The muscarinic receptor subtype antagonists 4-diphenylacetoxy-N-methylpiperidine methobromide (4-DAMP), 11-[(2-[diethylamino)methyl]-1- piperidinyl)acetyl]-5,11-dihydro-6H-pyrido[2,3- b][1,4]benzodiazepine-6-one (AF-DX 116) and pirenzepine were used to inhibit the outflow facility and accommodative and miotic responses to near-maximal intracameral doses of pilocarpine in the living rhesus monkey eye. The pharmacologic M3 antagonist 4-DAMP was the most potent inhibitor of all three responses, with IC50 values of 41.7 nM for outflow facility (vs. 40.9 microM pilocarpine), 19.8 nM for accommodation (vs. 40.9 or 81.8 microM pilocarpine) and 3.2 nM for miosis (vs. 4.1 microM pilocarpine). The M1 antagonist pirenzepine was at least 30-fold less potent, with IC50 values of 2.2 microM for outflow facility, 1.4 microM for accommodation and 0.1 microM for miosis. The M2 antagonist AF-DX 116 was the least potent by far, with IC50 values of 15.5 microM for outflow facility, 14.2 microM for accommodation and 1.5 microM for miosis. The results suggest that these three functional responses to pilocarpine are all mediated through an M3 receptor subtype.