(A) [Ca2+]ER-sensitive D1ER traces representing ER Ca2+ depletion with 10 mM caffeine, followed by ER refilling upon 1 mM [Ca2+]o addition in an mCherry-ER-3– (control, black) or TRIC-A-mCherry (TRIC-A, red)–transfected HEK293_RyR2 cell or diminished refilling in a 3 μM BTP2-incubated control cell (BTP2, blue). Bar graphs show mean ± SEM values for (B) ER refilling time and (C) ER refill rate in TRIC-A (+) cells (n = 45) versus controls (n = 51), and (D) ER Ca2+ release peak amplitude in TRIC-A (+) cells (n = 45) and BTP2-incubated cells (n = 36) versus controls (n = 51); *p < 0.05, **p < 0.01, ***p < 0.001. Underlying data in panels (A–D) are included in S1 Data. BTP2, N-[4-[3,5-Bis(trifluoromethyl)pyrazol-1-yl]phenyl]-4-methylthiadiazole-5-carboxamide; D1ER, genetically encoded ER-targeted Ca2+ sensor; ER, endoplasmic reticulum; HEK293, human embryonic kidney 293; ns, nonsignificant; RyR, ryanodine receptor; TRIC, trimeric intracellular cation.