Glomerular mesangial cells (MCs) regulate glomerular filtration rate (GFR) by regulating effective filtration surface area. Canonical transient receptor potential (TRPC) 6 is a Ca2+ permeable cation channel and playa an important role in renal physiology and pathophysiology. We have recently demonstrated that TRPC6 was expressed in MCs and modulated MC contractile function. The aim of the present study was to determine whether and how TRPC6 in MCs can regulate GFR. GFR in mice was estimated using a two-compartment clearance model. 5% FITC-inulin (3.74 μl/g body weight) was retroorbitally injected under anesthetized state and blood samples were periodically collected from the tail vein under the conscious state. GFR in rats was evaluated based on the plasma inulin clearance during intravenous infusion of 1% FITC-inulin at a rate of 1 ml/h/100 g BW. Blood samples were collected from the left common carotid artery and urine samples were collected from either the bladder or the urether under the anesthetized state (pentobarbital, 50 mg/kg BW, i.p). In vivo knockdown of TRPC6 in rat kidneys was achieved by injection of shRNA constructs against rat TRPC6 via the left renal artery under anesthetized state. Ca2+ imaging was employed to assess intracellular Ca2+ level. Furthermore, MC contractile response was assessed by measuring a change in the planar surface area of MCs in response to Ang II. We found that GFR in TRPC6 knockout mice (881 ± 238 μl/min, n=6) was remarkably elevated compared to wild type mice (177 ± 61 μl/min, n=7) (P<0.01, Student t-test). In rats, acute infusion of Ang II significantly reduced GFR (1.2 ± 0.1 vs. 0.7 ± 0.1 ml/min/100 g BW, before vs. after Ang II, n=8, P<0.05, paired t-test). However, the Ang II response was abolished by knockdown of TRPC6 in kidneys. Immunohistochemistry showed that MCs were the predominantly transfected glomerular cells. Consistently, the GFR of the transfected kidney significantly decreased by Ang II when transfected with control vectors (0.53 ± 0.21 vs. 0.33 ± 0.21 ml/min/100 g BW, before vs. after Ang II, P<0.05, paired t-test, n=5), but did not have a significant change when transfected with shRNA against TRPC6. There were no significant differences in blood pressure and renal blood flow between TRPC6-knocked down and control rats. Furthermore, Ang II-stimulated Ca2+ entry was significantly inhibited in MCs isolated from TRPC6 knockout mice and in human MCs with TRPC6 knocked down. Moreover, the Ang II-stimulated contraction was significantly suppressed in TRPC6 deficient mouse MCs. These results suggest that TRPC6 channel is an important component regulating GFR by modulating MC Ca2+ signaling and contraction.