Luminal Ca²⁺ and Mg²⁺ sensitivity of RyR2 channels. Channels were incorporated into bilayers by fusing heavy native SR microsomes. Cytosolic free Ca²⁺ concentration was constant (1 μM). The luminal Ca²⁺ or Mg²⁺ concentration was titrated from 10 μM to 10 mM. Holding potential was 0 mV. The luminal solution contained 100 mM Cs⁺ and net unit current was always in the lumen-to-cytosolic direction. (A) Example single channel recordings with zero current level marked. Recordings at left are from a control channel that was never exposed to the CSQ2 stripping process with no added CSQ2 in the luminal bathing solution. Recordings at right are from a channel that was first CSQ2 stripped before 0.5 μg/ml CSQ2-WT was added to the luminal solution. (B) Summary Po results (mean ± SEM) where the filled circles represent the Ca²⁺ sensitivity of 11 different control channels and the open circles represent the Mg²⁺ sensitivity of 10 different channels. The X-marked circles represent data collected from six different CSQ2-associated control channels in the presence of 3 mM cytosolic diBromoBAPTA.