Rationale: Adenosine (ADO) signaling is altered in both asthma and chronic obstructive pulmonary disease and the A_2B adenosine receptor (A_2B-R) may drive pulmonary inflammation. Accordingly, it has been proposed that specific inhibition of the A_2B-R could treat inflammatory lung diseases. However, stimulation of CFTR by ADO may be crucial in permitting the superficial epithelium to maintain airway surface liquid (ASL) volume, which is required to ensure hydrated and clearable mucus. Objective: To determine which ADO receptor (ADO-R) underlies ASL volume regulation in bronchial epithelia. Methods: We used PCR techniques to determine ADO-R expression in bronchial epithelia and used nasal potential difference measurements, Ussing chambers studies and XZ-confocal microscopy to look at Cl^- secretion and ASL volume regulation. Results: The A_2B-R was the mostly highly expressed ADO-R in donor specimens of human bronchial epithelia, and inhibition of ADO-R in vivo prevented activation of CFTR. A_2B-R was the only ADO-R detected in cultured human bronchial epithelial and inhibition of this receptor with specific A_2B-R antagonists resulted in ASL height collapse and a failure to effect ASL height homeostasis. Removal of ADO with adenosine deaminase and replacement with NECA resulted in dose-dependent changes in ASL height and suggested that the cell surface [ADO] may be in excess of 1 µM, which is sufficient to activate A_2B-R. Conclusions: A_2B-R are required for ASL volume homeostasis in human airways and therapies directed at inhibiting A_2B-R may lead to a cystic fibrosis-like phenotype with depleted ASL volume and mucus stasis.