Expression environment determines K + current properties: Kv1 and Kv4 α-subunit-induced K + currents in mammalian cell lines and cardiac myocytes

Abstract

Voltage-gated K+ channel (Kv) pore-forming (alpha) subunits of the Kv1 and Kv4 subfamilies have been cloned from heart cDNA libraries, and are thought to play roles in the generation of the transient outward K+ current, Ito. Heterologous expression of these subunits in Xenopus oocytes, however, reveals K+ currents that are quite distinct from Ito. In the experiments here, the detailed time- and voltage-dependent properties of the currents expressed in mammalian cell lines and in cardiac myocytes by Kv1.4 and Kv4.2 were examined and compared to previous findings in studies of oocytes, as well as to Ito characterized in various myocardial cells. As in oocytes, expression of Kv1.4 in HEK-293, Ltk- or neonatal rat ventricular cells reveals rapidly activating K+ currents. In contrast to the currents in oocytes, however, there are two components of inactivation of the Kv1.4-induced currents in mammalian cells, and both components are significantly slower in myocytes than in either HEK-293 or Ltk- cells. In addition, in all three cell types, recovery of Kv1.4 from steady-state inactivation is very slow, proceeding with mean time constants in the range of 6-8 s. The properties of Kv4.2-induced currents also vary with cell type and, importantly, the rates of activation, inactivation and recovery from inactivation are significantly faster in mammalian cells than in Xenopus oocytes. In HEK-293, Chinese hamster ovary (CHO) and neonatal rat ventricular cells, for example, the currents recover from steady-state inactivation with mean (+/-SD) time constants of 153+/-32 (n=12), 245+/-112 (n=10) and 86+/-38 (n=11) ms, respectively; therefore, recovery proceeds 5-10 times faster than observed for Kv4.2 in oocytes. These results emphasize the importance of the cellular expression environment in efforts to correlate endogenous K+ currents with heterologously expressed K+ channel subunits. In addition, the finding that Kv alpha subunits produce distinct K+ currents in different cells suggests that cell-type-specific associations with endogenous Kv alpha or accessory beta subunits and/or post-translational processing play roles in determining the properties of functional K+ channels.

Molecular Basis of Voltage-Dependent Potassium Currents in Porcine Granulosa Cells

“…The subunit composition of Kv channels influences the expression, and electrophysiological and pharmacological properties of the associated currents (Hopkins, 1998). Kv channel expression and function may be modulated not only by coassociation of the various pore-forming ␣-subunits with accessory (␤)-subunits and other regulatory proteins but also by post-translational modifications of either the Kv ␣-or Kv ␤-subunits (Martens et al, 1999;Petersen and Nerbonne, 1999). Several K ϩ channel subunits are typically expressed in a single cell.…”

“…Cumulative inactivation is not apparent. have been impossible to adequately test this hypothesis by measuring only electrophysiological and pharmacological characteristics of I Kur , because 1) the voltage dependence, gating kinetics, and toxin sensitivity of heteromeric K ϩ channels are not easily predicted and vary considerably with expression environment (Hopkins, 1998;Petersen and Nerbonne, 1999); and 2) there are no specific antagonists of Kv1.5. Accordingly, we performed additional experiments with standard molecular and biochemical techniques, as detailed below.…”

Cell Surface Targeting and Clustering Interactions between Heterologously Expressed PSD-95 and the Shal Voltage-gated Potassium Channel, Kv4.2

“…Percentages (from the biotinylation cell surface assay) were subjected to arcsin transformation to produce data with a normal distribution (30) (31,32), no voltage-gated currents were detected in whole-cell recordings from cells transfected with pEGFP-C1 alone (mock-transfected; n ϭ 6) (Fig. 1A).…”

Phenotypic differences in transient outward K+ current of human and canine ventricular myocytes: insights into molecular composition of ventricular Ito

“…For example, recent reports have elegantly implicated I to in the mechanism of arrhythmias associated with Brugada syndrome, whereby phase 2 reentrant excitation could occur between cells with recovered I to and those with inactive I to (3,9). Both Kv4.3 and Kv1.4 proteins are expressed in the myocardium of large mammals (29). Flecainide and H 2 O 2 have been used to determine the contribution of these two K ϩ channel subunits to native I to (12,36,39).…”