A new double-label autoradiographic glucose analog method for the sequential measurement of altered regional cerebral metabolic rates for glucose in the same animal is presented. This method is based on the sequential injection of two boluses of glucose tracer labeled with two different isotopes (short-lived 18F and long-lived 3H, respectively). An operational equation is derived which allows the determination of glucose utilization for the time period before the injection of the second tracer; this equation corrects for accumulation and loss of the first tracer from the metabolic pool occurring after the injection of the second tracer. An error analysis of this operational equation is performed. The double-label deoxyglucose method is validated in the primary somatosensory ("barrel") cortex of the anesthetized rat. Two different rows of whiskers were stimulated sequentially in each rat; the two periods of stimulation were each preceded by an injection of glucose tracer. After decapitation, dried brain slices were first exposed, in direct contact, to standard X-ray film and then to uncoated, "tritium-sensitive" film. Results show that the double-label deoxyglucose method proposed in this paper allows the quantification and complete separation of glucose utilization patterns elicited by two different stimulations sequentially applied in the same animal. The double-label deoxyglucose is of potential usefulness in sensory physiology since it makes possible the separate mapping of regional cerebral glucose utilization patterns elicited by two sequentially applied sensory stimulations in the same animal. The method allows the quantification of a step-like change in regional cerebral glucose utilization in the same animal. It could be used to study the cerebral metabolic effects induced by neuropharmacological agents or surgical interventions applied during the experiment. Using each animal as its own control eliminates intersubject variability. Thus experimental cost and effort can be saved, and the reliability of the results obtained can be increased.