OBJECTIVE Acidosis-induced [Ca2+]i transients emerged in heart ischemia is an old story.Currently, three mechanisms are involved in the Ca2+ transients in cardiomyocytes: a.the activation of Na +/H + exchanger and then the secondary stimulation of Na+/Ca2+ exchanger; b.Ca2+ release from intracellular Ca2+ stores; c.the opening of the Ca2+-permeable membrane ion channels.Although the above ways are well studied, the concrete mechanisms for acidosis-induced Ca2+ elevation in cardiomyocytes are largely unknown.In this study, we will try to identify the expression of the sensors of protons, including acid sensing ion channels (ASICs), vanilloid-1-transient potential receptor (TRPV1) and proton sensing G protein coupled receptors in cardiomyocytes, and explore the possibilities of these sensors to mediate the acidosis-induced [Ca2 +] i elevation in ventricular cardiomyocytes.METHODS RT-PCR, western blotting, immunofluorescence and Ca2+ imaging.RESULTS 1.The sensors of protons ASICs, TR-PV1 and proton sensing G protein were first identified in in vitro and in vivo cardiomyocytes.2.Rapid application of extracellular solutions (pH 5.0) markedly elevated [Ca2+]i in primary cardiomyocytes with the value of △F/F 2.64 ± 0.13.The protons-induced [Ca2+]i elevation was significantly inhibited by both the non-specific inhibitor (amiloride, 300 μmol·L-1) and the specific inhibitor of ASICs (PcTx1, 10 nmol·L-1).The inhibitory ratio was 50.07 ± 5.70% and 53.51 ± 3.31%, respectively.3.The inhibitor of TRPV1 (ruthenium red, 10 μ mol· L-1) also blunted the elevation of [Ca2+]i with the down-regulated ratio of 56.16 ± 6.22%.Interestingly, the down-regulated ratio increased to 82.61 ±4.18% after amiloride and ruthenium red co-incubation.4.When Ca2+ was depleted, robust elevation of [Ca2 +] i still could be observed.The elevation of [Ca2 +] i in the absence of extracellular Ca2 + was almost totally abolished by TG, the inhibitor of SR ATPase that can exhaust the Ca2 + content in SR Ca2+ stores.The inhibitory ratio was 90.59 ± 1.46 %.5.2-APB (200 μmol·L-1) and U73122 (3 μmol·L-1), both were regarded as the blocker of IP3 receptor in SR/ER and the inhibitor of PLC in cytoplasm, also significantly decreased the amplitude of [Ca2 +] i elevation in the absence of extracellular Ca2+.The inhibitory ratio was 49.80 ±4.85% and 53.93 ±5.14% in 2-APB and U73122-treated groups, respectively.6.Because the only protein OGR1 that was reported to activate Gq protein in the family of proton sensing G protein coupled receptors was detected in in vitro and in vivo cardiomyocytes.We applied Cu2+ to block the proton binding sites.Cu2+ (200 μmol· L-1) significantly attenuated the elevation of [Ca2+] i triggered by pH 5.0 acidic solutions with the inhibitory ratio of 47.91 ± 8.35%.CONCLUSION Our current study suggested that three kinds of H +-sensitive proteins, ASICs, TRPV1 and OGR1, mediate acidosis-induced Ca2+ transients in ventricular cardiomyocytes.Extracellular acidification mediated Ca2 + influx via rnembrane ASICs and TRPV1 and also mobilized intracellular Ca2+ from SR via OGR1-PLC-IP3-IP3R signaling pathway.This finding will raise the opportunities for us to understand the involvement of membrane ion channels and intracellular IP3 receptors in heart ischemia.