2me‐SATP activates P2X4R and induces a secondary increase of Na+ pump and Na+/Ca++ currents in cardiac myocytes of P2X4R overexpression transgenic mice

Abstract

2me‐SATP activates the P2X4 receptor (R) to induce an inward current at resting potentials, carried mostly by Na+ as well as some Ca++. This experiment tests if the Na+ pump current (Ip) or Na/Ca exchange current (Iex) would change after this increased Na+ entry caused by the activation of P2X4R. The whole cell voltage clamp was applied to myocytes from the P2X4R overexpressing transgenic mice at room temperature. Ip was measured at −20mV by rapidly switching the superfusing solution from 0 to 5.4mM K+ when the K+, Iex and ICa currents were blocked by CsCl and NiCl2 in pipette and superfusing solution. In 5 of 8 myocytes, 3 μM 2me‐ SATP induced an inward current (−1.20±0.28pA/pF) at −80mV. The average amplitude of Ip in these 5 cells increased from 0.12±0.03 in control to 0.14±0.03nA in 2me‐SATP (P=0.03). The integrated area of Ip also increased from 1371±379 to 1707±418 nA*ms (P=0.047). To measure Iex, 5μM ouabain instead of NiCl2 was used to block Ip. 10mM NiCl2 was applied to myocytes for 1 min by a rapid solution‐switching device and Ni‐sensitive current was taken as Iex. In 5 of 9 cells, 3 μM 2me‐SATP induced an inward current (−1.37±0.34pA/pF), the reversal potential of Iex shifted from −18 mV at control to −28 mV with ATP. Outward Iex increased significantly from 0.7±0.12 at control to 1.0±0.14 pA/pF after ATP at 50 mV (P=0.046), while the inward Iex changed little, from 0.98±0.07 at control to 1.03±0.03pA/pF with ATP at −80mV. These results indicate that 2me‐SATP, by activating P2X4R to increase the intracellular Na+ concentration, induces a secondary increase of Na pump activity and reverse‐mode Na+/Ca++ exchange.