Brain volume is regulated during acute hyperosmolality based, in part, on the tissue gain of Na, Cl, and K. This study evaluates the contribution of bulk flow of cerebrospinal fluid (CSF) into brain to the volume regulatory gain of electrolytes. Artificial CSF containing radiolabeled albumin and diethylenetriamine penta-acetic acid (DTPA) was perfused for 60 min through the ventricles and/or subarachnoid space of anesthetized rats, and tracer clearances from CSF to brain were measured as a function of plasma osmolality. Osmolality was elevated after 30 min of perfusion by intraperitoneal injection of hypertonic NaCl or mannitol. Albumin and DTPA clearances increased with osmolality at the same rate, despite a sevenfold difference in diffusion coefficient, consistent with osmotic stimulation of a bulk flow component of tracer influx into brain. The volume shift estimated on the basis of this data is 114 microliters CSF/g dry wt brain for a 60-mosmol/kg increase in osmolality. Results indicate that CSF is a major source of the volume regulatory gain of Na and Cl, but not of K.