To examine the reliability of quantitative positron emission tomography studies in the rat (Rat-PET), we assessed the influence of radioactivity accumulated in the Harderian glands on PET CMRglc determination. We measured CMRglc by PET and ex vivo dissection methods by using 2-[18F]fluoro-2-deoxy-D-glucose in rats with and without focal brain ischemia. The CMRglc values obtained by PET, after correcting with recovery coefficients, were higher than those measured by the ex vivo method at rostral slices, and reduction of the CMRglc in the ischemic brain was not demonstrated by PET in the frontal cortex. The radioactivity accumulated in the Harderian glands prevents the quantitative determination of CMRglc using Rat-PET.