Shirley H. Bryant

Chloride channel regulation in the skeletal muscle of normal and myotonic goats

External intercostal muscle biopsies from normal and congenitally myotonic goats were studied in vitro at 30° C using a two-microelectrode square-pulse cable analysis assisted by computer. The resting chloride conductance (Gcl) was estimated from the difference between the mean membrane conductance in chloride-containing and chloride-free bathing media. The protein kinase C (PKC) activator, 4-β-phorbol-12,13-dibutyrate, (0.1–2.0 μM) blocks a maximum of 76% of Gcl in normal goat fibers and induces myotonic hyperexcitability similar to that of congenitally myotonic goat fibers. The Gcl block was partially antagonized by pretreatment with the PKC inhibitor, staurosporine (10 μM). The “inactive” 4-αphorbol-12, 13,didecanoate had no effect at 50 μM, whereas the “active” 4-β isomer blocked 41% Gcl at 1 μM. The nearly absent Gcl of congenitally myotonic goat fibers was not restored by treatment with high concentrations of the PKC inhibitors staurosporine, 1-(5-isoquinolinesulfonyl)-2-methylpiperazine (H7), or tetrahydropapaveralone (THP). Also, forskolin and cholera toxin, which may increase cyclic adenosine monophosphate (cAMP) levels, or the R(+) clofibric acid enantiomers and taurine, which increase Gcl in normal fibers, were also unable to restore Gcl in myotonic goat fibers. The data suggest that PKC may be a chloride channel regulator in normal goat skeletal muscle fibers, however the molecular defect of congenitally myotonic fibers does not appear to be due to excessive activity of PKC.

Enantiomers of clofibric acid analogs have opposite actions on rat skeletal muscle chloride channels

The S-(−) isomers of a series of clofibric acid analogs produced only a block of chloride conductance of rat skeletal muscle fibers with increasing concentrations until block was nearly complete. The R-(+) isomers, on the other hand, at low concentrations increased chloride conductance by as much as 9% to 39% and at higher concentrations decreased chloride conductance, but never by more than 27% of the control value. The actions of the enantiomeric pairs to either produce or inhibit myotonic excitability paralleled their ability to block or increase chloride conductance, respectively.

The Toxic Effects of Clofibrate and Its Metabolite on Mammalian Skeletal Muscle: An Electrophysiologigal Study

Membrane electrical parameters of rat extensor digitorum longus muscles were analysed in vitro, with intracellular microelectrodes, after chronic treatment with clofibrate or after in vitro application of chlorophenoxyisobutyric acid, the in vivo metabolite of clofibrate. Clofibrate caused only a 38% decrease in the membrane chloride conductance while its metabolite induced a large decrease of chloride conductance and drastic changes in the excitability characteristics of the muscles sampled. The capability of clofibrate to induce two types of myotonia is discussed.

Antimyotonic effects of tocainide enantiomers on skeletal muscle fibers of congenitally myotonic goats.

Tocainide is effective in the symptomatic treatment of myotonic syndromes for its ability to reduce the high frequency discharges of action potentials typical of the disease, by blocking voltage-gated sodium channels. However, its use is restricted by serious side effects. In spite of its chiral structure, tocainide is clinically used as a racemic mixture. Since the optical isomers may differ in their efficacy and toxicity, the present study was aimed at evaluating the antimyotonic activity of the pure R(-) and S(+) enantiomers of tocainide, on the abnormal membrane hyperexcitability of external intercostal muscle fibers of congenitally myotonic goats. The excitability parameters were recorded in vitro by means of the standard two-microelectrode current-clamp technique before and after the addition of the compounds. The R(-) enantiomer of tocainide at concentrations as low as 10 microM potently counteracted the abnormal excitability of myotonic fibers, by increasing the threshold current, and decreasing the latency of the action potential and firing capability. Also, this concentration of R-(-) tocainide almost completely abolished the abnormal spontaneous electrical activity occurring in about 70-80% of the myotonic fiber. The S(+) enantiomer was remarkably less potent since up to 100 microM did not restore the normal excitability pattern. The results show that most of the antimyotonic activity of tocainide resides in the R(-) enantiomer suggesting that its clinical use may allow a significant reduction of the doses and possibly of the side effects.

