Expression of intermediate-conductance, Ca²⁺-activated K⁺ channel (KCNN4) in H441 human distal airway epithelial cells

Electrophysiological studies of H441 human distal airway epithelial cells showed that thapsigargin caused a Ca²⁺-dependent increase in membrane conductance (G_Tot) and hyperpolarization of membrane potential (V_m). These effects reflected a rapid rise in cellular K⁺ conductance (G_K) and a slow fall in amiloride-sensitive Na⁺ conductance (G_Na). The increase in G_Tot was antagonized by Ba²⁺, a nonselective K⁺ channel blocker, and abolished by clotrimazole, a KCNN4 inhibitor, but unaffected by other selective K⁺ channel blockers. Moreover, 1-ethyl-2-benzimidazolinone (1-EBIO), which is known to activate KCNN4, increased G_K with no effect on G_Na. RT-PCR-based analyses confirmed expression of mRNA encoding KCNN4 and suggested that two related K⁺ channels (KCNN1 and KCNMA1) were absent. Subsequent studies showed that 1-EBIO stimulates Na⁺ transport in polarized monolayers without affecting intracellular Ca²⁺ concentration ([Ca²⁺]_i), suggesting that the activity of KCNN4 might influence the rate of Na⁺absorption by contributing to G_K. Transient expression of KCNN4 cloned from H441 cells conferred a Ca²⁺- and 1-EBIO-sensitive K⁺ conductance on Chinese hamster ovary cells, but this channel was inactive when [Ca²⁺]_i was <0.2 μM. Subsequent studies of amiloride-treated H441 cells showed that clotrimazole had no effect on V_m despite clear depolarizations in response to increased extracellular K⁺ concentration ([K⁺]_o). These findings thus indicate that KCNN4 does not contribute to V_m in unstimulated cells. The present data thus establish that H441 cells express KCNN4 and highlight the importance of G_K to the control of Na⁺ absorption, but, because KCNN4 is quiescent in resting cells, this channel cannot contribute to resting G_K or influence basal Na⁺ absorption.