Serum cystatin C concentration correlates negatively with glomerular filtration rate as well as or better than that of serum creatinine, suggesting a constant formation, and elimination from extracellular fluid mainly by glomerular filtration. It is not known, however, how well the renal plasma clearance of this 13-kDa basic polypeptide matches the glomerular filtration rate. This was investigated in rats during control conditions and after reduced renal perfusion pressure. 125I-cystatin C and an indicator for glomerular filtration (51Cr-EDTA or 131I-aprotinin) were injected intravenously. The renal accumulation and urinary excretion of the tracers were recorded in periods of 2.5 to 20.0 min. The renal plasma clearance of 125I-cystatin C (Ccy) based on the renal content of 125I correlated well with the glomerular filtration rate (CCr-EDTA) in periods up to 6 min; i.e. Ccy = 0.94 x CCr-EDTA, r = 0.99. Less than 0.5% of the filtered amount appeared in the urine. During more prolonged periods, Ccy increasingly underestimated glomerular filtration rate, reaching about 0.4 x CCr-EDTA in a 20-min period. Free 125I relative to total plasma 125I activity increased from about 2% at 5 min to about 70% at 20 min. In nephrectomized rats, free 125I accumulated in plasma at a slower rate, accounting for about 15% of the total activity 20 min after injection of 125I-cystatin C. We conclude that cystatin C is (a) mainly removed from the extracellular fluid by the kidneys, (b) practically freely filtered in the glomeruli, and (c) completely absorbed and rapidly broken down by the proximal tubular cells.