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Open Access Publications from the University of California

Scholarly Works (24 results)

  • Article
  • Peer Reviewed
UCLA Previously Published Works (2021)
Loss-of-function mutations of SCN1A encoding the pore-forming α subunit of the NaV1.1 neuronal sodium channel cause a severe developmental epileptic encephalopathy, Dravet syndrome (DS). In this issue of the JCI, Chen, Luo, Gao, et al. describe a phenocopy for DS in mice deficient for posttranslational conjugation with neural precursor cell expressed, developmentally downregulated 8 (NEDD8) (neddylation), selectively engineered in inhibitory interneurons. Pursuing the possibility that this phenotype is also caused by loss of NaV1.1, Chen, Luo, Gao, and colleagues show that interneuron excitability and GABA release are impaired, NaV1.1 degradation rate is increased with a commensurate decrease of NaV1.1 protein, and NaV1.1 is a substrate for neddylation. These findings establish neddylation as a mechanism for stabilizing NaV1.1 subunits and suggest another pathomechanism for epileptic sodium channelopathy.
    Cover page: Epilepsy channelopathies go neddy: stabilizing NaV1.1 channels by neddylation.
    • Article
    • Peer Reviewed
    UCLA Previously Published Works (2020)
    Calcium gluconate has been empirically administered to hasten recovery of force during an episode of periodic paralysis. By using a genetically engineered mouse model, Uwera and colleagues show that low Ca2+ clearly promotes a loss of force in affected muscle, thereby providing the first evidence for the benefit of maintaining normal Ca2+ levels in this disorder.
      Cover page: A role for external Ca2+ in maintaining muscle contractility in periodic paralysis.
      • Article
      • Peer Reviewed
      UCLA Previously Published Works (2017)
      Cannon reviews new evidence supporting a key role for anomalous inward currents in the etiology of hypokalemic periodic paralysis.
        Cover page: An atypical CaV1.1 mutation reveals a common mechanism for hypokalemic periodic paralysis
        • Article
        • Peer Reviewed
        UCLA Previously Published Works (2015)
        Familial disorders of skeletal muscle excitability were initially described early in the last century and are now known to be caused by mutations of voltage-gated ion channels. The clinical manifestations are often striking, with an inability to relax after voluntary contraction (myotonia) or transient attacks of severe weakness (periodic paralysis). An essential feature of these disorders is fluctuation of symptoms that are strongly impacted by environmental triggers such as exercise, temperature, or serum K(+) levels. These phenomena have intrigued physiologists for decades, and in the past 25 years the molecular lesions underlying these disorders have been identified and mechanistic studies are providing insights for therapeutic strategies of disease modification. These familial disorders of muscle fiber excitability are "channelopathies" caused by mutations of a chloride channel (ClC-1), sodium channel (NaV1.4), calcium channel (CaV1.1), and several potassium channels (Kir2.1, Kir2.6, and Kir3.4). This review provides a synthesis of the mechanistic connections between functional defects of mutant ion channels, their impact on muscle excitability, how these changes cause clinical phenotypes, and approaches toward therapeutics.
          Cover page: Channelopathies of Skeletal Muscle Excitability
          • Article
          • Peer Reviewed
          UCLA Previously Published Works (2016)
          This scientific commentary refers to “Loss-of-function mutations in SCN4A cause severe foetal hypokinesia or ‘classical’ congenital myopathy”, by Zaharieva et al. (doi:10.1093/brain/awv352).
            Cover page: When all is lost…a severe myopathy with hypotonia from sodium channel mutations
            • Thesis
            • Peer Reviewed
            UCLA Electronic Theses and Dissertations (2020)

            The precise control of skeletal muscle excitation, contraction, and relaxation relies upon a highly choreographed sequence of ionic currents in the sarcolemmal membrane. Mutations of ion channels can lead to deleterious changes of ionic currents that impair muscle fiber excitability and thereby cause either weakness or involuntary after-contractions. The work presented here aims to define the functional consequences for newly identified disease mutations in a sodium channel and to establish the mechanism by which the partial loss of a potassium conductance creates susceptibility to episodic attacks of weakness.

