This study examines the effects of a low-protein diet on cholesterol metabolism in rats under constant darkness. The experiment was conducted in two-way layout methods with two factors: lighting conditions and dietary protein levels. Each factor has two levels: lighting condition has normal lighting (7:00-19:00; light period, 19:00-7:00; dark period, N) and constant darkens (D) ; dietary protein levels has 5% casein (5%) and 20% casein (20%) . Forty-eight Fisher strain 4-week-old female rats were divided into four groups based on the two factors and levels. At the end of the fourth week, three rats in each group were euthanized at 14:00, 20:00, 2:00, and 8:00. The effects of lighting condition and dietary protein level on the serum total and hepatic cholesterol concentrations were observed. The D-condition serum concentration was higher than that of the N-condition (p<0. 01) and that of the 5%-diet was lower than that of the 20%-diet (p<0. 01) . Conversely, the D-condition hepatic concentration was lower than that of the N-condition (p<0. 05) and that of the 5%-diet was higher than that of the 20%-diet (p<0. 01) . The effects of both factors on ApoC3, an apolipoprotein of VLDL, were observed. The ApoC3 mRNA expression of the D-condition was higher than that of the N-condition (p<0. 01) , and that of the 5%-diet was lower than that of the 20%-diet (p<0. 01) . We also observed lighting condition on mRNA expressions of Lrp1 (a receptor of lipoprotein) at 2:00 and Hmgcr (a rate-limiting enzyme of cholesterol synthesis) at 20:00. The mRNA expressions of both genes in the D-condition were lower than in the N-condition (p<0. 05) . There were no significant differences between lighting conditions and dietar y protein levels. Therefore, both factors were acting independently, and the decrease in the serum concentration by low-protein diet was related to ApoC3 and the increase by constant darkness was related to ApoC3, Lrp1, and Hmgcr.