EGTA Can Inhibit Vesicular Release in the Nanodomain of Single Ca²⁺ Channels

The exogenous Ca²⁺ chelator EGTA (ethylene glycol tetraacetic acid) has been widely used to probe the coupling distance between Ca²⁺ channels and vesicular Ca²⁺ sensors for neurotransmitter release. Because of its slow forward rate for binding, EGTA is thought to not capture calcium ions in very proximity to a channel, whereas it does capture calcium ions at the remote distance. However, in this study, our reaction diffusion simulations (RDSs) of Ca²⁺ combined with a release calculation using vesicular sensor models indicate that a high concentration of EGTA decreases Ca²⁺ and vesicular release in the nanodomain of single channels. We found that a key determinant of the effect of EGTA on neurotransmitter release is the saturation of the vesicular sensor. When the sensor is saturated, the reduction in the Ca²⁺ concentration by EGTA is masked. By contrast, when the sensor is in a linear range, even a small reduction in Ca²⁺ by EGTA can decrease vesicular release. In proximity to a channel, the vesicular sensor is often saturated for a long voltage step, but not for a brief Ca²⁺ influx typically evoked by an action potential. Therefore, when EGTA is used as a diagnostic tool to probe the coupling distance, care must be taken regarding the presynaptic Ca²⁺ entry duration as well as the property of the vesicular Ca²⁺ sensor.