Modulation of glioma BK channels via erbB2

Glioma cells show up-regulation and constitutive activation of erbB2, and its expression correlates positively with increased malignancy. A similar correlation has been demonstrated for the expression of gBK, a calcium-sensitive, large-conductance K+ channel. We show here that glioma BK channels are a downstream target of erbB2/neuregulin signaling. Tyrphostin AG825 was able to disrupt the constituitive erbB2 activation in a dose-dependent manner, causing a 30-mV positive shift in gBK channel activation in cell-attached patches. Conversely, maximal stimulation of erbB2 with a recombinant neuregulin (NRG-1 beta) caused a 12-mV shift in the opposite direction. RT-PCR studies reveal no change in the BK splice variants expressed in treated glioma cells. Furthermore, isolation of surface proteins through biotinylation did not show a change in gBK channel expression, and probing with phospho-specific antibodies showed no alteration in channel phosphorylation. However, fura-II Ca2+ fluorescence imaging revealed a 35% decrease in the free intracellular Ca2+ concentration after erbB2 inhibition and an increase in NRG-1 beta-treated cells, suggesting that the observed changes most likely were due to alterations in [Ca2+](i). Consistent with this conclusion, neither tyrphostin AG825 nor NRG-1 beta was able to modulate gBK channels under inside-out or whole-cell recording conditions when intracellular Ca2+ was fixed. Thus, gBK channels are a downstream target for the abundantly expressed neuregulin-1 receptor erbB2 in glioma cells. However, unlike the case in other systems, this modulation appears to occur via changes in [Ca2+](i) without changes in channel expression or phosphorylation. The enhanced sensitivity of gBK channels in glioma cells to small, physiological Ca2+ changes appears to be a prerequisite for this modulation. (c) 2005 Wiley-Liss, Inc.