Chronic in utero cyclooxygenase inhibition alters PGE2-regulated ductus arteriosus contractile pathways and prevents postnatal closure

Abstract

Although prostaglandin E2 (PGE2) vasodilates the ductus arteriosus, tocolysis with cyclooxygenase (COX) inhibitors delays postnatal ductus arteriosus closure. We used fetal mice and sheep to determine whether PGE2 has a role in the development of ductus contractility that is distinct from its function as a vasodilator. Prolonged exposure of fetal ductus to PGE2 in vitro increased the expression of CaL- and K+-channel genes (CaLalpha1c, CaLbeta2, Kir6.1, and Kv1.5, which regulate oxygen-induced constriction) without affecting the genes that regulate Rho-kinase-mediated calcium sensitization. Conversely, chronic exposure to COX inhibitors in utero decreased expression of CaL- and K+-channel genes, without affecting Rho-kinase-associated genes. Chronic COX inhibition in utero decreased the ductus' in vitro contractile response to stimuli that use CaL- and K+-channels (like O2 and K+), whereas the response to stimuli that act through Rho-kinase-mediated pathways (like U46619) was not significantly affected. Phosphodiesterase expression, which decreases the ductus' sensitivity to cAMP- or cGMP-dependent vasodilators, was increased by PGE2 exposure and decreased by COX inhibition, respectively. These studies identify potential downstream effectors of a PGE2-mediated, developmental program, regulating oxygen-induced ductus closure. Alterations in these effectors may explain the increased risk of patent ductus arteriosus (PDA) after in utero COX inhibition.

Figures

Figure 1

Response of the fetal and newborn ductus arteriosus to acute (COX-In d19) and chronic (COX-In d15–19) COX inhibition in vivo. DA=ductus and AO=transverse aorta lumen diameters. Acute COX inhibition constricted the fetal ductus at term gestation (n = 17 fetuses; 7 litters) compared with Control fetuses (n = 9; 6 litters). Chronic COX inhibition (n = 17; 6 litters) did not constrict the fetal ductus. Ductus closure in 4 hour-old newborn pups (chronically exposed to COX inhibitors in utero (n = 26; 11 litters) was significantly reduced (resulting in a PDA) compared to Control newborns (n = 12; 4 litters). *p<0.05 compared to fetal controls; § p<0.05 compared to newborn control.

Figure 2

Contractile response of isolated ductus from19 day-old fetal mice treated with or without chronic COX inhibitors. Ductus were incubated in deoxygenated buffer containing L-NAME and indomethacin and exposed to increasing oxygen (A, D), K+ (B, E), or U-46619 (C, F) concentrations. P = maximal relaxation with papaverine. Steady state oxygen tensions were 42.5±2., 57.3±2.1, 75.1±1.8, 118.8±1.7, and 181.0±5.6 mmHg when the buffer was bubbled with 0, 2, 5, 12, and 21% oxygen, respectively. The K+ and U-46619 dose-response experiments were performed in 0% oxygen. Degree of lumen closure = 1-(measured diameter/ diameter after papaverine relaxation). Open squares = Controls, solid squares = chronic COX inhibition. Animal numbers: O2 (A, D): Control=6, COX inhibition=6; K+ (B,E): Control=9, COX inhibition=5; U-46619 (C, F): Control=7, COX inhibition=6. *p<0.05 compared to Control.