Résumé :
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Communication n° 162. Introduction : The mechanisms underlying the high sarcolemmal permeability to Ca2+ and the perturbed Ca2+ homeostasis in muscle fibers of dystrophic mdx mouse are still debated. Objective. To perform a biophysical and pharmacological characterization of the channel potentially involved in Ca2+ deregulation in dystrophic fibers. Methods : Cell-attached patch-clamp recordings on enzimatically-dissociated native FDB muscle fibers from 8-10 week old male wt and mdx mice. Results : We confirmed a higher occurrence and density of a voltage-insensitive cationic current in mdx (~92 %) vs wt animals (~60 %). In both genotypes, the current was inhibited by Gd3+ and ruthenium red, as expected for a TRP-like current. In wt fibers, the application of insulin-like growth factor 1 (IGF-1, 3.3 nM) induced a channel activity or enhanced the pre-existing one without changing the single channel conductance (~ 30 pS). This effect was slowly reversible upon IGF-1 washout; however wt fibers previously exposed to IGF-1 showed a higher occurrence of active patches in drug-free solution. The IGF-1 effect was prevented by a prior incubation with wortmannin, suggesting the involvement of the PI3-kinase. Interestingly, application of IGF-1 on mdx muscle fibers did not further enhance channel activity. This difference between wt and mdx was confirmed on the sarcolemmal permeability of intact tendon-to-tendon fibers from EDL, measured by the Mn2+-quenching technique. Either IGF-1-induced and spontaneously active current observed in wt and/or mdx fibers were silenced by a cAMP-dependent mechanism, as shown by using db-cAMP or pentoxifylline, a phosphodiesterase inhibitor. Conclusion : IGF-1 increases, via PI3-kinase, the number of the channels in the sarcolemma, and the loss of integrity of cytoskeleton, as in dystrophin-deficient fibers, leads to the deregulation of this mechanism and abnormal channel activity. Channel regulation downstream the above mechanism may open possible therapeutic approaches. (Supported by Telethon-Italy # 1150).
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