Control of cAMP in lung endothelial cell phenotypes. Implications for control of barrier function

Pulmonary microvascular endothelial cells (PMVECs) form a more restrictive barrier to macromolecular flux than pulmonary arterial endothelial cells (P AECs); however, the mechanisms responsible for this intrinsic feature of PM VECs are unknown. Because cAMP improves endothelial barrier function, we hy pothesized that differences in enzyme regulation of cAMP synthesis and/or d egradation uniquely establish an elevated content in PMVECs. PMVECs possess ed 20% higher basal cAMP concentrations than did PAECs; however, increased content was accompanied by 93% lower ATP-to-cAMP conversion rates. In PMVEC s, responsiveness to beta-adrenergic agonist (isoproterenol) or direct aden ylyl cyclase (forskolin) activation was attenuated and responsiveness to ph osphodiesterase inhibition (rolipram) was increased compared with those in PAECs. Although both types of endothelial cells express calcium-inhibited a denylyl cyclase, constitutive PR NEC cAMP accumulation was not inhibited by physiological rises in cytosolic calcium, whereas PAEC cAMP accumulation w as inhibited 30% by calcium. Increasing either PMVEC calcium entry by maxim al activation of store-operated calcium entry or ATP-to-cAMP conversion wit h rolipram unmasked calcium inhibition of adenylyl cyclase. These data indi cate that suppressed calcium entry and low ATP-to-cAMP conversion intrinsic ally influence calcium sensitivity. Adenylyl cyclase-to-cAMP phosphodiester ase ratios regulate cAMP at elevated levels compared with PAECs, which like ly contribute to enhanced microvascular barrier function.