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Modulation of the gating of CIC-1 by S-(-) 2-(4-chlorophenoxy) propionic acid
E C Aromataris1, D S Astill, G Y Rychkov
1Department of Physiology, University of Adelaide, South Australia, Australia.
Abstract:
1. Using whole-cell patch-clamping and Sf-9 cells expressing the rat skeletal muscle chloride channel, rCIC-1, the cellular mechanism responsible for the myotonic side effects of clofibrate derivatives was examined. 2. RS-(+/-) 2-(4-chlorophenoxy)propionic acid (RS-(+/-) CPP) and its S-(-) enantiomer produced pronounced effects on CIC-1 gating. Both compounds caused the channels to deactivate more rapidly at hyperpolarizing potentials, which showed as a decrease in the time constants of both the fast and slow deactivating components of the whole cell currents. Both compounds also produced a concentration-dependent shift in the voltage dependence of channel apparent open probability to more depolarizing potentials, with an EC50 of 0.79 and 0.21 mM for the racemate and S-(-) enantiomer respectively. R-(+) CPP at similar concentrations had no effect on gating. RS-(+/-) CPP did not block the passage of Cl- through the pore of rCIC-1. 3. CIC-1 is gated by Cl- binding to a site within an access channel and S-(-) CPP alters gating of the channel by decreasing the affinity of this binding site for Cl-. Comparison of the EC50 for RS-(+/-) CPP and S-(-) CPP indicates that R-(+) CPP can compete with the S-(-) enantiomer for the site but that it is without biological activity. 4. RS-(+/-) CPP produced the same effect on rCIC-1 gating when added to the interior of the cell and in the extracellular solution. 5. S-(-) CPP modulates the gating of CIC-1 to decrease the membrane Cl- conductance (GCl), which would account for the myotonic side effects of clofibrate and its derivatives.
Insights
The S-(-) enantiomer of RS-(+/-) 2-(4-chlorophenoxy)propionic acid (CPP) alters rat skeletal muscle chloride channel 1 (rCIC-1) gating, decreasing chloride conductance and explaining myotonic side effects of clofibrate derivatives.
Area of Science:
- Pharmacology
- Molecular Biology
- Biophysics
Background:
- Myotonic side effects are associated with clofibrate derivatives.
- The cellular mechanisms underlying these side effects require elucidation.
Purpose of the Study:
- To investigate the mechanism by which clofibrate derivatives cause myotonic side effects.
- To examine the effects of RS-(+/-) 2-(4-chlorophenoxy)propionic acid (CPP) and its enantiomers on the rat skeletal muscle chloride channel rCIC-1.
Main Methods:
- Whole-cell patch-clamping technique was employed.
- Sf-9 cells expressing the rat skeletal muscle chloride channel, rCIC-1, were utilized.
- The effects of RS-(+/-) CPP and its S-(-) enantiomer on rCIC-1 gating kinetics and voltage dependence were analyzed.
Main Results:
- The S-(-) enantiomer of CPP significantly altered rCIC-1 gating, causing faster deactivation at hyperpolarizing potentials.
- Both RS-(+/-) CPP and S-(-) CPP induced a concentration-dependent shift in voltage dependence, with S-(-) CPP being more potent (EC50 = 0.21 mM).
- The R-(+) enantiomer had no effect, and RS-(+/-) CPP did not block Cl- passage, indicating a modulatory effect on gating.
Conclusions:
- S-(-) CPP decreases the affinity of the Cl- binding site within the rCIC-1 channel, altering its gating.
- This modulation reduces membrane chloride conductance (GCl), explaining the myotonic side effects observed with clofibrate derivatives.