Myogenic response, a phenomenon in which resistance size arteries and arterioles swiftly constrict or dilate in response to an acute elevation or reduction, respectively, in intravascular pressure is a key component of renal autoregulation mechanisms. Although it is well established that the renal system is functionally immature in neonates, mechanisms that regulate neonatal renal blood flow (RBF) remain poorly understood. In this study, we investigated the hypothesis that members of the transient receptor potential vanilloid (TRPV) channels are molecular components of renal myogenic constriction in newborns. We show that unlike TRPV1–3, TRPV4 channels are predominantly expressed in neonatal pig preglomerular vascular smooth muscle cells (SMCs). Intracellular Ca²⁺ concentration ([Ca²⁺]_i) elevation induced by osmotic cell swelling was attenuated by TRPV4, L-type Ca²⁺, and stretch-activated Ca²⁺ channel blockers but not phospholipase A₂ inhibitor. Blockade of TRPV4 channels reversed steady-state myogenic tone and inhibited pressure-induced membrane depolarization, [Ca²⁺]_i elevation, and constriction in distal interlobular arteries. A step increase in arterial pressure induced efficient autoregulation of renal cortical perfusion and total RBF in anesthetized and mechanically ventilated neonatal pigs. Moreover, intrarenal arterial infusion of the TRPV4 channel blockers HC 067047 and RN 1734 attenuated renal autoregulation in the pigs. These data suggest that renal myogenic autoregulation is functional in neonates. Our findings also indicate that TRPV4 channels are mechanosensors in neonatal pig preglomerular vascular SMCs and contribute to renal myogenic autoregulation.