Origin of delayed outward ionic current in charge movement traces from frog skeletal muscle

1. Non-linear membrane ionic current was studied in highly stretched cut frog twitch fibres in a double Vaseline-gap voltage clamp chamber, with the internal solution containing 0.1 mM EGTA and the external solution containing Cl- as the major anion. After the Na+ currents was abolished by TTX in the external solution and the K+ currents were suppressed by external TEA+ and Rb+ and internal Cs+, a delayed outward ionic current with a time course similar to that of the delayed rectifier current was observed during depolarization. 2. The delayed outward ionic current was resistant to 1 mM 3,4-diaminopyridine (3,4-DAP) in the external solution and was unaltered when a fraction of the internal Cs+ was replaced by K+ or Na+, suggesting that the current was not carried by cations flowing through the delayed rectifiers. 3. The delayed outward ionic current was greatly reduced by replacing the external Cl- with CH3SO3-,SO4(2-), glutamate or gluconate, indicating strongly that the current was carried by Cl- flowing through anion channels. The current was also suppressed by 1 mM external 9-anthracenecarboxylic acid (9-ACA). 4. The delayed outward ionic current was reduced by blockers of calcium-dependent Cl- channels, such as SITS and frusemide (furosemide), in a dose- and voltage-dependent manner and by increasing intracellular [EGTA] to 20 mM, suggesting that part of the Cl- current in the muscle fibres could be calcium dependent. 5. The total Cl- current could be dissected into calcium-dependent and calcium-independent components. Each component accounted for roughly half of the total Cl- current. The maximum slope conductance of the calcium-dependent Cl- channels was 60.9 +/- 6.0 microS microF-1 (mean +/- S.E.M., n = 4).