The reabsorption of Na⁺ in distal lung airways is considered to depend at least partially on a non-selective ion channel, presumably formed by ENaC subunits. Ussing chamber experiments on monolayers of dexamethasone treated NCI-H441 cells revealed an apical ion conductance of moderate selectivity, which contributes to transepithelial Na+ transport. This conductance was sensitive to amiloride derivatives with a potency in the following order benzamil = amiloride > (5-(N-ethyl-N-isopropyl)amiloride) EIPA (Ramminger et al., Am J Physiol Lung Cell Mol Physiol, 2004).

Whole cell patch clamp experiments on H441 cells revealed slightly outward rectifying whole cell currents with a reversal potentail close to 0 mV. Extra cellular Cl^- depletion did not alter the reversal potential, ruling out any Cl^- conductance. This current was blocked by amiloride derivatives, phenamil >> benzamil = amiloride > EIPA with IC₅₀ values 38 0.01 nM, 0.29 0.04 mM, 0.44 0.04 mM, 1.08 0.04 mM respectively. Thus, its sensitivity to amiloride derivatives equals the one of the reported apical Na⁺ conductance in H441 cells. Inhibitors of transient receptor potential (TRP) channels also blocked the identified current as followed: Capsazepine = SB366791 > La³⁺ > SKF96365 with IC₅₀ values 1.21 0.02 nM, 2.2 0.04 nM, 0.099 0.02 mM, 1.3 0.01 mM respectively.

Semi quantitative RT-PCR experiments revealed the highest expression level for aENaC subunits in H441 cells. The expression for ßENaC and gENaC subunits were approximately ten- and 500-fold reduced compared to aENaC respectively. dENaC transcripts could not be detected. However, transcripts encoding TRPV1, TRPC1 and TRPP2 were detected at reasonable expression levels.

Capsazepine and SB366791, the most potent blockers of H441 whole cell currents, were tested for their effect on transepithelial Na⁺ transport in Ussing chambers on M1 cells. Both compounds reduced the short circuit current (I_SC) at concentrations above 3 mM. At concentrations of 30 mM they reduced the I_SC by ca. 20% of the amiloride sensitive I_SC only. This suggests that the identified amiloride-sensitive whole cell currents in H441 cells are mediated by ion channels different from those responsible for the amiloride sensitive Na⁺ transport in M1 monolayers.

We demonstrate that the TRP channel blockers capsazepine and SB366791 do not affect amiloride sensitive I_SC, unless 500 fold higher concentrations are applied than those sufficient to block amiloride-sensitive whole cell currents in H441 cells. It is questionable therefore that amiloride-sensitive ENaC currents in distal airway epithelia solely contribute to pulmonary fluid clearance and edema prevention.

Supported by: DFG D1402, FWF P15743, the 6th framework of the EU, Pulmo-Net & Boehringer Ingelheim Pharma GmbH & CoKG.