Background: Proteolytic degradation of epithelial sodium channels (ENaC) assists in regulating net salt and water balance in lung epithelia. Results: H 2 O 2 increases surface expression of ␣-ENaC, transepithelial Na transport, and alveolar fluid clearance via redoxsensitive Nedd8. Conclusion: Redox-sensitive Nedd8 is involved in the ubiquitination of lung ENaC. Significance: Understanding ROS-mediated signaling of lung ENaC is crucial for understanding pulmonary physiology and pathology.

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Chronic alcohol consumption is associated with increased incidence of ICU-related morbidity and mortality, primarily from acute respiratory distress syndrome (ARDS). However, the mechanisms involved are unknown. One explanation is that alcohol regulates epithelial sodium channels (ENaC) via oxidant signaling to promote a pro- injury environment. We used small rodent models to mimic acute and chronic alcohol consumption and tested the hypothesis that ethanol (EtOH) would affect lung fluid clearance by up-regulating ENaC activity in the lung. Fluorescence labeling of rat lung slices and in vivo mouse lung revealed an increase in ROS production in response to acute EtOH exposure. Using western blots and fluorescein-5-maleimide labeling, we conclude that EtOH exposure modifies cysteines of α-ENaC while data from single channel patch clamp analysis confirm that 0.16% EtOH increased ENaC activity in rat alveolar cells. In vivo lung fluid clearance demonstrated a latent increase in fluid clearance in mice receiving EtOH diet. Ethanol mice given a tracheal instillation of LPS demonstrated early lung fluid clearance compared to caloric control mice and C57Bl/6 mice. Standard biochemical techniques reveal that chronic EtOH consumption resulted in greater protein expression of the catalytic gp91phox subunit and the obligate Rac1 protein. Collectively these data suggest that chronic EtOH consumption may lead to altered regulation of ENaC, contributing to a ‘pro-injury’ environment in the alcohol lung.

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Cav2.1 channels regulate Ca2+ signaling and excitability of cerebellar Purkinje neurons. These channels undergo a dual feedback regulation by incoming Ca2+ ions, Ca2+-dependent facilitation and inactivation. Endogenous Ca2+-buffering proteins, such as parvalbumin (PV) and calbindin D-28k (CB), are highly expressed in Purkinje neurons and therefore may influence Cav2.1 regulation by Ca2+. To test this, we compared Cav2.1 properties in dissociated Purkinje neurons from wild-type (WT) mice and those lacking both PV and CB (PV/CB−/−). Unexpectedly, P-type currents in WT and PV/CB−/− neurons differed in a way that was inconsistent with a role of PV and CB in acute modulation of Ca2+ feedback to Cav2.1. Cav2.1 currents in PV/CB−/− neurons exhibited increased voltage-dependent inactivation, which could be traced to decreased expression of the auxiliary Cavβ2a subunit compared with WT neurons. Although Cav2.1 channels are required for normal pacemaking of Purkinje neurons, spontaneous action potentials were not different in WT and PV/CB−/− neurons. Increased inactivation due to molecular switching of Cav2.1 β-subunits may preserve normal activity-dependent Ca2+ signals in the absence of Ca2+-buffering proteins in PV/CB−/− Purkinje neurons.