In order to develop a suitable radiotracer for the measurement of glial metabolism, we synthesized four different types of ester derivatives of [14C]acetate, namely, [14C]phenyl acetate, [14C]para-nitrophenyl acetate, [14C]2,4-dinitrophenyl acetate and [14C]benzyl acetate ([14C]BA), and evaluated their potencies in rats. Among the derivatives, the highest brain uptake at 30 s postinjection was observed for [14C]BA, which was more than 23 times higher than that of [14C]acetate itself. A long-term retention of [14C]BA radioactivity in the brain was observed, whereas rapid clearance of radioactivity was seen in the heart. [14C]BA was rapidly hydrolyzed in the intact rat brain, and less than 5% of radiolabeled parent was observed 1 min after the injection. Radiochemical analysis using thin-layer chromatography revealed that [14C]BA was rapidly converted to [14C]glutamine and [14C]glutamate in the cortex within 10 min after injection. Furthermore, the uptake of [14C]BA was significantly decreased following microinjection of fluorocitrate, a selective glial toxin. These results strongly suggest that [14C]BA may be a useful radiotracer for the measurement of glial metabolism in the intact rat brain.