Regulation of Alveolar Epithelial Na+ Channels by ERK1/2 in Chlorine-Breathing Mice

The mechanisms by which the exposure of mice to Cl-2 decreases vectorial Na+ transport and fluid clearance across their distal lung spaces have not been elucidated. We examined the biophysical, biochemical, and physiological changes of rodent lung epithelial Na+ channels (ENaCs) after exposure to Cl-2, and identified the mechanisms involved. We measured amiloride-sensitive short-circuit currents (I-amil) across isolated alveolar Type II (ATII) cell monolayers and ENaC single-channel properties by patching ATII and ATI cells in situ. alpha-ENaC, gamma-ENaC, total and phosphorylated extracellular signalrelated kinase (ERK) 1/2, and advanced products of lipid peroxidation in ATII cells were measured by Western blot analysis. Concentrations of reactive intermediates were assessed by electron spin resonance (ESR). Amiloride-sensitive Na+ channels with conductances of 4.5 and 18 pS were evident in ATI and ATII cells in situ of air-breathing mice. At 1 hour and 24 hours after exposure to Cl-2, the open probabilities of these two channels decreased. This effect was prevented by incubating lung slices with inhibitors of ERK1/2 or of proteasomes and lysosomes. The exposure of ATII cell monolayers to Cl-2 increased concentrations of reactive intermediates, leading to ERK1/2 phosphorylation and decreased I-amil and alpha-ENaC concentrations at 1 hour and 24 hours after exposure. The administration of antioxidants to ATII cells before and after exposure to Cl-2 decreased concentrations of reactive intermediates and ERK1/2 activation, which mitigated the decrease in I-amil and ENaC concentrations. The reactive intermediates formed during and after exposure to Cl-2 activated ERK1/2 in ATII cells in vitro and in vivo, leading to decreased ENaC concentrations and activity.