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PH-regulated Slo3 K+ channels: properties of unitary currents.

ABSTRACT: Here we have examined the voltage and pH dependence of unitary Slo3 channels and used analysis of current variance to define Slo3 unitary current properties over a broader range of voltages. Despite complexity in Slo3 channel openings that precludes simple definition of the unitary conductance, average current through single Slo3 channels varies linearly with voltage at positive activation potentials. Furthermore, the average Slo3 unitary current at a given activation potential does not change with pH. Consistent with macroscopic conductance estimates, the apparent open probability of Slo3 channel exhibits a pH-dependent maximum, with limiting values around 0.3 at the most elevated pH and voltage. Estimates of Slo3 conductance at negative potentials support a weaker intrinsic voltage dependence of gating than is observed for Slo1. For the pH-regulated Slo3 K(+) channel, the dependence of macroscopic conductance on pH suggests that the pH-sensitive mechanism regulates gating in an allosteric manner qualitatively similar to regulation of Slo1 by Ca(2+). Together, the results support the view that the regulation of macroscopic Slo3 currents by pH reflects regulation of gating equilibria, and not a direct effect of pH on ion permeation. Specifically, both voltage and pH regulate a closed-open conformational change in a largely independent fashion.

SUBMITTER: Zhang X

PROVIDER: S-EPMC2151565 | biostudies-literature | 2006 Sep

REPOSITORIES: biostudies-literature

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pH-regulated Slo3 K+ channels: properties of unitary currents.

Zhang Xue X Zeng Xuhui X Xia Xiao-Ming XM Lingle Christopher J CJ

The Journal of general physiology 20060901 3

Here we have examined the voltage and pH dependence of unitary Slo3 channels and used analysis of current variance to define Slo3 unitary current properties over a broader range of voltages. Despite complexity in Slo3 channel openings that precludes simple definition of the unitary conductance, average current through single Slo3 channels varies linearly with voltage at positive activation potentials. Furthermore, the average Slo3 unitary current at a given activation potential does not change w ...[more]

PMID: 16940554

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Project description:The mouse Slo3 gene (KCNMA3) encodes a K(+) channel that is regulated by changes in cytosolic pH. Like Slo1 subunits responsible for the Ca(2+) and voltage-activated BK-type channel, the Slo3 alpha subunit contains a pore module with homology to voltage-gated K(+) channels and also an extensive cytosolic C terminus thought to be responsible for ligand dependence. For the Slo3 K(+) channel, increases in cytosolic pH promote channel activation, but very little is known about many fundamental properties of Slo3 currents. Here we define the dependence of macroscopic conductance on voltage and pH and, in particular, examine Slo3 conductance activated at negative potentials. Using this information, the ability of a Horrigan-Aldrich-type of general allosteric model to account for Slo3 gating is examined. Finally, the pH and voltage dependence of Slo3 activation and deactivation kinetics is reported. The results indicate that Slo3 differs from Slo1 in several important ways. The limiting conductance activated at the most positive potentials exhibits a pH-dependent maximum, suggesting differences in the limiting open probability at different pH. Furthermore, over a 600 mV range of voltages (-300 to +300 mV), Slo3 conductance shifts only about two to three orders of magnitude, and the limiting conductance at negative potentials is relatively voltage independent compared to Slo1. Within the context of the Horrigan-Aldrich model, these results indicate that the intrinsic voltage dependence (z(L)) of the Slo3 closed-open equilibrium and the coupling (D) between voltage sensor movement are less than in Slo1. The kinetic behavior of Slo3 currents also differs markedly from Slo1. Both activation and deactivation are best described by two exponential components, both of which are only weakly voltage dependent. Qualitatively, the properties of the two kinetic components in the activation time course suggest that increases in pH increase the fraction of more rapidly opening channels.

Project description:Wild-type AMPA receptors display a characteristic rapidly desensitizing phenotype. Many studies point to the dimer interface between pairs of extracellular ligand binding domains as the key region controlling the rate at which the receptors desensitize. However, mutations at the extracellular end of the pore-forming regions (near the putative ion channel gate) have also been shown to alter desensitization. Here we report the behavior of single GluA4 receptors carrying one of two mutations that greatly reduce desensitization at the level of ensemble currents: the dimer interface mutation L484Y and the Lurcher mutation (A623T, GluA4-Lc) in the extracellular end of M3 (the second true transmembrane helix). Analysis of unitary currents in patches with just one active receptor showed that each mutation greatly prolongs bursts of openings without prolonging the apparent duration of individual openings. Each mutation decreases the frequency with which individual receptors visit desensitized states, but both mutant receptors still desensitize multiple times per second. Cyclothiazide (CTZ) reduced desensitization of wild-type receptors and both types of mutant receptor. Analysis of shut-time distributions revealed a form of short-lived desensitization that was resistant to CTZ and was especially prominent for GluA4-Lc receptors. Despite reducing desensitization of GluA4 L484Y receptors, CTZ decreased the amplitude of ensemble currents through GluA2 and GluA4 LY receptor mutants. Single-channel analysis and comparison of the GluA2 L483Y ligand binding domain dimer in complex with glutamate with and without CTZ is consistent with the conclusion that CTZ binding to the dimer interface prevents effects of the LY mutation to modulate receptor activation, resulting in a reduction in the prevalence of large-conductance substates that accounts for the decrease in ensemble current amplitudes. Together, the results show that similar nondesensitizing AMPA-receptor phenotypes of population currents can arise from distinct underlying molecular mechanisms that produce different types of unitary activity.

Dataset Information

PH-regulated Slo3 K+ channels: properties of unitary currents.

Publications

pH-regulated Slo3 K+ channels: properties of unitary currents.

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