The Calcium Channel A2D Subunit Increases the Gating Charges of Cav1.2 Channels

Abstract

Optically tracking the movement of individual voltage sensors (VSD) of human CaV1.2 revealed that association of α2δ-1 with α1C resulted in a substantial change in the intrinsic voltage-sensing properties of VSDs I-III (Savalli et al 2016, JGP 142, 147-159). Co-expression of α2δ-1 increased the sensitivity of activation for several VSDs indicating that this mostly extracellular subunit increased the electrical distances traversed by voltage-sensing charges. In this study we measured the slope of the voltage-dependence of the channel's probability of being open (Po) at extremely low probability using marcropatches of Xenopus oocytes expressing CaV1.2/CaVβ3 by themselves or in combination with α2δ-1. Patches containing from 50 to several 100 channels were recorded in 75 mM Ba2+ and in the presence of 1 µM (-) Bay K 8644 and held at several voltages ranging from −70 to −20 mV for 10 to 90 s. Po times the number of channels (NPo) were obtained by finding the most likely combination of normal distributions shifted by the single channel currents amplitude that described the data. The relative weight of the different distributions was then contrasted with what should be expected from a Poisson distribution of opening levels. The number of channels (N) was estimated from the noise analysis of hundreds of tail current traces at −40 mV following a depolarizing pulse to +80 mV. In the presence of α2δ-1, the limiting slope was about 4 elementary charges, close to the sum of the voltage-dependencies of VSDII and VSDIII activation and clearly less than the sum of all VSDs and thus consistent with the idea that only a subset of VSDs contribute to the effective charges for channel opening. When the α2δ-1 subunit was absent the limiting slope was reduced by about one elementary charge. This modest change can be accounted by a reduction in the electrical distance that gating charges need to cross for channel opening.Funding: FONDECYT 3140590 (GC) and 1120864 (AN), R01GM110276 (RO) and 16POST27250284 (NS).