title: Pregnancy-specific modulatory role of mitochondria on adenosine 5'-triphosphate-induced cytosolic [Ca2+] signaling in uterine artery endothelial cells.
abstract: Vascular endothelial cells respond to extracellular ATP by inositol 1,4,5-trisphosphate-mediated Ca2+ release from the endoplasmic reticulum followed by Ca2+ influx and subsequent synthesis of vasodilators. In this study, the contribution of mitochondria in shaping the ATP-induced Ca2+ increase was examined in ovine uterine artery endothelial cells from nonpregnant and pregnant (late gestation) ewes (NP- and P-UAEC, passage 4). The mitochondrial protonophore carbonyl cyanide m-chlorophenylhydrazone (CCCP) induced a rapid mitochondrial depolarization. CCCP also slowly increased cytosolic [Ca2+] ([Ca2+]c), which then gradually declined to 10-20 nM above resting level. Pretreatment with CCCP for 30 min significantly inhibited both ATP and thapsigargin-induced [Ca2+]c, with inhibition in NP-UAEC more effective than in P-UAEC. Pretreatment of mitochondrial permeability transition pore inhibitor cyclosporine A did not affect CCCP-induced mitochondrial depolarization, but delayed CCCP-induced [Ca2+]c for about 12-15 min (we termed this the "window of time"). During the cyclosporine A-delayed window of time of CCCP-induced [Ca2+]c, ATP induced a normal Ca2+ response, but after this window of time, ATP-induced [Ca2+]c was significantly inhibited. Pretreatment of oligomycin B to prevent intracellular ATP depletion by F0F1-ATPase did not reduce the inhibition of ATP-induced [Ca2+]c by CCCP. Ruthenium red, a mitochondrial Ca2+ uptake blocker, did not mimic the inhibition of Ca2+ signaling by CCCP. In conclusion, our data show that mitochondrial Ca2+ depletion after dissipation of mitochondrial membrane potential with CCCP inhibits ATP-induced [Ca2+]c, mediated at the level of Ca2+ release from the endoplasmic reticulum. Moreover, our data revealed that P-UAEC is more resistant to the inhibitory effect of CCCP on [Ca2+]c than NP-UAEC.
authors.affiliation: University of Wisconsin-Madison, Department of Obstetrics and Gynecology, Perinatal Research Laboratories, 7E Meriter Hospital/Park, 202 South Park Street, Madison, Wisconsin 53715, USA. fuxianyi@wisc.edu
authors.forename: Fu-Xian
authors.initials: FX
authors.lastname: Yi
authors.forename: Ian M
authors.initials: IM
authors.lastname: Bird
mesh_terms: D000255:Adenosine Triphosphate / Q000172:deficiency / Q000378:metabolism* / Q000494:pharmacology; D000818:Animals; D001158:Arteries; D002118:Calcium / Q000378:metabolism; D020013:Calcium Signaling / Q000502:physiology*; D002258:Carbonyl Cyanide m-Chlorophenyl Hydrazone / Q000494:pharmacology; D003600:Cytosol / Q000378:metabolism*; D004594:Electrophysiology; D042783:Endothelial Cells / Q000187:drug effects / Q000378:metabolism* / Q000502:physiology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005260:Female; D008928:Mitochondria / Q000187:drug effects / Q000502:physiology*; D009994:Osmolar Concentration; D011247:Pregnancy; D011270:Pregnancy, Animal / Q000502:physiology*; D012756:Sheep; D019284:Thapsigargin / Q000494:pharmacology; D014599:Uterus / Q000098:blood supply*
chemical_list: D004791:Enzyme Inhibitors; D002258:Carbonyl Cyanide m-Chlorophenyl Hydrazone; D019284:Thapsigargin; D000255:Adenosine Triphosphate; D002118:Calcium