Specificity of methoctramine in blocking muscarinic receptors which inhibit adenylate cyclase in cerebellar granule cells

Abstract

In primary cultures of cerebellar granule cells, activation of muscarinic receptors stimulates both hydrolysis of phosphatidylinositol (PI) and inhibition of adenylate cyclase. The specificity of three muscarinic receptor antagonists, pirenzepine, methoctramine and (−)quinuclidinyl xanthene-9-carboxylate [(−)QNX], in blocking carbachol-stimulated hydrolysis of PI and inhibition of adenylate cyclase were determined. Pirenzepine was found to be nonspecific in blocking the carbachol-stimulated hydrolysis of PI and inhibition of adenylate cyclase, while methoctramine specifically antagonized carbachol-stimulated inhibition of adenylate cyclase with 600 times greater potency than carbachol-stimulated hydrolysis of PI. (−)Quinuclidinyl xanthene-9-carboxylate was approximately 20 times more potent in blocking the carbachol-stimulated hydrolysis of PI than inhibition of adenylate cyclase. Studies of the ability of these three antagonists to block the binding of [ 3H]quinuclidinyl benzilate ([ 3H]QNB) to muscarinic sites on membranes from cerebellar granule cells, revealed that all three antagonists displayed binding characteristics, characteristic of two binding sites, possibly representing the two types of muscarinic receptors. However, the ratio of the affinities for each of the two binding sites was about ten for pirenzepine, 100 for methoctramine and 650 for (−)QNX. Thus, the specificity of these antagonists, in blocking the inhibition of adenylate cyclase and hydrolysis of PI did not correlate with their specificities obtained with the binding studies with [ 3H]QNB. Since four or possibly five muscarinic receptive proteins have been described, it is possible that this discrepancy can be explained by the high affinity binding of each antagonist to a different subset of muscarinic receptive proteins, some of which are coupled to receptors stimulating the hydrolysis of PI and some to receptors inhibiting adenylate cyclase. Methoctramine seems specific for those muscarinic receptive proteins coupled to the inhibition of adenylate cyclase.