PHARMACOLOGICAL PROFILES FOR RAT CORTICAL M1 AND M2 MUSCARINIC RECEPTORS USING SELECTIVE ANTAGONISTS - COMPARISON WITH N1E-115 MUSCARINIC RECEPTORS

We previously showed that M1 and M2 muscarinic receptors in dissociated cells of the adult rat cortex couple to phosphoinositide (PI) and cyclic AMP (cAMP) metabolism, respectively. To further classify these receptors according to probable subtype, we have employed a group of selective muscarinic antagonists to obtain pharmacological profiles of the cortical M1 and M2 receptors, and to compare them with the muscarinic receptors in N1E-115 cells, which contain M1 receptors mediating cyclic GMP elevation and M4 receptors inhibiting cAMP levels. The M2-mediated inhibition of cAMP levels in cortex was blocked by 4-diphenylacetoxy-N-methyl piperidine methiodide (4-DAMP) with higher potency (0.29 nM) than for reported potency in cardiac tissue (approximately 10 nM), indicating that this cortical response is probably not mediated by the m2 gene product. Similarly, the potency of hexahydrosiladiphenidol (HSD) at the cortical M2 receptor (159 nM) was somewhat greater than the reported potency in cardiac tissue (295 nM). The cardioselective drugs AF-DX 116 and methoctramine blocked the cortical M2 response less potently (135 nM and 229 nM, respectively) than would be expected for involvement of the m2 gene product. Thus, the potencies of AF-DX 116, methoctramine, 4-DAMP and HSD suggest that the cortical M2 response, like the striatal M2 receptor, is mediated by a noncardiac M2 receptor, perhaps by the m4 gene product. This postulate was supported by the significant correlations between cortical and striatal M2 receptors as compared to the M4 receptor in N1E-115 cells (r = 0.92 and 0.99, respectively, P < 0.25). Although HSD was more potent at blocking the cortical M1 receptor (K(i) = 20 nM) as compared to the M2 receptor (K(i) = 159 nM), the drugs 4-DAMP, secoverine and AF-DX 116 were equipotent in their blockade of these two receptor-effector systems. Para-fluoro-HSD, a putative M3-selective antagonist, competitively blocked the cortical M1 receptor with lower potency (K1 = 56 nM) than would be expected for involvement of an m3 gene product, and was ineffective in blocking the cortical M2 receptor. In a comparison using antagonist potencies, it appears that the cortical M1 receptor and the M1 receptor in N1E-115 cells are very similar (r = 0.99, P < .01). However, pirenzepine inhibited the carbachol-mediated phosphoinositide response in cortical dissociated cells with a Hill slope significantly less than unity, indicating partial involvement of a second muscarinic receptor in this response. When these data were fitted with a two-site model, the results were consistent with a view that the majority (73%) of the cortical PI response is mediated by the m1 gene product (pirenzepine K(i) = 5.5 nM); the remaining fraction may be mediated by the m3 or other gene products. Similarly, the bulk (74%) of muscarinic receptor-mediated PI turnover in rat hippocampus was shown to involve a receptor with M1-type affinity for pirenzepine (31 nM). Our studies thus indicate that the cortical and striatal muscarinic receptors that couple to cAMP inhibition are noncardiac M2 receptors, possibly m4 gene products, whereas PI metabolism in cortex is mediated primarily, but not entirely, by the m1 gene product. Using antagonist potencies, the pharmacological profiles of cortical and N1E-115 muscarinic receptors appear very similar.