Recent study indicates that endogenous 5-hydroxyindole-acetic acid (5-HIAA) clearance can be used as an alternative procedure to para-amino hippurate (PAH) clearance for the estimation of renal plasma flow in human patients. In view of the limitations of PAH clearance measurements in newborn infants we made an attempt to validate the technique of measuring renal blood flow with 5-HIAA quantitatively against PAH clearance. Thirty-four simultaneous determinations of PAH and 5-HIAA clearances were performed in 14 newborn rabbits. 5-HIAA concentrations in plasma and urine were measured by using HPLC coupled with electrochemical detection (Beckman). Renal blood flow was found to range between 0.60 and 6.90 ml/min/kg (mean: 3.39 ml/min/kg) for 5-HIAA and from 0.93 to 6.61 ml/min/kg (mean: 3.68 ml/min/kg) for PAH clearances. There was a significant positive correlation between the values obtained by the two techniques (r = 0.84, P < 0.001). When 5-HIAA clearance was analyzed as a function of plasma 5-HIAA level only a weak, but statistically significant correlation could be detected (r = 0.33, P < 0.05). Plasma 5-HIAA measurement alone, therefore, does not reflect renal blood flow in newborn rabbits. It is concluded that endogenous 5-HIAA clearance might serve as a reliable estimate of renal blood flow in the neonate under different physiologic and pathologic conditions.