Voltage and concentration dependence of Ca²⁺ permeability in recombinant glutamate receptor subtypes

Abstract

The channels associated with glutamate receptor (GluR) subtypes, namely N‐methyl‐d‐aspartate receptors (NMDARs), and Ca2+‐permeable α‐amino‐3‐hydroxy‐5‐methyl‐4‐isoxazolepropionate receptors (AMPARs) and kainate receptors (KARs), are to varying degrees permeable to Ca2+. To compare the mechanism of Ca2+ influx, we measured Ca2+ permeability relative to that of Na+ (PCa/PNa) using fractional Ca2+ currents (Pf) and reversal potential measurements over a wide voltage and Ca2+ concentration range in recombinant NMDAR NR1‐NR2A, AMPAR GluR‐A(Q) and KAR GluR‐6(Q) channels. For NR1‐NR2A channels, PCa/PNa derived from Pf measurements was voltage independent but showed a weak concentration dependence. A stronger concentration dependence was found when PCa/PNa was derived from changes in reversal potentials on going from a Na+ reference solution to a solution with Ca2+ as the only permeant ion (‘biionic’ condition). In contrast, PCa/PNa was concentration independent when derived from changes in reversal potentials on going from a Na+ reference solution to the same solution with added Ca2+ (‘high monovalent’ condition). For GluR‐A(Q) channels, PCa/PNa derived from all three approaches was concentration independent, and for the reversal potential‐based approaches were of comparable magnitude. Their most distinctive property was that PCa/PNa derived from Pf measurements was strongly voltage dependent. For GluR‐6(Q) channels, PCa/PNa derived from Pf measurements was weakly voltage dependent. On the other hand, PCa/PNa derived from all three approaches was the most strongly concentration dependent of any GluR subtype and, except for low Ca2+ concentrations, the values were of comparable magnitude. Thus, the three Ca2+‐permeable GluR subtypes showed unique patterns of Ca2+ permeability, indicating that distinct biophysical and molecular events underlie Ca2+ influx in each subtype.

Contribution of Calcium Ions to P2X Channel Responses

“…2) because these are well characterized channels that are known to transport Ca 2ϩ across membranes in neurons (Burnashev, 1998;MacDermott et al, 1999;Montell, 2001;Reeves and Lummis, 2002). Our Pf % values for the human ␣4␤2, chick ␣4␤2, and rat NR1/NR2A were 3.1 Ϯ 0.6 (n ϭ 5), 3.1 Ϯ 0.8 (n ϭ 4), and 14.1 Ϯ 0.9% (n ϭ 15) and are in excellent agreement with published reports (Ragozzino et al, 1998;Jatzke et al, 2002;Lax et al, 2002;Watanabe et al, 2002). The Pf % values of the rat 5-HT 3A and rat VR1 channels are not reported previously, and we find them to be 4.7 Ϯ 0.3% (n ϭ 8) and 3.5 Ϯ 0.3% (n ϭ 30), respectively.…”

Glutamate Receptor Ion Channels: Structure, Regulation, and Function

“…Fractional Ca 2ϩ currents (P f ) are the percentage of the total current carried by Ca 2ϩ and are measured using whole-cell currents and high concentrations of fura-2 to ensure that all incoming Ca 2ϩ is captured by dye. P f values are concentration-and voltage-dependent with the magnitude of these effects dependent on specific subtypes Jatzke et al, 2002). The values shown were either recorded at Ϫ60 mV and 1.8 mM Ca 2ϩ or adjusted to these conditions assuming GHK (see Jatzke et al, 2002).…”

“…Only values derived from recombinant channels are shown; see Dingledine et al (1999) for more details and examples of native channels. P Ca /P X values are derived from the Lewis equation (see Jatzke et al, 2002) based on changes in reversal potentials given in articles typically measured under bionic conditions where a reference solution (X, usually Cs ϩ or Na ϩ ) is replaced with a solution containing high Ca 2ϩ (typically Ϸ100 mM). P Ca /P X (from P f ) is derived from P f measurements using the GHK equation .…”

Quantifying Ca2+ Current and Permeability in ATP-gated P2X7 Receptors

“…The Pf % method has distinct advantages over traditional reversal potential measurements of permeability; it is a direct and model-independent measure of Ca 2ϩ entry obtained using physiological concentrations of extracellular ions (51)(52)(53). By contrast, reversal-based techniques do not measure the physiologically relevant Ca 2ϩ current, require high concentrations of extracellular Ca 2ϩ , and make the tenuous assumption that the Ca 2ϩ current follows the assumptions of the Goldman-Hodgkin-Katz equation (54,55).…”