Change of cytosolic Ca(2+) mobility in cultured bovine corneal endothelial cells by endothelin-1.
The effect of endothelin-1 (ET-1) on the intracellular free Ca(2+) ([Ca(2+)](i)) in cultured bovine corneal endothelial cells was studied after loading with fura-2-AM. In Ca(2+)-containing buffer and Ca(2+)-free buffer, ET-1 induced a significant rise in [Ca(2+)](i) at concentrations from 10(-9) to 10(-7) M. In Ca(2+)-free buffer, pretreatment of the cells with ET-1 inhibited thapsigargin-induced [Ca(2+)](i) increase and carbonylcyanide m-chlorophenylhydrazone (CCCP)-induced Ca(2+) release by 99% and 62%, respectively. Pretreatment of the cells with thapsigargin or CCCP also inhibited ET-1-induced [Ca(2+)](i) rise by 36% and 92%, respectively. In Ca(2+)-containing buffer, the ET(A) receptor antagonist (BQ123) and ET(B) receptor antagonist (BQ788) partially inhibited ET-1-induced [Ca(2+)](i) by 92% and 98%, respectively. Nifedipine and La(3+) also inhibited ET-1-induced [Ca(2+)](i) increase by 26% and 91%, respectively. The intracellular calcium release caused by ET-1 was partially inhibited by phospholipase C inhibitor (U73122). After incubation of the cells with ET-1 in Ca(2+)-free buffer, the addition of 5 mM CaCl(2) increased Ca(2+) influx, implying that release of Ca(2+) from internal stores caused by ET-1 further induced capacitative Ca(2+) entry. These data suggest that ET-1-induced [Ca(2+)](i) rise in bovine corneal endothelial cells are mediated by ET(A) receptor, ET(B) receptor, La(3+)-sensitive Ca(2+) pump and L-type voltage-operated Ca(2+) channel, leading to Ca(2+) influx. ET-1 also increased the internal Ca(2+) release from endoplasmic reticulum and mitochondria Ca(2+) stores followed by capacitative Ca(2+) entry. ET-1-induced intracellular Ca(2+) release was modulated by phospholipase C-coupled events.
Hong
Show-Jen
SJ
Department of Pharmacology, Kaohsiung Medical University, Kaohsiung, Taiwan.
Wu
Kwou-Yeung
KY
Wang
Hwei-Zu
HZ
Fong
Jim C
JC
D000818:Animals; D001692:Biological Transport / Q000187:drug effects; D002021:Buffers; D002118:Calcium / Q000378:metabolism* / Q000494:pharmacology; D020746:Calcium Channels, L-Type / Q000502:physiology; D000252:Calcium-Transporting ATPases / Q000187:drug effects / Q000378:metabolism; D002417:Cattle; D002460:Cell Line; D003600:Cytosol / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D019332:Endothelin-1 / Q000008:administration & dosage / Q000494:pharmacology*; D004728:Endothelium, Corneal / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism*; D005456:Fluorescent Dyes; D016257:Fura-2 / Q000031:analogs & derivatives*; D007811:Lanthanum / Q000494:pharmacology; D009994:Osmolar Concentration; D010738:Type C Phospholipases / Q000378:metabolism
D002021:Buffers; D020746:Calcium Channels, L-Type; D019332:Endothelin-1; D005456:Fluorescent Dyes; C049925:fura-2-am; D007811:Lanthanum; D010738:Type C Phospholipases; D000252:Calcium-Transporting ATPases; D002118:Calcium; D016257:Fura-2