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 Ca²⁺ concentration ([Ca²⁺]_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, P2X₃, P2X₅, and P2X₇ receptor subtype mRNA was increased in differentiated cells. On the other hand, the expression of G-protein-coupled P2Y₂ mRNA was downregulated in differentiated cells. Increases in [Ca²⁺]_i evoked by αβ-methylene ATP (αβ-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 P2X₁, P2X₂, P2X₃, and P2X₇ 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.