Effects on K+ currents in rat cerebellar granule neurones of a membrane-permeable analogue of the calcium chelator BAPTA

Abstract

1 Whole cell recordings of voltage-activated K+ currents were made with the amphotericin B perforated patch technique from cerebellar granule (CG) neurones of 6-8 days rats that had been in culture for 1 to 16 days. By use of appropriate voltage protocols, the effects of the membrane-permeant form of BAPTA, 1,2-bis-(2-amino-phenoxy)ethane-N,N,N',N'-tetraacetic acid acetoxymethyl ester (BAPTA-AM), on the transient A current (I(KA)), the delayed rectifier current (I(Kv)) and a standing outward current (I(Kso)) were investigated. 2 Bath application of 25 μM BAPTA-AM inhibited both I(Kv) and I(Kso) in cultured neurones, but did not seem to affect I(KA) Neither 25 μM BAPTA (free acid) nor 25 μM ethylenediaminetetraacetic acid acetoxymethyl ester (EDTA-AM) had any significant effect on the magnitude of I(Kso). Similarly in short-term (1-2 days) cultured CG neurones I(Kv), but not I(Kso), was inhibited by 25 μM BAPTA-AM. 3 BAPTA-AM (2.5 μM) reduced I(Kv) in short-term culture CG neurones, with further inhibition being seen when the perfusate was changed to one containing 25 μM BAPTA-AM. 4 Tetraethylammonium ions (TEA) (10 mM) reversibly inhibited I(Kv) in these cells with a similar rate of block of I(Kv) to that induced by 25 μM BAPTA-AM. 5 The degree of inhibition of I(Kv) by 25 μM BAPTA-AM was both time- and voltage-dependent, in contrast to the inhibition of this current by TEA. 6 These data indicate that BAPTA-AM reduces K+ currents in cerebellar granule neurones and that this inhibition cannot be explained in terms of intracellular Ca2+ chelation, but is a direct effect on the underlying channels.