The effects of extracellular nucleotides on [Ca2+]i signalling in a human-derived renal proximal tubular cell line (HKC-8).
HKC-8 cells are a human-derived renal proximal tubular cell line and provide a useful model system for the study of human renal cell function. In this study, we aimed to determine [Ca(2+)](i) signalling mediated by P2 receptor in HKC-8. Fura-2 and a ratio imaging method were employed to measure [Ca(2+)](i) in HKC-8 cells. Our results showed that activation of P2Y receptors by ATP induced a rise in [Ca(2+)](i) that was dependent on an intracellular source of Ca(2+), while prolonged activation of P2Y receptors induced a rise in [Ca(2+)](i) that was dependent on intra- and extracellular sources of Ca(2+). Pharmacological and molecular data in this study suggests that TRPC4 channels mediate Ca(2+) entry in coupling to activation of P2Y in HKC-8 cells. U73221, an inhibitor of PI-PLC, did not inhibit the initial ATP-induced response; whereas D609, an inhibitor of PC-PLC, caused a significant decrease in the initial ATP-induced response, suggesting that P2Y receptors are coupled to PC-PLC. Although P2X were present in HKC-8, The P2X agonist, alpha,beta me-ATP, failed to cause a rise in [Ca(2+)](i). However, PPADS at a concentration of 100 microM inhibits the ATP-induced rise in [Ca(2+)](i). Our results indicate the presence of functional P2Y receptors in HKC-8 cells. ATP-induced [Ca(2+)](i) elevation via P2Y is tightly associated with PC-PLC and TRP channel.
Turvey
Matthew R
MR
Department of Physiology, University of Birmingham, Birmingham, UK.
Wang
Yanyun
Y
Gu
Yuchun
Y
D000255:Adenosine Triphosphate / Q000378:metabolism; D020013:Calcium Signaling / Q000502:physiology*; D002460:Cell Line; D045604:Extracellular Fluid / Q000737:chemistry; D015870:Gene Expression; D006801:Humans; D007687:Kidney Tubules, Proximal / Q000378:metabolism*; D011685:Purine Nucleotides / Q000378:metabolism*; D012333:RNA, Messenger / Q000032:analysis; D018048:Receptors, Purinergic P2 / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D050052:TRPC Cation Channels / Q000378:metabolism; D010738:Type C Phospholipases / Q000378:metabolism
D011685:Purine Nucleotides; D012333:RNA, Messenger; D018048:Receptors, Purinergic P2; D050052:TRPC Cation Channels; C489571:TRPC4 ion channel; D000255:Adenosine Triphosphate; D010738:Type C Phospholipases; C105598:phosphatidylcholine-specific phospholipase C