Expression in mammalian cells and electrophysiological characterization of two mutant Kv1.1 channels causing episodic ataxia type 1 (EA‐1)

Abstract

Episodic ataxia type 1 (EA-1) is a rare neurological disorder and was the first ionic channel disease to be associated with defects in a potassium channel. Until now 10 different point mutations in the KCNA1-gene have been reported to cause this disorder. We have investigated the functional consequences of two mutations leading to amino acid substitutions in the first and sixth transmembrane segments of a Kv1.1 channel subunit, by means of the patch-clamp technique; we injected cRNA coding for, respectively, F184C and V408A mutant Kv1.1 channels into mammalian cells and compared the resulting currents with those in the wild-type. The expression levels of F184C and V408A mutant channels relative to that of the wild-type was 38 and 68%, respectively. Since the single-channel conductance of the F184C mutant was similar to that of the wild-type (12 pS) without an apparent change in the maximum open probability, we conclude that the lower expression level in the F184C mutant channels is due to a reduced number of functional channels on the cell surface. F184C activated slower, and at more depolarized potentials, and deactivated faster compared with the wild-type. V408A channels deactivated and inactivated faster compared with the wild-type. Studies with different extracellular cations and tetraethylammonium gave no indication that the pore structure was changed in the mutant channels. Acetazolamide, that is helpful in some patients suffering from EA-1, was without effect on Kv1.1 wild-type or mutant channels. This study confirms and extends earlier studies on the functional consequences of Kv1.1 mutations associated with EA-1, in an attempt to understand the pathophysiology of the disease.

Ion channels and epilepsy

“…Functional expression in Xenopus oocytes or mammalian cells resulted in a reduction of the K currents either by diminished expression or shifts in voltage-dependence. According to these studies, both dominant negative effects on WT channels and haploinsuf®ciency can cause EA-1 [Adelman et al, 1995;Zerr et al, 1998;Bretschneider et al, 1999;Zuberi et al, 1999;Eunson et al, 2000]. A speci®c defect for those mutations going along with an epilepsy phenotype could not be found [Zuberi et al, 1999;Eunson et al, 2000].…”

Episodic ataxia type 1 mutations differentially affect neuronal excitability and transmitter release

“…Kv1.1-containing channels exhibit variable kinetics depending on the other pore-forming subunits and associated cytoplasmic  subunits (Rettig et al, 1994;Lai and Jan, 2006). The functional consequences of KCNA1 mutations have been examined by expression of mutant and/or wild-type (WT) Kv1.1 in non-neuronal cells (Adelman et al, 1995;D' Adamo et al, 1998;Zerr et al, 1998a;Zerr et al, 1998b;Zuberi et al, 1999;Eunson et al, 2000;Bretschneider et al, 1999;Boland et al, 1999;Rea et al, 2002) . Mutations confer a loss of Kv1.1 function through altered kinetics or reduced current density, in some cases with impaired trafficking (Rea et al, 2002;Manganas et al, 2001).…”

Episodic Ataxia Type-1 Mutations in the Kv1.1 Potassium Channel Display Distinct Folding and Intracellular Trafficking Properties

“…Genetic linkage studies in EA-1 patients have identified at least 14 mutations in the mammalian KCNA1 gene, which encodes the Kv1.1 ␣-subunit, as the underlying cause of EA-1 (2)(3)(4). Most of the KCNA1 mutations in EA-1 patients lead to Kv1.1 channels with altered biophysical characteristics, although certain mutations eliminate any detectable channel activity (5)(6)(7)(8)(9)(10)(11)(12)(13).…”