3. In the absence of self-maintained activity both myotonic fibres and normal fibres in chloride-free Ringer show an after-depolarization which is proportional to the number of driven impulses. The half-time for the decay of this after-potential is about 0 5 sec.4. Tubular potassium accumulation resulting from the initially driven activity and the known low chloride conductance of myotonic muscle fibres appear to account for the initiation of the myotonic discharge.

SUMMARY1. Cable parameters, component conductances, excitability and membrane potentials in isolated external intercostal fibre bundles at 380 C from normal and myotonic goats were measured in normal and low-chloride Ringer, and in the presence of monocarboxylic aromatic acids that produce myotonic responses in mammalian muscle.2. The mean resting chloride conductance in gzmho/cm2 of myotonic fibres (range 0-147) was significantly less than that of normal fibres (range 376-951). The mean resting potassium conductance was higher in myotonic fibres (range 123-285) than in normal fibres (range 44-132).Potassium conductance increased about 10 ,smho/cm2 per mV increase in absolute resting potential.3. In normal fibres in normal Ringer 3-chloro-2,5,6-trimethylbenzoic acid; 5,6-dihydro-5,5-dimethyl-7-carboxybenz[c]acridine; phenanthrene-9-carboxylic acid; and anthracene-9-carboxylic acid at 10-5-104 M decreased membrane conductance without consistently changing diameter or capacitance. In low-chloride Ringer 3-chloro-2,5,6-trimethylbenzoic acid (5 x 10-5 M) increased potassium conductance in myotonic and normal fibres. It is concluded that these compounds block chloride conductance.4. The carboxylic acids produced myotonia in normal fibres similar to that in untreated myotonic fibres.5. Anthracene-9-carboxylic acid intravenously (8 mg/kg) in normal goats produced acutely a condition resembling myotonia congenita. The carboxylic acids produced no myotonic effects in frog muscle.

The single-channel blocking kinetics of tetrodotoxin (TTX), saxitoxin (STX), and several STX derivatives were measured for various Na-channel subtypes incorporated into planar lipid bilayers in the presence of batrachotoxin. The subtypes studied include Na channels from rat skeletal muscle and rat brain, which have high affinity for TTX/STX, and Na channels from denervated rat skeletal muscle and canine heart, which have about 20-60-fold lower affinity for these toxins at 22 degrees C. The equilibrium dissociation constant of toxin binding is an exponential function of voltage (e-fold per 40 mV) in the range of -60 to +60 mV. This voltage dependence is similar for all channel subtypes and toxins, indicating that this property is a conserved feature of channel function for batrachotoxin-activated channels. The decrease in binding affinity for TTX and STX in low-affinity subtypes is due to a 3-9-fold decrease in the association rate constant and a 4-8-fold increase in the dissociation rate constant. For a series of STX derivatives, the association rate constant for toxin binding is approximately an exponential function of net toxin charge in membranes of neutral lipids, implying that there is a negative surface potential due to fixed negative charges in the vicinity of the toxin receptor. The magnitude of this surface potential (-35 to -43 mV at 0.2 M NaCl) is similar for both high- and low-affinity subtypes, suggesting that the lower association rate of toxin binding to toxin-insensitive subtypes is not due to decreased surface charge but rather to a slower protein conformational step. The increased rates of toxin dissociation from insensitive subtypes can be attributed to the loss of a few specific bonding interactions in the binding site such as loss of a hydrogen bond with the N-1 hydroxyl group of neosaxitoxin, which contributes about 1 kcal/mol of intrinsic binding energy.

1. In preparations of about 200 fibres each from thirty‐nine biopsies of external intercostal muscle taken from nine myotonic and six nonmyotonic goats, cable properties were determined at 38° C for individual fibres with a pair of intracellular micro‐electrodes. 2. In each preparation the mean fibre dimensions, determined histologically and corrected for shrinkage, were used to calculate the mean membrane resistance, Rm, fibre capacitance, Ct, and myoplasmic resistivity, Ri. In the 124 nonmyotonic fibres the mean values were: Rm, 1897 Ω.cm2, Ct, 4·1 μF/cm2, and Ri, 112 Ω.cm. In 151 myotonic fibres Rm was 5589 Ω.cm2, Ct, 4·4 μF/cm2, and Ri, 103 Ω.cm. 3. Conductance of the fibre core times unit length increased with cross‐sectional area, and fibre capacitance per unit length increased with perimeter. There was little correlation of membrane resistance per unit length of fibre with either fibre perimeter or resting potential. 4. The principal abnormality of cable properties in the myotonic fibre is its threefold higher membrane resistance, which accounts for its decreased electrical current rheobase.

