PURPOSE. The purpose of this study was to investigate the relationship between circadian rhythm and intraocular pressure (IOP), and to explore whether electrical stimulation of cervical sympathetic ganglia (SCG) can regulate IOP via neurotransmitter distribution around the Schlemm's canal (SC) in rats. METHODS. Sprague Dawley rats were housed under normal (N-normal), constant dark (N-dark), and constant light (N-light) rhythms (n = 6 per group). Electrical stimulation (intermittent wave [20 hertz {Hz}, 2 mA, 10 minutes]) was used to stimulate the SCG. Atropine sulfate eye gel was applied three times a day. DiI was injected into the SCG and anterior chamber. The cross-sectional area and circumference of SC were evaluated using hematoxylin-eosin staining. Immunofluorescence staining was used to evaluate dopamine-beta-hydroxylase (D beta H) expression in SC endothelial (SCE) cells. RESULTS. N-Dark increased the IOP, decreased the cross-sectional area of SC, and increased D beta H levels in SCE cells. Nerve projection between SC and SCG was detected, and electrical stimulation of SCG upregulated D beta H expression in SCE cells. Under normal and constant light rhythms, electrical stimulation of SCG increased D beta H and decreased the cross-sectional area and circumference of SC, while simultaneously increasing IOP and decreasing IOP fluctuations. After paralyzing the ciliary muscles, electrical stimulation of SCG decreased the cross-sectional area and circumference of SC under normal and constant light rhythms. CONCLUSIONS. N-Dark increased D beta H in SCE cells, reduced the cross-sectional area of SC, and increased IOP. Under the normal and light rhythms, electrical stimulation of SCG increased D beta H in SCE cells, reduced the cross-sectional area and circumference of SC, and in turn elevated IOP and decreased IOP fluctuations.