The suprachiasmatic nucleus (SCN), the biological clock, is responsible for a 24-h rhythm in plasma glucose concentrations, with the highest concentrations toward the beginning of the activity period. To investigate whether the SCN is also responsible for daily fluctuations in glucose uptake and to examine how these fluctuations relate to the rhythm in plasma glucose concentrations, SCN-intact rats and SCN-lesioned rats were injected intravenously with a glucose bolus at different time points. We found an increase in glucose uptake toward the beginning of the activity period, followed by a gradual reduction in glucose uptake toward the end of the activity period. The daily variation in glucose tolerance seemed not to be caused by fluctuations in insulin responses of the pancreas but by a daily variation in insulin sensitivity. Lesioning the SCN resulted in the disappearance of the daily fluctuation in glucose uptake and insulin sensitivity. Interestingly, SCN-lesioned rats showed an enhancement in glucose tolerance that could not be explained by higher insulin responses or enhanced insulin sensitivity. Therefore, these findings suggest a role for the SCN in insulin-independent glucose uptake. The present results further show that the daily rhythm in glucose tolerance follows the same pattern as the daily rhythm in plasma glucose concentrations. We hypothesized that the biological clock prepares the individual for the upcoming activity period by two separate mechanisms: increasing plasma glucose concentrations and making tissue more tolerant to glucose.