Cholesterol modulates the recruitment of Kv1.5 channels from Rab11-associated recycling endosome in native atrial myocytes

doi:10.1073/pnas.0902809106

Cholesterol modulates the recruitment of Kv1.5 channels from Rab11-associated recycling endosome in native atrial myocytes

^aInstitut National de la Santé et de la Recherche Médicale, Unité Mixte de Recherche Scientifique-956, 75013 Paris, France;
^bUniversité Pierre et Marie Curie, Paris-6, Unité Mixte de Recherche Scientifique-956, 75013 Paris, France;
^cPlate-forme imagerie cellulaire IFR14, 75013 Paris, France; and
^dDepartment of Anesthesiology, Pharmacology and Therapeutics, University of British Columbia, Vancouver, BC, Canada V6T 1Z3

Edited by Lily Y. Jan, University of California, San Francisco, CA, and approved June 19, 2009 (received for review March 17, 2009)

Abstract

Cholesterol is an important determinant of cardiac electrical properties. However, underlying mechanisms are still poorly understood. Here, we examine the hypothesis that cholesterol modulates the turnover of voltage-gated potassium channels based on previous observations showing that depletion of membrane cholesterol increases the atrial repolarizing current I_Kur. Whole-cell currents and single-channel activity were recorded in rat adult atrial myocytes (AAM) or after transduction with hKv1.5-EGFP. Channel mobility and expression were studied using fluorescence recovery after photobleaching (FRAP) and 3-dimensional microscopy. In both native and transduced-AAMs, the cholesterol-depleting agent MβCD induced a delayed (≈7 min) increase in I_Kur; the cholesterol donor LDL had an opposite effect. Single-channel recordings revealed an increased number of active Kv1.5 channels upon MβCD application. Whole-cell recordings indicated that this increase was not dependent on new synthesis but on trafficking of existing pools of intracellular channels whose exocytosis could be blocked by both N-ethylmaleimide and nonhydrolyzable GTP analogues. Rab11 was found to coimmunoprecipitate with hKv1.5-EGFP channels and transfection with Rab11 dominant negative (DN) but not Rab4 DN prevented the MβCD-induced I_Kur increase. Three-dimensional microscopy showed a decrease in colocalization of Kv1.5 and Rab11 in MβCD-treated AAM. These results suggest that cholesterol regulates Kv1.5 channel expression by modulating its trafficking through the Rab11-associated recycling endosome. Therefore, this compartment provides a submembrane pool of channels readily available for recruitment into the sarcolemma of myocytes. This process could be a major mechanism for the tuning of cardiac electrical properties and might contribute to the understanding of cardiac effects of lipid-lowering drugs.

Footnotes

¹To whom correspondence should be addressed at:
Unité Mixte de Recherche Scientifique-956, Faculté de Médecine Pierre et Marie Curie, Paris-6, 91 boulevard de l'Hôpital, 75013 Paris, France.
E-mail: stephane.hatem{at}upmc.fr
Author contributions: E.B., A.C., and S.N.H. designed research; E.B., S.E.-H., G.D., and A.C. performed research; A.D., J.E., and D.F. contributed new reagents/analytic tools; E.B., S.E.-H., A.C., and S.N.H. analyzed data; and E.B., J.E., D.F., A.C., and S.N.H. wrote the paper.
The authors declare no conflict of interest.
This article is a PNAS Direct Submission.
This article contains supporting information online at www.pnas.org/cgi/content/full/0902809106/DCSupplemental.