The present study investigated the drug distribution and elimination profiles in ocular tissues of pirenzepine, a selective M1 muscarinic antagonist known to inhibit myopia. Results demonstrate that (1) Intravitreal injections of the M1 selective antagonist pirenzepine were more effective at preventing form-deprivation myopia than subconjunctival injections. (2) Maximum drug levels were reached within 1 hr for both retina and sclera following intravitreal (28 and 11 nanomole) and subconjunctival (0.25 and 1 nanomole) injection. Intravitreal injection proved a more effective route of drug delivery to all ocular tissues compared to subconjunctival injection. (3) Elimination times of pirenzepine from ocular tissues were much shorter than those reported for blood plasma. (4) Histological examination revealed no evidence of gross toxic effects at doses effective in inhibiting induced axial myopia. In conclusion, pirenzepine was effective at reducing form-deprivation myopia in a dose-dependent manner with no evidence of disruption to the retina. However, results were not conclusive as to where pirenzepine may have its site of action in preventing form-deprivation myopia.