            The majority of this work focuses on Andersen-Tawil syndrome, a form of periodic paralysis caused by loss-of-function mutations of the inward rectifying potassium channel Kir2.1. While a genetic basis was established for the disease, it was unknown how this defect creates susceptibility to episodes weakness, or whether as in other forms of periodic paralysis, the attacks are exacerbated by low or high potassium. To investigate this, we generated two different mouse models of ATS, one genetic and the other pharmacologic, that enabled us to explore the mechanistic relationship between the loss of a potassium conductance and the functional deficits in muscle excitation and contraction. We found that loss of muscle force was consistently induced by a high-K challenge, whereas only in the setting of a severely reduced the inward rectifier current (e.g. 6% remaining) did the muscle also exhibit weakness in a low-K challenge. Simulations of a model fiber provided insights on this divergent K-dependence for eliciting weakness and also identified approaches for manipulating conductances, transporters, or pumps to reduce the likelihood of an episode of paralysis. We tested a variety of agents and found that KATP channel openers may provide a means for pharmacologic intervention.

            The remainder of the work characterizes two newly identified mutations in the voltage-gated sodium channel (NaV1.4). The first is a novel loss-of-function mutation found in recessive congenital myopathy plus fluctuating weakness. The second study identified gain-of-function changes for a NaV1.4 mutation associated with an unusually early onset of myotonia with congenital anomalies, and defined a new syndrome, myotonic myopathy with secondary joint and bone anomalies.

