The store-operated calcium-release-activated calcium current, ICRAC, is a major mechanism for calcium entry into non-excitable cells. ICRAC refills calcium stores and permits sustained calcium signalling. The relationship between inositol 1,4,5-trisphosphate receptor (InsP3R)-containing stores and ICRAC is not understood. A model of global InsP3R store depletion coupling with ICRAC activation may be simplistic, since intracellular stores are heterogeneous in their release and refilling activities. Here we use a ligand-gated calcium channel, TRPV1 (transient receptor potential channel, vanilloid subfamily member 1), as a new tool to probe store heterogeneity and define intracellular calcium compartments in a mast cell line. TRPV1 has activity as an intracellular release channel but does not mediate global calcium store depletion and does not invade a store coupled with ICRAC. Intracellular TRPV1 localizes to a subset of the InsP3R-containing stores. TRPV1 sensitivity functionally subdivides the InsP3-sensitive store, as does heterogeneity in the sarcoplasmic/endoplasmic-reticulum Ca2+-ATPase isoforms responsible for store refilling. These results provide unequivocal evidence that a specific ‘CRAC store’ exists within the InsP3-releasable calcium stores and describe a novel methodology for manipulation of intracellular free calcium.
Abbreviations used: ICRAC, calcium-release-activated calcium current; ER, endoplasmic reticulum; FBS, foetal bovine serum; fura 2/AM, fura 2 acetoxymethyl ester; InsP3, inositol 1,4,5-trisphosphate; InsP3R, InsP3 receptor; MOI, multiplicity of infection; pfu, plaque-forming units; RBL, rat basophilic leukaemia; SERCA, sarcoplasmic/endoplasmic-reticulum Ca2+-ATPase; SOC, store-operated calcium; TRPV1, transient receptor potential channel, vanilloid subfamily member 1.
Present address: Laboratory of Cell Biology and Immunology, Center for Biomedical Research at the Queen's Medical Center, Honolulu, HI, U.S.A.