Microdomain [Ca²⁺] near ryanodine receptors as reported by L-type Ca²⁺ and Na⁺/Ca²⁺ exchange currents

During Ca²⁺ release from the sarcoplasmic reticulum triggered by Ca²⁺ influx through L-type Ca²⁺ channels (LTCCs), [Ca²⁺] near release sites ([Ca²⁺]_nrs) temporarily exceeds global cytosolic [Ca²⁺]. [Ca²⁺]_nrs can at present not be measured directly but the Na⁺/Ca²⁺ exchanger (NCX) near release sites and LTCCs also experience [Ca²⁺]_nrs. We have tested the hypothesis that I_CaL and I_NCX could be calibrated to report [Ca²⁺]_nrs and would report different time course and values for local [Ca²⁺]. Experiments were performed in pig ventricular myocytes (whole-cell voltage-clamp, Fluo-3 to monitor global cytosolic [Ca²⁺], 37°C). [Ca²⁺]_nrs-dependent inactivation of I_CaL during a step to +10 mV peaked around 10 ms. For I_NCX we computationally isolated a current fraction activated by [Ca²⁺]_nrs; values were maximal at 10 ms into depolarization. The recovery of [Ca²⁺]_nrs was comparable with both reporters (>90% within 50 ms). Calibration yielded maximal values for [Ca²⁺]_nrs between 10 and 15 μmol l^-1 with both methods. When applied to a step to less positive potentials (-30 to -20 mV), the time course of [Ca²⁺]_nrs was slower but peak values were not very different. In conclusion, both I_CaL inactivation and I_NCX activation, using a subcomponent analysis, can be used to report dynamic changes of [Ca²⁺]_nrs. Absolute values obtained by these different methods are within the same range, suggesting that they are reporting on a similar functional compartment near ryanodine receptors. Comparable [Ca²⁺]_nrs at +10 mV and -20 mV suggests that, although the number of activated release sites differs at these potentials, local gradients at release sites can reach similar values.