forming bridges between the junctional membranes. ACLase was activated by NaF and isoproterenol. Alloxan was effective as an inhibitor of ACLase. GCLase was stimulated by the in vitro acetylcholine treatment.