No abstract

External intercostal muscle biopsies from normal and congenitally myotonic goats were studied in vitro at 30 degrees 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-beta-phorbol-12,13-dibutyrate. (0.1-2.0 microM) 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 microM). The "inactive" 4-alpha-phorbol-12,13-didecanoate had no effect at 50 microM, whereas the "active" 4-beta isomer blocked 41% Gcl at 1 microM. 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 fiber does not appear to be due to excessive activity of PKC.

A B S T R A C T In isolated fiber bundles of external intercostal muscle from each of 13 normal volunteers and each of 6 patients with myotonia congenita, some or all of the following were measured: concentrations of Na+, K+, and Cl-, extracellular volume, water content, K+ efflux, fiber size, fiber cable parameters, and fiber resting potentials.Muscle from patients with myotonia congenita differed significantly (0.001 < P< 0.025) with respect to the following mean values (myotonia congenita vs. normal): the membrane resistance was greater (5729 vs. 2619 U. cm2), the internal resistivity was less (75.0 vs. 123.2 Q-cm), the water content was less (788.2 vs. 808.2 ml/kg wet weight), and the mean resting potential was greater (68 vs. 61 mv).No significant differences were found with respect to the following variables: K+ content (73.5 vs. 66.7 mEq/kg wet weight) and the calculated intracellular K+ concentration (215 vs. 191 mEq/liter fiber water), fiber capacitance (5.90 vs. 5.15 Mf/cm2), Na+ content (97.7 vs. 94.1 mEq/kg wet weight), 74.7 mEq/kg wet weight), mannitol extracellular volume (45.1 vs. 46.6 cc/100 g wet weight), and K+ efflux (23.2 vs. 21.5 moles X 10-12 cm-2.sec-1).These abnormalities of skeletal muscle in human myotonia congenita are like those of skeletal muscle in goats with hereditary myotonia. We tentatively conclude that a decreased Cl-permeability accounts for some of the abnormal electrical properties of skeletal muscle in myotonia congenita. We now report findings in isolated external intercostal muscle from patients with myotonia congenita and from normal volunteers. We have compared our human results with similar data we obtained in isolated external intercostal muscle from the goat (1, 2). METHODS Normal volunteers. Our normals are 13 males aged 21-33 yr with no evidence of neuropathy or myopathy. One (R. G.) had diabetes mellitus; another (T. P.) had probable pulmonary sarcoidosis (minimal pulmonary fibrosis on chest X-ray and scalene node biopsy positive for noncaseating granuloma). At the time of muscle biopsy, T. P. had no symptoms and his chest X-ray was clearer (without therapy) than 1 yr earlier. No volunteer had taken any medication for at least 5 days before the external intercostal muscle biopsy except for the patient with diabetes, who received insulin up to and including the day before biopsy. The remaining 11 subjects were healthy and the results of laboratory studies (chest X-ray, electrocardiogram, hemoglobin, hematocrit, white blood cell count and differential, urinalysis, serum urea nitrogen, fasting blood sugar, Wasserman, serum creatinine, serum ions [Na+, K+, Cl-], serum Ca and Mg, total serum proteins, serum alkaline phosphatase, serum glutamic oxalacetic transaminase, and creatinine phosphokinase) were normal. Biopsy techniques. Biopsies of the anterior border of external intercostal muscle (2.5-5 cm in length) were obtained under local anesthesia from the right eighth intercostal space. Preoperative medications were pentobarbital (from 100 to 200 mg), and morphin...

In isolated bundles of external intercostal muscle from normal goats and goats with hereditary myotonia the following were determined: concentrations and unidirectional fluxes of Na+, K+, and C-, extracellular volume, water content, fiber geometry, and core-conductor constants. No significant difference between the two groups of preparations was found with respect to distribution of fiber size, intracellular concentrations of Na+ or Cl-, fiber water, resting membrane potential, or overshoot of action potential. The intracellular C1-concentration in both groups of preparations was 4 to 7 times that expected if C1-were distributed passively between intracellular and extracellular water. The membrane permeability to K (Ps) calculated from efflux data was (a) at 38°C, 0.365 X 10 -6 cm sec -1 for normal and 0.492 X 10-6 for myotonic muscle, and (b) at 250°C, 0.219 X 10-6 for normal and 0.199 X 10-6 for myotonic muscle. From C1-washout curves of normal muscle usually only three exponential functions could be extracted, but in every experiment with myotonic muscle there was an additional, intermediate component. From these data Pcl could be calculated; it was 0.413 X 10-6 cm sec-1 for myotonic fibers and was > 0.815 X 10-6 cm sec-1 for normal fibers. The resting membrane resistance of myotonic fibers was 4 to 6 times greater than that of normal fibers.