O2-sensitive K+ channels in immortalised rat chromaffin-cell-derived MAH cells

The regulation of K+ channels by O-2 levels is a key link between hypoxia and neurotransmitter release in neuroendocrine cells. Here, we examined the effects of hypoxia on K+ channels in the immortalised v-myc, adrenal-derived HNK1(+) (MAH) cell line. MAH cells possess a K+ conductance that is sensitive to Cd2+, iberiotoxin and apamin, and which is inhibited by similar to24 % when exposed to a hypoxic perfusate (O-2 tension 20 mmHg). This conductance was attributed to high-conductance Ca2+-activated K+ (BK) and small-conductance Ca2+-activated K+ (SK) channels, which are major contributors to the O-2-sensitive K+ conductance in adrenomedullary chromaffin cells. Under low [Ca2+](i) conditions that prevented activation of Ca2+-dependent K+ conductances, a rapidly activating and slowly inactivating K+ conductance, sensitive to both TEA and 4-aminopyridine (4-AP), but insensitive to 100 nM charybdotoxin (CTX), was identified. This current was also reduced (by similar to25 %) when exposed to hypoxia. The hypoxia-sensitive component of this current was greatly attenuated by 10 mM 4-AP, but was only slightly reduced by 10 mM TEA. This suggests the presence of delayed-rectifier O-2-sensitive channels comprising homomultimeric Kv1.5 or heteromultimeric Kv1.5/Kv1.2 channel subunits. The presence of both Kv1.5 and Kv1.2 alpha-subunits was confirmed using immunocytochemical techniques. We also demonstrated that these K+ channel subunits are present in neonatal rat adrenomedullary chromaffin cells in situ. These data indicate that MAH cells possess O-2-sensitive K+ channels with characteristics similar to those observed previously in isolated chromaffin cells, and therefore provide an excellent model for examining the cellular mechanisms of O-2 sensing in adrenomedullary chromaffin cells.