Cardiac Microvascular Endothelial Cells Express a Functional Ca²⁺-Sensing Receptor

The mechanism whereby extracellular Ca²⁺ 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 Ca²⁺-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 Ca²⁺-sensitive fluorochrome, Fura-2/AM. A number of CaSR agonists, such as spermine, Gd³⁺, La³⁺ and neomycin, elicited a heterogeneous intracellular Ca²⁺ signal, which was abolished by disruption of inositol 1,4,5-trisphosphate (InsP₃) signaling and by depletion of intracellular stores with cyclopiazonic acid. The inhibition of the Na⁺/Ca²⁺ exchanger upon substitution of extracellular Na⁺ unmasked the Ca²⁺ signal triggered by an increase in extracellular Ca²⁺ levels. Finally, aromatic amino acids, which function as allosteric activators of CaSR, potentiated the Ca²⁺ response to the CaSR agonist La³⁺. These data provide evidence that CMEC express CaSR, which is able to respond to physiological agonists by mobilizing Ca²⁺ from intracellular InsP₃-sensitive stores.