Myogenic response and tubuloglomerular feedback are the two primary mechanisms for renal blood flow autoregulation by regulating the afferent arteriolar vascular resistance. However, the mechanisms of mechanotransduction in pressure induced myogenic response are not well defined in afferent arterioles. We have previously demonstrated that integrin‐mediated mechanical force increases the occurrence of Ca2+ sparks in freshly isolated renal vascular smooth muscle cells (VSMCs). To test whether generation of Ca2+ sparks is a downstream signal of mechanotransduction in pressure induced myogenic constriction, the relationship between Ca2+ spark frequency and transmural perfusion pressure was investigated in intact VSMCs of afferent arterioles isolated from Sprague‐Dawley rats. Spontaneous Ca2+ sparks were found in VSMCs when afferent arterioles were perfused at 80 mmHg. The Ca2+ spark frequency was significantly increased when perfusion pressure was increased by 40 mmHg. Similar increase of Ca2+ spark frequency was also found when arterioles were stimulated with β1‐integrin activating antibody. Spontaneous membrane current recorded with whole cell perforated‐patch in renal VSMCs indicated that there were more spontaneous transient inward currents (STICs) than spontaneous transient outward currents (STOCs) when holding potential was set close to physiological resting membrane potential. These observations indicated that Ca2+ sparks activity might couple to Ca2+‐activated Cl− (ClCa) channel and trigger pressure induced myogenic constriction via membrane depolarization. Real‐time PCR and immunofluorescence confirmed the presence of the ClCa channel TMEM16A in renal VSMCs. Inhibition of TMEM16A with the specific antagonist T16Ainh‐A01 impaired pressure induced myogenic contraction in the perfused afferent arterioles. It is known that myogenic response is enhanced in spontaneously hypertensive rat (SHR). TMEM16A abundance and spontaneous Ca2+ sparks were more prevalent in renal arterioles of SHR than those of Sprague‐Dawley rats, which are consistent with notion that Ca2+ spark activity is part of the signal transduction process in myogenic response. In conclusion, Ca2+ sparks were detected in intact VSMCs of afferent arterioles for the first time and their frequencies were positively modulated by the perfusion pressure. The coupling Ca2+ sparks to ClCa might contribute to the mechanical signal transduction process in pressure induced myogenic constriction.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.