The Journal of Membrane Biology

, Volume 123, Issue 3, pp 223–233 | Cite as

Response of chloride efflux from skeletal muscle ofRana pipiens to changes of temperature and membrane potential and diethylpyrocarbonate treatment

  • Bruce C. Spalding
  • Patricia Taber
  • John G. Swift
  • Paul Horowicz


Efflux of36Cl from frog sartorius muscles equilibrated in two depolarizing solutions was measured. Cl efflux consists of a component present at low pH and a pH-dependent component which increases as external pH increases.

For temperatures between 0 and 20°C, the measured activation energy is 7.5 kcal/mol for Cl efflux at pH 5 and 12.6 kcal/mol for the pH-dependent Cl efflux. The pH-dependent Cl efflux can be described by the relationu=1/(1+10n(pK a -pH)), whereu is the Cl efflux increment obtained on stepping from pH 5 to the test pH, normalized with respect to the increment obtained on stepping from pH 5 to 8.5 or 9.0. For muscles equilibrated in solutions containing 150mm KCl plus 120mm NaCl (internal potential about −15 mV), the apparent pK a is 6.5 at both 0 and 20°C, andn=2.5 for 0°C and 1.5 for 20°C. For muscles equilibrated in solutions containing 7.5mm KCl plus 120mm NaCl (internal potential about −65 mV), the apparent pK a at 0°C is 6.9 andn is 1.5. The voltage dependence of the apparent pK a suggests that the critical pH-sensitive moiety producing the pH-dependent Cl efflux is sensitive to the membrane electric field, while the insensitivity to temperature suggests that the apparent heat of ionization of this moiety is zero. The fact thatn is greater than 1 suggests that cooperativity between pH-sensitive moieties is involved in determining the Cl efflux increment on raising external pH.

The histidine-modifying reagent diethylpyrocarbonate (DEPC) applied at pH 6 reduces the pH-dependent Cl efflux according to the relation, efflux=exp(−k·[DEPC]·t), wheret is the exposure time (min) to DEPC at a prepared initial concentration of [DEPC] (mm). At 17°C,k−1=188mm·min. For temperatures between 10 and 23°C,k has an apparent Q10 of 2.5. The Cl efflux inhibitor SCN at a concentration of 20mm substantially retards the reduction of the pH-dependent Cl efflux by DEPC. The findings that the apparent pK a is 6.5 in depolarized muscles, that DEPC eliminates the pH-dependent Cl efflux, and that this action is retarded by SCN supports the notion that protonation of histidine groups associated with Cl channels is the controlling reaction for the pH-dependent Cl efflux.

Key Words

skeletal muscle Cl efflux Cl channel pH muscle membrane temperature diethylpyrocarbonate 


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Copyright information

© Springer-Verlag New York Inc. 1991

Authors and Affiliations

  • Bruce C. Spalding
    • 1
  • Patricia Taber
    • 1
  • John G. Swift
    • 1
  • Paul Horowicz
    • 1
  1. 1.Department of Physiology, School of Medicine and DentistryUniversity of RochesterRochester

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