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Link to original content: https://pubmed.ncbi.nlm.nih.gov/29674667
A Mixed Periodic Paralysis & Myotonia Mutant, P1158S, Imparts pH-Sensitivity in Skeletal Muscle Voltage-gated Sodium Channels - PubMed Skip to main page content
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. 2018 Apr 19;8(1):6304.
doi: 10.1038/s41598-018-24719-y.

A Mixed Periodic Paralysis & Myotonia Mutant, P1158S, Imparts pH-Sensitivity in Skeletal Muscle Voltage-gated Sodium Channels

Mohammad-Reza Ghovanloo  1 , Mena Abdelsayed  1 , Colin H Peters  1 , Peter C Ruben  2
Affiliations

A Mixed Periodic Paralysis & Myotonia Mutant, P1158S, Imparts pH-Sensitivity in Skeletal Muscle Voltage-gated Sodium Channels

Mohammad-Reza Ghovanloo et al. Sci Rep. .

Abstract

Skeletal muscle channelopathies, many of which are inherited as autosomal dominant mutations, include myotonia and periodic paralysis. Myotonia is defined by a delayed relaxation after muscular contraction, whereas periodic paralysis is defined by episodic attacks of weakness. One sub-type of periodic paralysis, known as hypokalemic periodic paralysis (hypoPP), is associated with low potassium levels. Interestingly, the P1158S missense mutant, located in the third domain S4-S5 linker of the "skeletal muscle", Nav1.4, has been implicated in causing both myotonia and hypoPP. A common trigger for these conditions is physical activity. We previously reported that Nav1.4 is relatively insensitive to changes in extracellular pH compared to Nav1.2 and Nav1.5. Given that intense exercise is often accompanied by blood acidosis, we decided to test whether changes in pH would push gating in P1158S towards either phenotype. Our results suggest that, unlike in WT-Nav1.4, low pH depolarizes the voltage-dependence of activation and steady-state fast inactivation, decreases current density, and increases late currents in P1185S. Thus, P1185S turns the normally pH-insensitive Nav1.4 into a proton-sensitive channel. Using action potential modeling we predict a pH-to-phenotype correlation in patients with P1158S. We conclude that activities which alter blood pH may trigger the noted phenotypes in P1158S patients.

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Conflict of interest statement

The authors declare no competing interests.

Figures

Figure 1
Figure 1
Homology model of sodium channel. (A–C) Show the views of the location of P1158 from the side, intracellular, and extracellular sides. The structure is colored by domain. P1158 is highlighted in red. (D,E) Show the hinge angle difference between having proline (84.9°) or serine (87.2°) at position 1158. (F) Show the location of the P1158S mutation on a generic sodium channel schematic (highlighted in red), and the conservation of this part of Nav1.4, aligned with the rest of sodium channels.
Figure 2
Figure 2
Normalized conductance plotted against membrane potential. (A–D) Show overlaps of WT (blue squares) and P1158S (grey circles) conductance at pH6.4, 7.0, 7.4, and 8.0. (E,F) Show conductance of each WT and P1158S. (G) Shows the voltage-dependence of activation as a function of pH and fitted with Hill equation (P1158S) or a flat line (WT). (H) Shows pulse protocol.
Figure 3
Figure 3
Current and conductance densities measured in pA/pF and nS/pF, respectively. (A–H) Sample macroscopic sodium currents elicited by depolarizations between −100 and +80 mV. (I) Average current (left Y-axis) and conductance (right Y-axis) densities of P1158S and WT at extracellular pH between 6.4 and 8.0. The solid bars represent current density and patterned bars represent conductance density. **Indicate p < 0.01, *indicate p < 0.05 (P1158S at pH6.4 and 7.0 are significantly different from pH7.4 and 8.0; P1158S at physiological pH of 7.4 is significantly different from WT at pH7.4). (J) Sample proton block trace. Maximal current amplitude is plotted over a series of depolarizing pulses. External solution pH is 8.0 at the start of the trace. pH6.4 gets perfused at the pulses indicated by arrows. (K) Sample current amplitude at pH8.0 and 6.4 in WT and P1158S.
Figure 4
Figure 4
Voltage-dependence of steady-state fast inactivation as normalized current plotted against membrane potential. (A–D) Show the voltage-dependence of fast inactivation of WT (blue squares) and P1158S (grey circles) at pH6.4, 7.0, 7.4, and 8.0. (E,F) Show voltage-dependence of fast inactivation of each WT and P1158S. (G) Shows the voltage-dependence of steady-state fast inactivation as a function of pH and fitted with a Hill equation (P1158S) or a flat line (WT). (H) Shows voltage protocol.
Figure 5
Figure 5
Late sodium current. (A–D) Representative normalized current traces of late currents in WT and P1158S at extracellular pH between 6.4 and 8.0. The inset in panel (A) shows the pulse protocol. (E) Shows average late sodium current as a percentage of peak sodium current for WT and P1158S at extracellular pH between 6.4 and 8.0. **Indicate p < 0.01 (P1158S is significantly different from WT).
Figure 6
Figure 6
Use-dependent inactivation. (A–D) Show normalized current decay plotted as a function of time. The inset in panel (A) shows the pulse protocol. (E–H) Show normalized current decay fitted with an exponential curve.
Figure 7
Figure 7
Immunocytochemistry measurements of channel trafficking and expression. (A–C) Representative images of WT, P1158S, and negative control cells taken from the middle confocal plane. (D,E) Show the distribution of channels, measured by using pixel grey value, across the cell. Both WT and P1158S channels are localized around the edges of cells, in proximity to cell membrane (distances 0–4 µm, and 9–14 µm). (F) Average pixel intensity across all confocal images per cell area (n = 3 per condition).
Figure 8
Figure 8
Skeletal muscle action potential modeling. (A) Shows single action potential simulations of WT and P1158S channels at the physiological pH of 7.4. The stimulation pulse of 50 µA/cm2 was applied for 2.0 ms (WT) and 1.2 ms (P1158S). (B) Shows the T-Tubule potassium concentration in WT and P1158S. The potassium concentration is proportional to amount of sodium late currents. (C,D) Show long stimulation pulse from 50–350 ms in WT, and P1158S at all four pH points. (E–H) Show P1158S at pH6.4, 7.0, 7.4, and 8.0.
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