Characterization of a Cardiac Transient Outward Current

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EXCEED THE SPACE PROVIDED. The transient outward K+ current, Ito, is a major contributor to atrial and ventricular repolarization and is thought to be an important but unexplored therapeutic target for arrhythmia suppression. However, there are multipledistinctcardiac Ito phenotypes with heterogeneous expression patterns in the heart. There are also multiple K+ channel (X subunits (Kvl.4, Kv4.2, and Kv4.3) that serve as potential molecular substrates for native Ito phenotypes. We demonstrated heterogeneity of Kvl .4 and Kv4.2/4.3 CC subunit expression of 2 distinct Ito phenotypes (a rapidly recovering Ito which correlates with Kv4.2/4.3 expression, and a slowly recovering Ito in LV endocardial myocytes which correlates with Kvl.4 expression). Several important scientific and clinically relevant questions remain. Do other channels and/or ancillary subunits contribute to this current and/or form the basis forthe heterogeneity of distinct cardiac Ito phenotypes? Which members of this class are present in heart and what is their regional distribution? What is the role of Ito in different atrial myocyte types? Why do heterologously expressed Kv4.2/4.3 a subunits fail to reconstitute the Ito with rapid recovery kinetics? To address these important questions, we have focused on a class of Ca2+-binding Kv channel interacting proteins (KCMPs) that are integral components of native Kv4.x channel complexes. KCMPs haverecently been discovered to accelerate the recovery kinetics of heterologously expressed Ky4.x pc subunits. To determine if KChlPs are the missing modulatory factors needed to fully reconstitute the rapidly recovering !, phenotype, we will use a combination of molecularbiological, immunofluorescent (IF) localization, insitu hybridization, biochemical,and biophysical techniques to elucidate the mechanisms by which KChlPs modulate the gating characteristics of both expressed Kv4.2/4.3 ¿ subunits (Xenopus oocytes, HEK 293 cells) and native Ito phenotypes in ferret myocytes. We will define the regional distribution patterns of Kvl .4, Kv4.2, Kv4.3, and KChlPs in ferret ventricular and atrial tissue cross sections and then relate this molecular data to functional patch clamp analysis of Ito in myocytes isolated from specific anatomical regions of the atrium and ventricle. We will also conduct a similar but limited IF localizationstudy of these same subunits in the human ventricle. PERFORMANCE SITE ========================================Section End===========================================