In vivo characterization of vasodilating muscarinic-receptor subtypes in humans

The role of muscarinic (M)-receptor subtypes in the regulation of vascular tone has not yet been defined in humans. To analyze the role of M-receptor subtypes in the forearm resistance vasculature of normotensive volunteers (n = 20), we infused acetylcholine (ACh) and methacholine (MCh) in the presence of saline and the antagonists atropine (nonselective), pirenzepine (M1 selective), and AF-DX 116 (M2 selective), using automated R-wave-triggered venous occlusion plethysmography. Schild analysis was applied by calculating plasma concentrations of the infused compounds and determining EC50 values. ACh and MCh both caused dose-dependent vasodilation, with EC50 values of 537 and 52 nmol/L, respectively. The apparent 10-fold higher potency of MCh compared with ACh may be explained by rapid degradation of ACh in plasma. The concentration-response curve of MCh was shifted to the right by atropine, pirenzepine, and AF-DX 116, with apparent pA2 values of 8.03 +/- 0.03, 6.71 +/- 0.08, and 5.32 +/- 0.05, respectively, and slopes not different from unity. The present technique enabled us to perform M-receptor characterization by Schild analysis in humans. The affinity constants and rank order of potency--atropine > pirenzepine > AF-DX 116-suggest that cholinergic vasodilation in this vascular bed is predominantly mediated by the M3-receptor subtype. The EC50 value of MCh and the pA2 values of pirenzepine and AF-DX 116 are comparable to values reported for in vitro experiments.