ACTIVATION OF GUANOSINE 3',5'-CYCLIC-MONOPHOSPHATE (CGMP)-DEPENDENT PROTEIN-KINASE IN RAT VAS-DEFERENS AND DISTAL COLON IS NOT ACCOMPANIED BY INHIBITION OF CONTRACTION

There is good evidence that in vascular smooth muscle, the relaxant ef fects of sodium nitroprusside (SNP) are mediated by increases in cGMP levels and activation of cGMP-dependent protein kinase (PKG). However, in rat vas deferens and rat distal colon, cGMP-elevating agents such as SNP and atrial natriuretic factor (ANF) have been shown to elevate cGMP without inducing relaxation. The lack of relaxation might be expl ained by either lack of activation of PKG by these agents or low level s of PKG in these tissues. The object of the present study was to inve stigate these possibilities by simultaneously monitoring cGMP levels, PKG activity and contractility in isolated strips of rat vas deferens, rat proximal colon and distal colon exposed to high concentrations of SNP or ANF. Verification of the specificity of the assay for PKG was obtained using MonoQ chromatography to resolve soluble smooth muscle e xtracts, followed by immunoblotting with a PKG-specific antibody to id entify the kinase. In rat vas deferens, 5 mM SNP increased cGMP levels (14-fold) and PKG activity ratios (3.4-fold) but did not inhibit phen ylephrine-induced contractions. In both rat proximal and rat distal co lon, 100 nM ANF significantly elevated cGMP levels and PKG activity ra tios, but only in the proximal colon was inhibition of spontaneous con tractions observed. Total PKG activity was much lower (approximate to 16 pmol PO4/min/mg protein) in rat vas deferens, which was not relaxed by SNP, than in rabbit aorta (approximate to 148 pmol PO4/min/mg), wh ich was relaxed. However, in the rat proximal colon, despite low PKG l evels (approximate to 11 pmole/min/mg), ANF did inhibit contractions. Thus the inability of the cGMP-elevating agents SNP and ANF to inhibit contractions in rat vas deferens and rat distal colon cannot be expla ined by either of the possibilities suggested above.