Fluid movement between airspace and vascular compartments is important in normal lung function and pulmonary edema. This proposal is focused on biophysical (aim 1) and molecular (aim 2) water transporting mechanisms, and the role of molecular water channels in lung physiology (aim 3). The proposal consists of three specific aims:
In Specific Aim 1 they will apply novel optical methods to measure water permeability in intact lung epithelia, endothelia and individual cell membranes. Optical methods were developed in their laboratories to measure: a) airspace-to-capillary osmotic water permeability (Pf) in intact lung; b) transepithelial Pf in airways; and c) membrane Pf in epithelial layers. New optical approaches will be applied to quantify the role of epithelial vs. endothelial barriers in airspace-to-capillary water movement, of type I vs. type II cells in transalveolar water permeability, and of apical vs. basolateral Pf in airway and alveolar epithelia.
In Specific Aim 2 they will identify and characterize molecular water channels (aquaporins) that confer high water permeability in lung. Four aquaporins are expressed in lung: AQP3 (trachea) and AQP4 (airway basolateral membrane), cloned by the principal investigator, AQP1 (endothelium) and AQP5 (alveolar apical membrane) cloned by Dr. Agre. Water channels have not been identified at the alveolar basolateral membrane and airway apical membrane. The following cloning strategies will be used to identify these proteins: PCR-homology cloning using mRNA from alveolar and airway cells, and chromosome-specific exon amplification. Water channels will be studied by Xenopus oocyte expression and immunocytochemistry.
In Specific Aim 3 they will define the role of water channels in lung physiology using transgenic knock-out mice. The investigators recently generated the first animal knock-out models for water channels AQP4 and AQP1 deficient mice. Homozygous transgenic knock-out mice will be generated for AQP3 and AQP5, and characterized in terms of gross phenotype, lung water permeability, clearance of alveolar and interstitial edema, and airway resistance and reactivity.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL059198-02
Application #
6056460
Study Section
Special Emphasis Panel (ZRG2-RAP (01))
Project Start
1998-09-01
Project End
2003-08-31
Budget Start
1999-09-01
Budget End
2000-08-31
Support Year
2
Fiscal Year
1999
Total Cost
Indirect Cost
Name
University of California San Francisco
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
073133571
City
San Francisco
State
CA
Country
United States
Zip Code
94143
Verkman, A S (2013) Aquaporins. Curr Biol 23:R52-5
Verkman, Alan S; Ratelade, Julien; Rossi, Andrea et al. (2011) Aquaporin-4: orthogonal array assembly, CNS functions, and role in neuromyelitis optica. Acta Pharmacol Sin 32:702-10
Crane, J M; Tajima, M; Verkman, A S (2010) Live-cell imaging of aquaporin-4 diffusion and interactions in orthogonal arrays of particles. Neuroscience 168:892-902
Crane, Jonathan M; Haggie, Peter M; Verkman, A S (2009) QUANTUM DOT SINGLE MOLECULE TRACKING REVEALS A WIDE RANGE OF DIFFUSIVE MOTIONS OF MEMBRANE TRANSPORT PROTEINS. Proc SPIE Int Soc Opt Eng 7489:
Namkung, Wan; Song, Yuanlin; Mills, Aaron D et al. (2009) In situ measurement of airway surface liquid [K+] using a ratioable K+-sensitive fluorescent dye. J Biol Chem 284:15916-26
Verkman, Alan S; Galietta, Luis J V (2009) Chloride channels as drug targets. Nat Rev Drug Discov 8:153-71
Ruiz-Ederra, Javier; Levin, Marc H; Verkman, A S (2009) In situ fluorescence measurement of tear film [Na+], [K+], [Cl-], and pH in mice shows marked hypertonicity in aquaporin-5 deficiency. Invest Ophthalmol Vis Sci 50:2132-8
Yang, Baoxue; Zhao, Dan; Verkman, A S (2009) Hsp90 inhibitor partially corrects nephrogenic diabetes insipidus in a conditional knock-in mouse model of aquaporin-2 mutation. FASEB J 23:503-12
Crane, Jonathan M; Verkman, Alan S (2009) Determinants of aquaporin-4 assembly in orthogonal arrays revealed by live-cell single-molecule fluorescence imaging. J Cell Sci 122:813-21
Tradtrantip, Lukmanee; Sonawane, N D; Namkung, Wan et al. (2009) Nanomolar potency pyrimido-pyrrolo-quinoxalinedione CFTR inhibitor reduces cyst size in a polycystic kidney disease model. J Med Chem 52:6447-55

Showing the most recent 10 out of 168 publications