Crucial role of type 1, but not type 3, inositol 1,4,5-trisphosphate (IP3)receptors in IP3-induced Ca2+ release, capacitative Ca2+ entry, and proliferation of A7r5 vascular smooth muscle cells

,Stimulation of G protein- or tyrosine kinase-coupled receptors regulates c ell proliferation through intracellular Ca2+ ([Ca2+](i)) signaling. In A7r5 cells, we confirmed that inositol 1,4,5-trisphosphate (IP3) mediates vasop ressin (VP)-evoked Ca2+ release from intracellular stores and showed that t ypes 1 (IP3R1) and 3 (IP3R3) IP3 receptors were expressed. Using antisera s elective for IP3R1 or IP3R3 and another that interacted equally well with b oth subtypes, together with membranes from Sf9 cells expressing only single IP3R subtypes to calibrate immunoblotting, we established that A7r5 cells express 81% IP3R1 and 19% IP3R3. To elucidate the contributions of IP3R1 an d IP3R3 to Ca2+ signaling and proliferation, stable clones expressing promo ter-inducible antisense cDNA fragments (-90 to +9) corresponding to the two IP3R subtypes were selected. Mild inhibition of IP3R1 (71+/-8% of control level) slightly attenuated the IP3-evoked Ca2+ release (IICR) induced by VP but significantly decreased the subsequent capacitative Ca2+ entry (CCE) a nd proliferation. Moderate inhibition (34+/-6%) strongly decreased both IIC R and CCE and further blocked proliferation. Complete inhibition almost abo lished IICR and CCE and arrested proliferation entirely. Complete inhibitio n of IP3R3 expression slightly attenuated IICR without affecting CCE or pro liferation In cells microinjected with a low dose of heparin, VP-induced CC E was more susceptible than IICR to mild inhibition of both IP3R1 and IP3R3 . A high dose of heparin had a similar effect to complete inhibition of IP3 R1 expression: it blocked VP-evoked IICR entirely and CCE by 90%. We conclu de that IP3R1, but not IP3R3, is crucial for IICR, CCE, and proliferation o f vascular smooth muscle cells.