Aconitine-induced repetitive firing in frog skeletal muscle and the effect on cable properties

Abstract The repetitive firing in curarized frog sartorius muscle caused by aconitine was found to be associated with a dose-dependent membrane depolarization and a decrease in membrane resistance (R m ). Shortly after aconitine treatment, when repetitive discharges began to occur spontaneously in a few fibers (the pre-burst state), the muscle fibers were hypersensitive to mechanical and electrical stimulation. After a repetitive discharge (the post-burst state), the membrane potential fell to the 40–50 mV range. Cable measurements demonstrated that R m decreased by 27% in the pre-burst state and by 47% in the post-burst state. Substitution of choline for sodium in the post-burst state restored the membrane potential to normal and raised the R m toward the control level. It is proposed that aconitines production of repetitive discharges in skeletal muscle is partly the result of maintaining the sodium conductance g Na at a higher-than-normal level once excitation has been initiated.

Enantiomeric effects on excitation-contraction coupling in frog skeletal muscle by a chiral phenoxy carboxylic acid.

Aromatic monocarboxylic acids are known to significantly potentiate the mechanical response of skeletal muscle fibers. In this study we investigated the effects of enantiomers of 2-(4-chlorophenoxy)propionic acid, chemically one of the simplest aromatic monocarboxylic acids with chiral properties, on mechanical threshold and charge movement in frog skeletal muscle. The R(+), but not the S(-), enantiomer lowered rheobase mechanical threshold and shifted charge movement to more negative potentials. The R(+) enantiomer also significantly slowed charge movement kinetics, with pronounced delays of the OFF charge transitions. These effects required high temperature for their production. The stereospecific actions of the R(+) enantiomer are interpreted in terms of a specific interaction of this compound at an anion-sensitive site involved in excitation-contraction coupling, most likely on the dihydropryidine-sensitive voltage sensor in the T-system.

Effect of Taurine on Chloride Conductance and Excitability of Rat Skeletal Muscle Fibers

Taurine is found in mammalian skeletal muscle fibers and all other excitable tissues (17), but its function is not well understood (16). Abnormal taurine contents whether pathologically or experimentally induced are often associated with changes of excitability in heart (9), nerve (8,21), and retina (4). The taurine content is increased in skeletal muscle from the dystrophic chicken (23).

Effects of Veratridine on Na and Ca Currents in Frog Skeletal Muscle

1. Voltage-clamp experiments were performed to determine the effects of veratridine on Na and Ca currents in frog skeletal muscle fibres. 2. Veratridine (1 microM) did not affect the kinetics of the fast Na current but it did induce a slowly inactivating tetrodotoxin-sensitive inward current that was apparent after Na current inactivation. This slow current had a peak amplitude of 6.7 +/- 0.7 microA/cm2 at -20 mV and decayed monoexponentially with a time constant of 606 +/- 77 ms. 3. The slow current had a voltage-dependence for activation that was similar to that of the fast Na current. Single depolarizing prepulses that induced complete inactivation of the fast Na channels, prevented development of the slow current. Trains of brief depolarizations at increasing frequencies increased the amplitude of the slow current. These results suggest that the slow current may be mediated by veratridine modified Na channels that must be in the open position. 4. The low concentration of veratridine (1 microM) did not affect the Ca current, while 100 microM veratridine reversibly suppressed the Ca current and shifted its peak current-voltage relation towards more negative potentials. Thus, veratridine appears not to be a selective fast Na channel modifier as it may also alter Ca channel gating properties in skeletal muscle fibres.

Effects of veratrine on ion currents in single rabbit cardiomyocytes

1. Voltage-clamp experiments were performed to determine the effects of veratrine (1 microgram/ml) on Na and K currents in isolated rabbit ventricular cardiomyocytes. 2. Veratrine did not affect the inward rectifier K current, increased the inactivation time constant of the transient outward current (I(to)) and induced a slowly decaying inward current component (Iv), which was sensitive to tetrodotoxin. 3. Inactivation of fast Na channels by application of short depolarizing prepulses to potentials between -90 and -50 mV prevented the development of Iv.Iv decayed biexponentially with time constants equal to 139 +/- 9.0 ms and 776 +/- 47 ms. The net amplitude of Iv and the time constants for its rapidly and slowly inactivating components were little affected by trains of conditioning prepulses to 0 mV. The contributions, however, of the fast and slow components to the net current were significantly altered by repetitive depolarizations. 4. These components of Iv are likely due to modification of open cardiac Na channels by veratrum alkaloids.