Differential modulation of N‐type α_1B and P/Q‐type α_1A calcium channels by different G protein β subunit isoforms

Abstract

Using transient calcium phosphate transfection into the human embryonic kidney tsa‐201 cell line and subsequent whole‐cell patch‐clamp protocols, we examined the tonic modulation of cloned N‐ and P/Q‐type calcium channels by five different G protein β subunits via strong depolarizing voltage prepulses. For N‐ and P/Q‐type channels, the magnitude of inhibition was dependent on the Gβ subtype co‐expressed. Both the absolute and relative magnitudes of Gβ subunit‐induced inhibition of P/Q‐type channels differed from those observed with the N‐type channel. For each calcium channel subtype, kinetics of both the prepulse‐mediated recovery from inhibition and the re‐inhibition following the prepulse were examined for each of the Gβ subunits by varying either the duration between the pre‐ and the test pulse or the length of the prepulse. For each channel subtype, we observed a differential Gβ subunit rank order with regard to the rates of re‐inhibition and recovery from inhibition. On average, P/Q‐type channels exhibited more rapid rates of recovery from inhibition than those observed with N‐type channels. Different Gβ subtypes mediated different degrees of slowing of activation kinetics. The differential modulation of P/Q‐ and N‐type channels by various Gβ subtypes may provide a mechanism for fine tuning the amount of calcium entering the presynaptic nerve termini.

A Minimal Model for G Protein–Mediated Synaptic Facilitation and Depression

“…Wild-type calcium channel cDNA constructs were supplied by Dr. Terry Snutch. The G protein cDNA constructs used were the same as those described by us previously (Arnot et al 2000;Feng et al 2001).…”

“…There are 5 known G␤ isoforms and 11 known G␥ isoforms (Betty et al 1998), and transient transfection studies have shown that different isoform combinations produce different inhibitory effects on Ca 2ϩ channels (Arnot et al 2000;Diversé-Pierluissi et al 2000;García et al 1998;Ruiz-Velasco and Ikeda 2000;Zhou et al 2000). In most cases, G␤␥ dissociates from channels when the membrane is depolarized (Boland and Bean 1993;Hille 1994;Zamponi and Snutch 1998).…”

G protein inhibition of CaV2 calcium channels

“…For example, the rank order of inhibition produced by the five different Gβ subunits (Gβ [1][2][3][4][5] ), all paired with the same Gγ 2 subunit, differs for Ca V 2.1 and Ca V 2.2 channels. 103 In another study, point mutations on the Gβ 1 subunit had distinct effects on the inhibition of Ca V 2.1 and Ca V 2.2 channels. 104 Functional differences might also arise from variable interactions or crosstalk with other signaling pathways.…”

Molecular Determinants of Syntaxin 1 Modulation of N-type Calcium Channels

“…We have previously described the cloning of G␤ 5 from rat brain via reverse transcriptase-PCR (29). Using XhoI and KpnI sites, the G␤ 5 insert was excised from pGEM T-Easy and subcloned in-frame into pTrcHisA.…”