RT info:eu-repo/semantics/article T1 Caffeine chelates calcium in the lumen of the endoplasmic reticulum A1 Rojo Ruiz, Jonathan A1 Rodríguez Prados, Macarena A1 Río Lorenzo, Alba del A1 Alonso Alonso, María Teresa A1 García-Sancho Martín, Francisco Javier K1 Calcio K1 Calcium AB Cytosolic Ca2+ signals are often amplified by massive calcium release from the endoplasmic reticulum (ER). This calcium-induced calcium release (CICR) occurs by activation of an ER Ca2+ channel, the ryanodine receptor (RyR), which is facilitated by both cytosolic- and ER Ca2+ levels. Caffeine sensitizes RyR to Ca2+ and promotes ER Ca2+ release at basal cytosolic Ca2+ levels. This outcome is frequently used as a readout for the presence of CICR. By monitoring ER luminal Ca2+ with the low-affinity genetic Ca2+ probe erGAP3, we find here that application of 50 mM caffeine rapidly reduces the Ca2+ content of the ER in HeLa cells by ∼50%. Interestingly, this apparent ER Ca2+ release does not go along with the expected cytosolic Ca2+ increase. These results can be explained by Ca2+ chelation by caffeine inside the ER. Ca2+-overloaded mitochondria also display a drop of the matrix Ca2+ concentration upon caffeine addition. In contrast, in the cytosol, with a low free Ca2+ concentration (10−7 M), no chelation is observed. Expression of RyR3 sensitizes the responses to caffeine with effects both in the ER (increase in Ca2+ release) and in the cytosol (increase in Ca2+ peak) at low caffeine concentrations (0.3–1 mM) that have no effects in control cells. Our results illustrate the fact that simultaneous monitoring of both cytosolic- and ER Ca2+ are necessary to understand the action of caffeine and raise concerns against the use of high concentrations of caffeine as a readout of the presence of CICR. PB Portland Press SN 0264-6021 YR 2018 FD 2018 LK http://uvadoc.uva.es/handle/10324/35192 UL http://uvadoc.uva.es/handle/10324/35192 LA eng NO Biochemical Journal, 2018, 475 (22) 3639-3649 NO Producción Científica DS UVaDOC RD 14-ene-2025