The blood-brain transport and regional distribution of a tritium-labeled, brominated benzodiazepine (BFB) was determined for the rat brain in vivo. The unidirectional transport constant from blood to brain was measured by a graphical, integral method and was found to be 0.83 ml/g/min, a value which indicates that transport is essentially flow-dependent. The apparent volume of distribution increased linearly during the measurement period, suggesting that back transport from brain to blood was zero and that BFB was trapped in the tissue, possibly by specific receptors or acceptors. Under the conditions of these experiments, autoradiography of brain tissue sections indicated a regional distribution of [3H]BFB similar to that expected for regional cerebral blood flow. These results indicate that BFB is a useful blood flow tracer in brain and suggest that BFB radiobrominated with bromine-75 may, under appropriate conditions, be a suitable tracer for in vivo regional blood flow measurement or benzodiazepine receptor mapping by positron-emission tomography.