PT - JOURNAL ARTICLE AU - Ildiko Kovacs AU - Henry I. Yamamura AU - Sue L. Waite AU - Eva V. Varga AU - William R. Roeske TI - Pharmacological Comparison of the Cloned Human and Rat M2 Muscarinic Receptor Genes Expressed in the Murine Fibroblast (B82) Cell Line DP - 1998 Feb 01 TA - Journal of Pharmacology and Experimental Therapeutics PG - 500--507 VI - 284 IP - 2 4099 - http://jpet.aspetjournals.org/content/284/2/500.short 4100 - http://jpet.aspetjournals.org/content/284/2/500.full SO - J Pharmacol Exp Ther1998 Feb 01; 284 AB - The coding sequence of the human m2 receptor gene was amplified by polymerase chain reaction and stably transfected into a murine fibroblast cell line (B82). We have compared the human M2 clonal cell line (HM2-B10) with the previously established B82 cell line (M2LKB2–2) expressing the rat M2 receptor to assess drug specificity, drug selectivity and effector coupling. Both transfected cell lines showed a high level of specific, saturable [3H](−)-N-methyl-3-quinuclidinyl benzilate binding withKd values of 243 pM (155–352 pM) and 345 pM (234–539 pM) and Bmax values of 97 ± 4 and 338 ± 16 fmol/106 cells, respectively. Inhibition of [3H](−)-N-methyl-3-quinuclidinyl benzilate binding to HM2-B10 cells and M2LKB2–2 cells showed the same rank order of potency for the antagonists: atropine > dexetimide > 4-diphenylacetoxy-N-methylpiperidine methiodide > himbacine > methoctramine > 11-[[2-[(diethylamino) methyl]-1-piperidinyl]acetyl]-5,11-dihidro-6H-pyrido-[2,3-b](1,4)-benzodiazepine-6-one > hexahydro-sila-difenidol hydro-chloride > pirenzepine. Correlation analysis of the pKi values indicate that the expressed human and rat M2 receptors have nearly identical ligand-binding characteristics. Carbachol inhibited forskolin-stimulated cAMP formation with similar potency in both cell lines [EC50 = 2.4 μM (0.2–2.8) and 1.1 μM (0.2–5.3) for the human and rat M2 receptor, respectively]. In the M2LKB2–2 cells, carbachol slightly stimulated the [3H]inositol monophosphate formation but had no significant effect in HM2-B10 cells. In conclusion, the human and rat M2 receptors expressed in the B82 cell line have very similar binding properties but exhibit slight differences in effector coupling mechanisms. The American Society for Pharmacology and Experimental Therapeutics