2-deoxy-D-[14C]-glucose (2DG) autoradiography

After MRI, three rats per group were randomly picked for 2DG autoradiography. Rats were anesthetized with isoflurane (3% with 700cc/min O₂) and [¹⁴C]−2DG (American Radiolabeled Chemicals Inc., St Louis, MO) was delivered as an intravenous bolus injection of 125µCi/kg body weight via the tail vein. Animals were awake for a tracer circulation time of 45 minutes, and then euthanized by isoflurane overdose, decapitated and brains were rapidly removed. The brains were then submerged shortly in 0.3 M sucrose on the ice, sectioned into 3 mm sections, frozen on dry ice and stored at −80 °C. Blocks were cut in a cryostat at −15 °C in 20 μm thick coronal sections then dried on glass slides. Slides were covered with scintillating sheets and exposed for 18 hours together with [¹⁴C] standard (ARC-146C(PL); St. Louis, MO) then analyzed in Microimager (Biospace lab, France). Measurements were performed on selected regions of interest (ROIs) placed in gray matter from CT and white matter in CC. Counts obtained from each slide were calibrated on the [¹⁴C] standard and values were then converted to nCi/g tissue.