              Cover page: Mouse Models of Periodic Paralysis in Andersen Tawil Syndrome and Functional Studies of Novel Nav1.4 mutations in Myotonia & Myopathy
              • Article
              • Peer Reviewed
              UCLA Previously Published Works (2000)
              Skeletal muscle dihydropyridine (DHP) receptors function both as voltage-activated Ca(2+) channels and as voltage sensors for coupling membrane depolarization to release of Ca(2+) from the sarcoplasmic reticulum. In skeletal muscle, the principal or alpha(1S) subunit occurs in full-length ( approximately 10% of total) and post-transcriptionally truncated ( approximately 90%) forms, which has raised the possibility that the two functional roles are subserved by DHP receptors comprised of different sized alpha(1S) subunits. We tested the functional properties of each form by injecting oocytes with cRNAs coding for full-length (alpha(1S)) or truncated (alpha(1SDeltaC)) alpha subunits. Both translation products were expressed in the membrane, as evidenced by increases in the gating charge (Q(max) 80-150 pC). Thus, oocytes provide a robust expression system for the study of gating charge movement in alpha(1S), unencumbered by contributions from other voltage-gated channels or the complexities of the transverse tubules. As in recordings from skeletal muscle, for heterologously expressed channels the peak inward Ba(2+) currents were small relative to Q(max). The truncated alpha(1SDeltaC) protein, however, supported much larger ionic currents than the full-length product. These data raise the possibility that DHP receptors containing the more abundant, truncated form of the alpha(1S) subunit conduct the majority of the L-type Ca(2+) current in skeletal muscle. Our data also suggest that the carboxyl terminus of the alpha(1S) subunit modulates the coupling between charge movement and channel opening.
                Cover page: Cooh-Terminal Truncated Alpha1S Subunits Conduct Current Better than Full-Length Dihydropyridine Receptors
                • Article
                • Peer Reviewed
                UCLA Previously Published Works (2014)
                Missense mutations at arginine residues in the S4 voltage-sensor domains of NaV1.4 are an established cause of hypokalemic periodic paralysis, an inherited disorder of skeletal muscle involving recurrent episodes of weakness in conjunction with low serum K(+). Expression studies in oocytes have revealed anomalous, hyperpolarization-activated gating pore currents in mutant channels. This aberrant gating pore conductance creates a small inward current at the resting potential that is thought to contribute to susceptibility to depolarization in low K(+) during attacks of weakness. A critical component of this hypothesis is the magnitude of the gating pore conductance relative to other conductances that are active at the resting potential in mammalian muscle: large enough to favor episodes of paradoxical depolarization in low K(+), yet not so large as to permanently depolarize the fiber. To improve the estimate of the specific conductance for the gating pore in affected muscle, we sequentially measured Na(+) current through the channel pore, gating pore current, and gating charge displacement in oocytes expressing R669H, R672G, or wild-type NaV1.4 channels. The relative conductance of the gating pore to that of the pore domain pathway for Na(+) was 0.03%, which implies a specific conductance in muscle from heterozygous patients of ∼ 10 µS/cm(2) or 1% of the total resting conductance. Unexpectedly, our data also revealed a substantial decoupling between gating charge displacement and peak Na(+) current for both R669H and R672G mutant channels. This decoupling predicts a reduced Na(+) current density in affected muscle, consistent with the observations that the maximal dV/dt and peak amplitude of the action potential are reduced in fibers from patients with R672G and in a knock-in mouse model of R669H. The defective coupling between gating charge displacement and channel activation identifies a previously unappreciated mechanism that contributes to the reduced excitability of affected fibers seen with these mutations and possibly with other R/X mutations of S4 of NaV, CaV, and KV channels associated with human disease.
                  Cover page: Disrupted coupling of gating charge displacement to Na+ current activation for DIIS4 mutations in hypokalemic periodic paralysis
                  • Article
                  • Peer Reviewed
                  UCLA Previously Published Works (2007)
                  The heritable muscle disorder hypokalemic periodic paralysis (HypoPP) is characterized by attacks of flaccid weakness, brought on by sustained sarcolemmal depolarization. HypoPP is genetically linked to missense mutations at charged residues in the S4 voltage-sensing segments of either CaV1.1 (the skeletal muscle L-type Ca(2+) channel) or NaV1.4 (the skeletal muscle voltage-gated Na(+) channel). Although these mutations alter the gating of both channels, these functional defects have proven insufficient to explain the sarcolemmal depolarization in affected muscle. Recent insight into the topology of the S4 voltage-sensing domain has aroused interest in an alternative pathomechanism, wherein HypoPP mutations might generate an aberrant ionic leak conductance by unblocking the putative aqueous crevice ("gating-pore") in which the S4 segment resides. We tested the rat isoform of NaV1.4 harboring the HypoPP mutation R663H (human R669H ortholog) at the outermost arginine of S4 in domain II for a gating-pore conductance. We found that the mutation R663H permits transmembrane permeation of protons, but not larger cations, similar to the conductance displayed by histidine substitution at Shaker K(+) channel S4 sites. These results are consistent with the notion that the outermost charged residue in the DIIS4 segment is simultaneously accessible to the cytoplasmic and extracellular spaces when the voltage sensor is positioned inwardly. The predicted magnitude of this proton leak in mature skeletal muscle is small relative to the resting K(+) and Cl(-) conductances, and is thus not likely to fully account for the aberrant sarcolemmal depolarization underlying the paralytic attacks. Rather, it is possible that a sustained proton leak may contribute to instability of V(REST) indirectly, for instance, by interfering with intracellular pH homeostasis.
                    Cover page: A Na+ Channel Mutation Linked to Hypokalemic Periodic Paralysis Exposes a Proton-selective Gating Pore
                    • Article
                    • Peer Reviewed
                    UCLA Previously Published Works (2002)
                    Slow inactivation of voltage-gated Na channels is kinetically and structurally distinct from fast inactivation. Whereas structures that participate in fast inactivation are well described and include the cytoplasmic III-IV linker, the nature and location of the slow inactivation gating mechanism remains poorly understood. Several lines of evidence suggest that the pore regions (P-regions) are important contributors to slow inactivation gating. This has led to the proposal that a collapse of the pore impedes Na current during slow inactivation. We sought to determine whether such a slow inactivation-coupled conformational change could be detected in the outer pore. To accomplish this, we used a rapid perfusion technique to measure reaction rates between cysteine-substituted side chains lining the aqueous pore and the charged sulfhydryl-modifying reagent MTS-ET. A pattern of incrementally slower reaction rates was observed at substituted sites at increasing depth in the pore. We found no state-dependent change in modification rates of P-region residues located in all four domains, and thus no change in aqueous accessibility, between slow- and nonslow-inactivated states. In domains I and IV, it was possible to measure modification rates at residues adjacent to the narrow DEKA selectivity filter (Y401C and G1530C), and yet no change was observed in accessibility in either slow- or nonslow-inactivated states. We interpret these results as evidence that the outer mouth of the Na pore remains open while the channel is slow inactivated.
                      Cover page: Slow Inactivation Does Not Block the Aqueous Accessibility to the Outer Pore of Voltage-gated Na Channels
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