Estrogen levels increase during pregnancy and clinical evidence has long suggested that melanocytes are estrogen-responsive. We hypothesized that nevi from pregnant patients would exhibit increased expression of estrogen receptor β (ERβ) and thus enhanced potential to respond to altered estrogen levels. Normal, dysplastic and congenital nevi (n = 212) were collected from pregnant and non-pregnant women ranging from 18 to 45 years of age. Immunohistochemical staining was performed on these nevi using antibodies specifically directed against estrogen receptor α (ERα) and ERβ. ERα was not observed in any lesions; thus, ERβ was the predominant estrogen receptor in melanocytic cells from all types of nevi. Enhanced positivity for ERβ in normal nevi during pregnancy was noted, compared with non-pregnant controls including nevocytes residing in both the epidermal and dermal micro-environments (P = 0.005 and P = 0.001 respectively). Nevi with increasingly melanocytic atypia showed increased ERβ in nevocytes nested within the epidermis. No additional increase in ERβ in atypical nevi was observed during pregnancy. For normal and congenital nevi, regardless of pregnancy status, dermally associated nevocytes tended to have greater ERβ immunoreactivity. Significant decreases in ERβ immunoreactivity were observed in congenital nevi from pregnant women compared with normal and dysplastic nevi from pregnant women. Our data suggest that nevi possess the capacity to be estrogen-responsive. Factors such as pregnancy and degree of atypia are associated with enhanced ERβ with the exception of congenital nevi where the melanocytes were unique in their response to pregnancy.