Cardiac microvascular endothelial cells express a functional Ca2+ -sensing receptor

The mechanism whereby extracellular Ca 2+ exerts the endothelium- dependent control of vascular tone is still unclear. In this study, we assessed whether cardiac microvascular endothelial cells (CMEC) express a functional extracellular Ca 2+ - sensing receptor (CaSR) using a variety of techniques. CaSR mRNA was detected using RT-PCR, and CaSR protein was identified by immunocytochemical analysis. In order to assess the functionality of the receptor, CMEC were loaded with the Ca 2+ -sensitive fluorochrome, Fura-2/AM. A number of CaSR agonists, such as spermine, Gd 3+ , La 3+ and neomycin, elicited a heterogeneous intracellular Ca 2+ signal, which was abolished by disruption of inositol 1,4,5-trisphosphate (InsP 3 ) signaling and by depletion of intracellular stores with cyclopiazonic acid. The inhibition of the Na + /Ca 2+ exchanger upon substitution of extracellular Na + unmasked the Ca 2+ signal triggered by an increase in extracellular Ca 2+ levels. Finally, aromatic amino acids, which function as allosteric activators of CaSR, potentiated the Ca 2+ response to the CaSR agonist La 3+ . These data provide evidence that CMEC express CaSR, which is able to respond to physiological agonists by mobilizing Ca 2+ from intracellular InsP 3 -sensitive stores.