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Published ahead of print on November 7, 2003, doi:10.1165/rcmb.2003-0325OC

Am. J. Respir. Cell Mol. Biol., Volume 30, Number 5, May 2004, 720-728

A more recent version of this article appeared on May 1, 2004
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Submitted on September 5, 2003
Revised on November 6, 2003

REGULATION OF AMILORIDE-SENSITIVE Na+ TRANSPORT BY BASAL NITRIC OXIDE

Karin M Hardiman1, Carmel M McNicholas-Bevensee1, James Fortenberry2, Carpantato T Myles3, Bela Malik4, Douglas C Eaton4, and Sadis Matalon5*

1 Physiology and Biophysics, University of Alabama, Birmingham, AL, USA, 2 Gregory Fleming James Cystic Fibrosis Research Center, University of Alabama, Birmingham, AL, USA, 3 Anesthesiology, University of Alabama, Birmingham, AL, USA, 4 Physiology, Emory University, Atlanta, GA, USA; Center for Cell and Molecular Signaling, Emory University, Atlanta, GA, USA, 5 Anesthesiology, University of Alabama, Birmingham, AL, USA; Physiology and Biophysics, University of Alabama, Birmingham, AL, USA

* To whom correspondence should be addressed. E-mail: sadis{at}uab.edu.

We investigated the mechanisms of endogenous nitric oxide (NO) modulation of lung sodium (Na+) transport. C57BL/6 mice injected intra-peritoneally with the specific iNOS inhibitor 1400W (10 mg/kg every 8 hr for 72 h), exhibited decreased alveolar nitrite levels and Na+-dependent amiloride-sensitive alveolar fluid clearance as compared to mice injected with vehicle. Similarly, pretreatment of mouse tracheal epithelial (MTE) cells with 1400W abolished the inhibitory effects of amiloride on their Na+ short circuit currents. On the other hand, MTE cells pretreated with 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ), a specific inhibitor of guanylate cyclase, had lower levels of cGMP, but normal values of amiloride-sensitive Na+ currents. Amiloride also inhibited whole-cell Na+ currents across A549 cells treated with vehicle ( Ki = 249 nM) but had no effect in A549 cells treated with 1400W. Western blotting studies showed significantly lower levels of {alpha} and {gamma}ENaC in lung tissues and alveolar type II (ATII) cells from iNOS(-/-) as well as iNOS (+/+) mice treated with 1400W, as compared to the corresponding values from vehicle treated iNOS (+/+) mice. Similar values for ratios of {alpha}, {beta}, and {gamma}enac to gapdh were obtained by real time PCR for iNOS(+/+) mice and iNOS(-/-) mice. We concluded that NO derived from iNOS under basal conditions is necessary for amiloride-sensitive Na+ transport across lung epithelial cells and modulates the amount of {alpha} and {gamma} ENaC via post-transcriptional, cGMP-independent mechanisms.




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