Abstract After secretion by the heart, atrial natriuretic factor (ANF) circulates in plasma, whereas C-type natriuretic peptide (CNP), which is found in abundance in the endothelium, may regulate vascular tone in a paracrine manner. However, there is little information on the effect of CNP on renal microvessels. We hypothesized that CNP dilates the afferent arteriole via the nitric oxide pathway, whereas ANF acts directly on vascular smooth muscle cells. When we perfused rat kidneys with minimal essential medium and bovine serum albumin at 100 mm Hg and examined the juxtamedullary afferent arterioles, neither CNP nor ANF was found to have any effect. When the peptides were added to arterioles preconstricted with norepinephrine, CNP and ANF dilated them in a similar fashion; diameters increased by 25±4% (n=7) and 29±6% (n=6) at 10 −7 mol/L, respectively ( P <.008). Pretreatment with 10 −4 mol/L N -nitro- l -arginine methyl ester (L-NAME) or 5×10 −6 mol/L indomethacin blocked CNP-induced dilation; dilation by ANF was unaffected by indomethacin (52±25%, n=5) and potentiated by L - NAME (73±14%, n=5). Thus, CNP dilates the afferent arterioles via the prostaglandin/nitric oxide pathway, whereas ANF dilates them directly. This difference may be important in controlling glomerular hemodynamics.