Utilizing the in vitro blood-perfused juxtamedullary nephron preparation, we examined the effects of alterations in renal arterial pressure on afferent arteriolar blood flow. With video microscopy and cross-correlation techniques, arteriolar inside diameters and centerline erythrocyte velocity were measured to estimate single afferent arteriolar blood flow. In response to random changes in perfusion pressure, afferent arteriolar diameter (n = 8) varied inversely (-0.53 ± 0.02%/mmHg), and erythrocyte velocity was directly related (1.4 ± 0.1%/mmHg). Above 95 mmHg, the slope of the relationship between perfusion pressure and afferent arteriolar blood flow did not differ from zero (0.081 ± 0.053%/mmHg), suggesting efficient autoregulation. When the tubuloglomerular feedback pathway was interrupted by the addition of furosemide (n = 9) or papillectomy (n = 7), there was attenuation of pressure-induced afferent arteriolar constriction, with impairment in blood flow autoregulation (0.60 ± 0.05%/mmHg). Superfusion with diltiazem abolished autoregulatory responses in afferent arteriolar diameter and blood flow (1.5 ± 0.2%/mmHg). These data demonstrate the autoregulation of blood flow of individual afferent arterioles in juxtamedullary nephrons and suggest that both tubulo-glomerular feedback-dependent and -independent mechanisms are required for autoregulatory responses.