Ca<sup>2+</sup>-sensitivity of human Na<sub>V</sub>1.4<sub>F1705I</sub> inactivation.

<p>(<b>A</b>) A schematic of the structured region of the C-terminus of hNa<sub>V</sub>1.4 between amino acids residues 1788 and 2040, the predicted helices are labeled H1–H6. The location of the EFL residues in and around H1 harbors species specific variations in the key Ca<sup>2+</sup> sensing residues in hNa<sub>V</sub>1.4 (G1613S and A1636D) compared with the rat isoform. The CaM binding motif IQ in H6 and, the cold aggravated myotonia mutation F1705I (rat: F1698I) in H5 are illustrated. (<b>B</b>) Representative whole-cell currents through wild type and mutant hNa<sub>V</sub>1.4<sub>F1705I</sub> channels expressed in HEK293 cells in [Ca<sup>2+</sup>]<sub>i</sub> free conditions. Na<sup>+</sup> currents were elicited by the protocol in the inset. (<b>C</b>) [Ca<sup>2+</sup>]<sub>i</sub> does not alter the I–V relationship of hNa<sub>V</sub>1.4<sub>F1705I</sub>. (<b>D</b>) Representative steady-state inactivation currents from different holding potentials through mutant hNa<sub>V</sub>1.4<sub>F1705I</sub> channels in the presence of 0.5 µM or absence of Ca<sup>2+</sup>. (<b>E</b>) Activation and steady-state inactivation curves of wild type and hNa<sub>V</sub>1.4<sub>F1705I</sub> channels in the absence and presence of 0.5 µM intracellular Ca<sup>2+</sup>. The V<sub>1/2</sub> of inactivation of hNa<sub>V</sub>1.4<sub>F1705I</sub> is sensitive to [Ca<sup>2+</sup>]<sub>i</sub> and significantly shifted in the hyperpolarizing direction in the presence of Ca<sup>2+</sup> (p<0.005). The activation relationships are fitted with dotted lines, the V<sub>1/2</sub> of activation are unaffected by change in [Ca<sup>2+</sup>]<sub>i</sub>. (<b>F</b>) and (<b>G</b>) illustrate the window currents through wild type and hNa<sub>V</sub>1.4<sub>F1705I</sub> in presence (<b>F</b>) and absence (<b>G</b>) of intracellular Ca<sup>2+</sup>, respectively. Dotted lines represent the wild type channel. The symbols and color are the same in plots C, E, F, and G.</p>



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