Characterization of multiple P2X receptors in cultured normal human epidermal keratinocytes.
ATP-gated ion channels (P2X) are expressed in human epidermis and cultured keratinocytes. The aim of this study was to characterize native P2X receptors in normal human epidermal keratinocytes (NHEK) using whole-cell patch clamp technique, RT-PCR, and determination of intracellular Ca(2+) concentration ([Ca(2+)](i)). Application of ATP resulted in an inward current with a reversal potential of 0 mV. Response to ATP showed two types of currents: the slowly desensitizing response and the rapidly desensitizing response. The slowly desensitizing response was blocked by iso-pyridocaphosphate-6-azophenyl-2', 5' disulfonic acid (PPADS), a P2X receptor antagonist. We found that the expression of multiple P2X(2), P2X(3), P2X(5), and P2X(7) receptor subtype mRNA was increased in differentiated cells. On the other hand, the expression of G-protein-coupled P2Y(2) mRNA was downregulated in differentiated cells. Increases in [Ca(2+)](i) evoked by alphabeta-methylene ATP (alphabeta-meATP) and 2', 3'-O-(4-benzoylbenzoyl) ATP (BzATP) were elevated, whereas elevation of [Ca(2+)](i) evoked by uridine 5'-triphosphate (UTP) was decreased in differentiated cells. Application of ATP or UVB radiation increased the expression of P2X(1), P2X(2), P2X(3), and P2X(7) receptors in NHEK. Changes in the expression levels and cation influx via multiple P2X receptors might be involved in the regulation of differentiation and one of the epidermal external sensors.
Inoue
Kaori
K
Shiseido Research Center, Yokohama, Japan. kaori.inoue@to.shiseido.co.jp
Denda
Mitsuhiro
M
Tozaki
Hidetoshi
H
Fujishita
Kayoko
K
Koizumi
Schuichi
S
Inoue
Kazuhide
K
D000255:Adenosine Triphosphate / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism; D002454:Cell Differentiation / Q000502:physiology; D002455:Cell Division / Q000502:physiology; D002478:Cells, Cultured; D000078404:Epidermal Cells; D015870:Gene Expression / Q000187:drug effects / Q000528:radiation effects; D006801:Humans; D015603:Keratinocytes / Q000166:cytology / Q000502:physiology*; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D012333:RNA, Messenger / Q000032:analysis; D018048:Receptors, Purinergic P2 / Q000235:genetics* / Q000502:physiology*; D058469:Receptors, Purinergic P2X; D058476:Receptors, Purinergic P2X2; D058477:Receptors, Purinergic P2X3; D058485:Receptors, Purinergic P2X5; D058486:Receptors, Purinergic P2X7; D014466:Ultraviolet Rays
C546401:P2RX2 protein, human; C546407:P2RX3 protein, human; C546465:P2RX5 protein, human; C546468:P2RX7 protein, human; D012333:RNA, Messenger; D018048:Receptors, Purinergic P2; D058469:Receptors, Purinergic P2X; D058476:Receptors, Purinergic P2X2; D058477:Receptors, Purinergic P2X3; D058485:Receptors, Purinergic P2X5; D058486:Receptors, Purinergic P2X7; D000255:Adenosine Triphosphate; D002118:Calcium