AIM: To investigate the effects of green flickering light on refractive development and expression of muscarinic acetylcholine receptor (mAChR) M1 in the eyes of guinea pigs. METHODS: Thirty guinea pigs (15-20 days old) were randomly divided into three groups (n=10/group). Animals in group I were raised in a completely closed carton with green flickering light illumination. Those in group II were kept in the open top closed carton under normal natural light. Guinea pigs were raised in a sight-widen cage under normal natural light in group III. The refractive status and axial length were measured before and after 8 weeks' illumination. Moreover, total RNA extracted from retinal, choroidal, and scleral tissues were determined by real-time reverse transcription polymerase chain reaction (RT-PCR). The expressions of the receptor M1 were also explored in the retina, choroid, and sclera using immunohistochemistry. RESULTS: There was a remarkable reduction in refractive error and increase in axial length after 8-weeks' green flickering light stimulation (P<0.001). The expression of M1 receptor mRNA in sclera and retina in myopia group were remarkably lower than that in group II and III (P<0.01). Significant reduced expression of M1 receptor stimulated by green flickering light in retina and sclera tissues were also observed (P<0.05). However, there was no M1 receptor expression in choroid in 3 groups. CONCLUSION: Myopia can be induced by 8 weeks' green flickering light exposure in the animal model. M1 receptor may be involved causally or protectively in myopia development.