blocking activity of different toxins against potassium channels Kv3.4 in RLE cells

ABSTRACT

K[+] channel toxins are essential tools for the first purifications, analysis of subunit structures and brain localization of voltage-gated K[+] [Kv] channels. The effects of a lot of toxins on Kv are not fully known. Using whole-cell patch clamping technique the action of a series of toxins on Kv3.4 current in rat liver cells with expressed Kv3.4 channels [RLE] cloned cells was investigated. The cells were grown in Williams E medium and after 6-8 days, they were suitable for patch clamping. A family of currents was recorded during voltage-clamp steps to various potentials applied from a holding potential of-60 mV to 60-80 mV in 10 mV increments. Upon depolarization, all channels were opened with a sigmoidal time course, reached to the peak within a few 10[th] of milliseconds and then slowly inactivated. Bath application of tetraethyl ammonium [TEA] or 3, 4-diaminopyridine [DAP] reduced the current dose dependently and inhibited it completely at 3 mM and 25 micro M respectively. The Bunodosoma granulifera [BgK] and Heteractis magniflca [HmK] toxins at concentrations up to 30 and 10 micro M respectively could not completely inhibit the current. On the hand, toxins such as beta-bungarotoxin, corotoxin, novel toxin and dendrotoxins I [DIP] and K [DPK] even in high concentrations [up to 100 mM] had not any significant effect on Kv3.4 current. Comparison of chemical structures of these effective agents with other reported effective toxins such as blood depressing substances [BDS 1 and II] show no homology between them, but specially the potency of 3, 4-DAP is comparable with these toxins. These results showed that, the Kv3.4 is more sensitive than other K[+] channels to 3, 4-DAP. The sensitivity of this channel to the TEA is low [at mM concentration]. More investigation is necessary to find more selective and potent inhibitor of Kv3.4 channels