Noninvasive in vivo fluorescence measurement of airway-surface liquid depth, salt concentration, and pH

The concentration of salt in the thin layer of fluid at the surface of larg e airways, the airway-surface liquid (ASL), is believed to he of central im portance in airway physiology and in the pathophysiology of cystic fibrosis . Invasive sampling methods have yielded a wide range of ASL [NaCl] from 40 to 180 mM. We have developed novel fluorescent probes and microscopy metho ds to measure ASL thickness, salt concentration, and pH quantitatively in c ell-culture models and in the trachea in vivo. By rapid z-scanning confocal microscopy, ASL thickness was 21 +/- 4 mum in well-differentiated cultures of bovine tracheal epithelial cells grown on porous supports at an air-liq uid interface. By ratio imaging fluorescence microscopy using sodium, chlor ide, and pH-sensitive fluorescent indicators, ASL [Na+] was 97 +/- 5 mM, [C l-] In as 118 +/- 3 mM, and pH was 6.94 +/- 0.03. In anesthetized mice in w hich a transparent window was created in the trachea, ASL thickness was 45 +/- 5 mum, [Na+] was 115 +/- 4 mM, [Cl-] was 140 +/- 5 mM, and pH was 6.95 +/- 0.05. Similar ASL tonicity and pH were found in cystic fibrosis (CFTR-n ull) mice. In freshly harvested human bronchi, ASL thickness was 55 +/- 5 m um, [Na+] was 103 +/- 3 mM, [Cl-] was 92 +/- 4 mM, and pH was 6.78 +/- 0.2. These results establish by a noninvasive approach the key properties of th e ASL and provide direct evidence that the ASL is approximately isotonic an d not saltier in cystic fibrosis.