To prevent bone loss, OPG/RANK/RANKL signalling pathway is a key in keeping the balance between the action of osteoblasts and osteoclasts. Aim of this study is to assess the influence of long-term nicotine exposure on bone mineral density (BMD) scores, RANKL and OPG levels of plasma and RANKL and OPG immunoreactivities of tissue in rats. In this study, totally 36 Swiss Albino rats (70±10 g) were used in three groups. Whereas normal drinking water was given for the control group (n:12), 0.4 mg/kg/day and 6.0 mg/kg/day nicotine was added to drinking water for low-dose nicotine (LDN) group (n:12) and high-dose nicotine (HDN) group (n:12), respectively for 12 months. At the end of 12^th month, BMD scores were measured via X-ray absorptiometry and then bone turnover was assessed via measuring both RANKL, OPG levels in plasma and RANKL, OPG immunoreactivities in tail vertebrae of all rats. Lumbar spine and femoral regions BMD scores of the control group and the nicotine groups were not significantly difference. In HDN group, OPG levels of plasma were found significantly higher when compared with the control and LDN groups (p=0.001) unlike RANKL levels of plasma. RANKL and OPG immunoreactivities of tissue were found significantly lower in both LDN and HDN groups (p<0.001, p=0.004, respectively) in comparison to control group. No correlation was found between plasma levels and tissue immunoreactivities of RANKL and OPG. As a result, this study indicates that nicotine is not primarily responsible for the decline of BMD frequently seen in smokers.