Functional properties of heteromeric P2X1/5 receptors expressed in HEK cells and excitatory junction potentials in guinea-pig submucosal arterioles

Abstract

P2X receptors are ATP-gated cation channels; they form as homomers or heteromers from a family of seven related subunits. In particular, heteromeric channels comprising P2X₂ and P2X₃ subunits, or P2X₁ and P2X₅ subunits, show distinctive physiological and pharmacological properties in heterologous expression systems. There is substantial evidence that one of the native P2X receptors in sensory neurones corresponds to the P2X_2/3 heteromer, but there is no evidence for P2X_1/5 heteromers in native tissue. We recorded currents in response to activation of heteromeric P2X_1/5 receptors expressed in HEK293 cells to characterize further their functional properties. The ATP concentration–response curve had a threshold concentration of 1 nM, and a Hill slope of one. TNP-ATP was a weak partial agonist, and a non-competitive antagonist which inhibited maximal ATP currents by 60%. Increasing or decreasing pH from 7.3 shifted the ATP concentration–response curves to the right by fivefold and decreased the maximum current by 40%. Calcium permeability was lower than that observed for other P2X receptors (P_Ca/P_Na ratio=1.1). The nanomolar sensitivity of this receptor revealed a steady release of ATP from HEK293 cells, providing an extracellular concentration which ranged from 3 to 300 nM. Noradrenaline (0.3–30 μM) increased ATP-evoked currents by 35%; this facilitation occurred within 20 ms. We also recorded excitatory junction potentials (EJPs) from guinea-pig submucosal arterioles. EJPs were inhibited by suramin and PPADS (IC₅₀s of 0.2 μM and 20 μM) but TNP-ATP (0.1–10 μM) inhibited EJPs by <30%. Noradrenaline (0.3–30 μM in the presence of phentolamine and propranolol) decreased EJPs in control preparations but facilitated EJPs by 5–20% in submucosal arterioles from reserpinized guinea-pigs. These properties are discussed in relation to P2X receptors underlying EJPs at autonomic neuroeffector junctions.