The right cerebral hemisphere of the rat was perfused in situ by retrograde infusion of HCO3 saline or blood into the right external carotid artery. Infusion rate was adjusted to minimize the contribution of systemic blood to flow in the hemisphere. During perfusion with whole or artificial blood, regional cerebral blood flow and blood volume were comparable to respective values in the conscious rat, whereas perfusion with HCO3 saline increased regional flow three- to fourfold due to the low viscosity of the saline perfusate. Perfusion with whole blood for 300 S or with HCO3 saline for 60 S did not alter the permeability of the blood-brain barrier. Cerebrovascular permeability coefficients of eight nonelectrolytes ranged from 10(-8) to 10(-4) cm X S-1 and were directly proportional to the octanol-water partition coefficient of the solute. Thus the in situ brain perfusion technique is a sensitive new method to study cerebrovascular transfer in the rat and permits absolute control of perfusate composition.