Naltriben

To measure SOCE, ER Ca²⁺ store depletion was stimulated by pre-incubation with 1 μM of thapsigargin (Sigma-Aldrich, USA; #:T9033) for 20 minutes prior to the experimental protocols in Ca²⁺-free Ringer’s solution. Subsequently, under fluorescence recordings, cells were continuously perfused for 120 seconds with Ca²⁺-free Ringer’s solution, followed by a re-addition of 2 mM of extracellular Ca²⁺ in normal Ringer’s solution to measure the delta (Δ) SOCE peak. To block SOCE, cells were also preincubated with pharmacological CRAC antagonists synta-66 (5 μM) (Sigma-Aldrich, USA; #:1949) or with 3,5-bis trifluoromethyl pyrazole-BTP-2 (20 μM) (Sigma-Aldrich, USA; #:203890) for two hours [10 (link),14 (link),39 (link)]. synta-66 concentration was determined after the construction of the concentration-effect curve (Fig. S2, B). The TRPM7 agonist, naltriben (NAL, 100 μM) (Tocris Bioscience, USA; #:0892) [31 (link),35 (link)], was perfused simultaneously with the Ca²⁺ re-addition for 720 seconds. In other sets of experiments, the TRPM7 antagonist NS8593 (30 μM) (Tocris Bioscience, USA; #:4597) was simultaneously perfused during the Ca²⁺ re-addition, in the absence or presence of naltriben (100 μM), as described [35 (link)].