581 J. Gen. Physiol. © The Rockefeller University Press • 0022-1295/99/04/581/20 $2.00 Volume 113 April 1999 581–600 http://www.jgp.org Distinct Transient Outward Potassium Current (I to ) Phenotypes and Distribution of Fast-inactivating Potassium Channel Alpha Subunits in Ferret Left Ventricular Myocytes Mulugu V. Brahmajothi,* Donald L. Campbell,* Randall L. Rasmusson, ‡ Michael J. Morales,* James S. Trimmer, § Jeanne M. Nerbonne, i and Harold C. Strauss* ¶ From the *Department of Pharmacology, ‡ Department of Biomedical Engineering, and ¶ Department of Medicine, Duke University Medical Center, Durham, North Carolina 27710; § Department of Biochemistry and Cell Biology, State University of New York, Stony Brook, Stony Brook, New York 11794; and i Department of Molecular Biology and Pharmacology, Washington University School of Med- icine, St. Louis, Missouri 63110 abstract The biophysical characteristics and a subunits underlying calcium-independent transient outward potassium current (I to ) phenotypes expressed in ferret left ventricular epicardial (LV epi) and endocardial (LV endo) myocytes were analyzed using patch clamp, fluorescent in situ hybridization (FISH), and immunofluores- cent (IF) techniques. Two distinct I to phenotypes were measured (21–228C) in the majority of LV epi and LV endo myocytes studied. The two I to phenotypes displayed marked differences in peak current densities, activation thresholds, inactivation characteristics, and recovery kinetics. I to,epi recovered rapidly [t rec, 270 mV 5 51 6 3 ms] with minimal cumulative inactivation, while I to,endo recovered slowly [t rec, 270 mV 5 3,002 6 447 ms] with marked cumu- lative inactivation. Heteropoda toxin 2 (150 nM) blocked I to,epi in a voltage-dependent manner, but had no effect on I to,endo . Parallel FISH and IF measurements conducted on isolated LV epi and LV endo myocytes demonstrated that Kv1.4, Kv4.2, and Kv4.3 a subunit expression in LV myocyte types was quite heterogenous: (a) Kv4.2 and Kv4.3 were more predominantly expressed in LV epi than LV endo myocytes, and (b) Kv1.4 was expressed in the majority of LV endo myocytes but was essentially absent in LV epi myocytes. In combination with previous mea- surements on recovery kinetics (Kv1.4, slow; Kv4.2/4.3, relatively rapid) and Heteropoda toxin block (Kv1.4, insensi- tive; Kv4.2, sensitive), our results strongly support the hypothesis that, in ferret heart, Kv4.2/Kv4.3 and Kv1.4 a subunits, respectively, are the molecular substrates underlying the I to,epi and I to,endo phenotypes. FISH and IF mea- surements were also conducted on ferret ventricular tissue sections. The three I to a subunits again showed distinct patterns of distribution: (a) Kv1.4 was localized primarily to the apical portion of the LV septum, LV endocar- dium, and approximate inner 75% of the LV free wall; (b) Kv4.2 was localized primarily to the right ventricular free wall, epicardial layers of the LV, and base of the heart; and (c) Kv4.3 was localized primarily to epicardial lay- ers of the LV apex and diffusely distributed in the LV free wall and septum. Therefore, in intact ventricular tissue, a heterogeneous distribution of candidate I to a subunits not only exists from LV epicardium to endocardium but also from apex to base. key words: cardiac repolarization • patch clamp • Kv1.4 • Kv4.2 • Kv4.3 introduction Due to their primary importance in both initiating and modulating repolarization of the cardiac action poten- tial, extensive experimental effort has been devoted to analysis of potassium channels as potential targets for antiarrhythmic agents (Rasmusson et al., 1994; Tseng, 1995; Tamkun et al., 1995; Barry and Nerbonne, 1996; Sanguinetti and Spector, 1997). In particular, the cal- cium-independent transient outward potassium cur- rent I to (also referred to as I to,1 ; Antzelevitch et al., 1995; Campbell et al., 1995) has recently received much experimental attention. I to is present in nearly all mammalian working cardiac myocyte types, including both human atrial (Escande et al., 1987; Shibata et al., 1988) and ventricular myocytes (Näbauer et al., 1993, 1996; Beuckelmann et al., 1993; Wettwer et al., 1993, 1994; Amos et al., 1996). Due to its rapid activation ki- netics, I to is the major current responsible for the early “notch” or phase 1 repolarization characteristic of the mammalian ventricular action potential. In addition, due to its slower inactivation kinetics, I to can also signif- icantly modulate the plateau and early phase 3 repolar- ization (Campbell et al., 1993a,b, 1995). Despite the nearly universal presence of I to in mam- R.L. Rasmusson’s present address is Department of Medicine, Alle- gheny University of the Health Sciences, Pittsburgh, PA 15212. D.L. Campbell’s, M.V. Brahmajothi’s, M. Morales’s, and H.C. Strauss’s present address is Department of Physiology and Biophysics, School of Medicine and Biomedical Sciences, University at Buffalo, SUNY, Buffalo, NY 14214. Address correspondence to D.L. Campbell, Department of Physi- ology and Biophysics, School of Medicine and Biomedical Sciences, University at Buffalo, SUNY, 124 Sherman Hall, Buffalo, NY 14214. Fax: 716-829-2344. Downloaded from http://rupress.org/jgp/article-pdf/113/4/581/1772327/gp-7767.pdf by guest on 28 July 2023