Abstract:The presence of specific melatonin binding sites in the Harderian gland of Syrian hamsters was studied using [125I]melatonin. Saturation binding experiments conducted with [125I] melatonin at 37°C using Harderian glands of both male and female Syrian hamsters revealed a single nanomolar‐affinity site. The dissociation constants (Kd) were 6.47 and 6.94 nM for males and females, respectively. The concentration of the binding sites was 7.58 fmol/mg protein for males and 13.50 fmol/mg protein for females. Castration of male hamsters resulted in a significant increase in [125I] melatonin binding sites while chronic melatonin administration did not modify the binding properties. The results confirm the presence of melatonin binding sites in the Harderian glands of rodents. The gender‐associated differences found together with the effects of castration in male hamsters suggest an androgenic control in [125I] melatonin binding sites of the Syrian hamster Harderian g