Cystic fibrosis is caused by dysfunction of the cystic fibrosis transmembrane conductance regulator (CFTR) protein, leading to altered ion transport, chronic infection, and excessive inflammation. Here we investigated regulation of CFTR in airway cell monolayers by adenosine, adenosine receptors, and arachidonic acid. Our studies demonstrate that the A2B adenosine receptor is expressed at high levels relative to the other adenosine receptor subtypes, with a characteristic low-affinity profile for adenosine-stimulated CFTR Cl- currents in both Calu-3 cells and CFBE41o- airway cell monolayers stably transduced with wild-type CFTR. The levels of adenosine found in sputum from patients with cystic fibrosis with moderate to severe lung disease stimulated apical prostaglandin release in Calu-3 and CFBE41o- cells, implicating adenosine regulation of phospholipase A2 (PLA2) activity. A2B adenosine receptor and arachidonic acid stimulation produced CFTR-dependent currents in airway monolayers and increased cAMP levels that were sensitive to cyclooxygenase inhibition. Arachidonic acid demonstrated dual regulation of CFTR, stimulating CFTR and Cl- currents in intact airway monolayers, and potently inhibiting PKA-activated Cl- currents in excised membrane patches. Cl- currents produced by arachidonic acid were sensitive to inhibition of PKA, cyclooxygenase, and 5-lipoxygenase. Together, the results provide a converging mechanism to link regulation of CFTR and airway cell inflammation through adenosine and adenosine receptors.