OBJECTIVE To investigate the expression of the muscarinic receptor (mAChR) subtypes (m1, m2, m3, m4, and m5) in the human ciliary muscle. METHODS cDNA probes specific for the m1 to m5 subtypes were used to characterize the expression of subtype-specific mRNAs by in situ hybridization and Northern blot analysis on a cell line derived from human ciliary muscle tissue (H7CM), sectioned tissue from four donors, and fresh human ciliary muscle tissue from three donors. RESULTS Messenger RNA for m2, m3, and m5 was detected in situ hybridization and Northern blot analysis in the H7CM cell line and in human ciliary muscle tissue from the donors. In human ciliary muscle tissues, the expression of mRNA of the m2 and m3 subtypes was more prominent in the circular portion than in the longitudinal portion. In contrast, mRNA for the m5 subtype was greater in the longitudinal portion than in the circular portion. Additionally, Northern blot analysis showed that m1 and m4 were expressed in ciliary muscle tissue. CONCLUSIONS These studies show that the human ciliary muscle definitely expresses the mRNA of subtypes m2, m3, and m5, and may also express the mRNA of m1 and m4. Although the expression of mRNA for the m2, m3, and m5 subtypes differed between the circular and longitudinal portions of the ciliary muscle, the strong expression of all three subtypes of muscarinic receptors by both portions demonstrates that the differential distribution of muscarinic subtypes probably is not responsible for the dissociation between outflow facility and accommodation that can occur under certain conditions.