Extracellular ATP mediates cellular K+/Na+ homeostasis in two contrasting poplar species under NaCl stress

Abstract

eATP mediates cellular K + and Na + homeostasis in two contrasting poplar species differing in salt tolerance. Using the non-invasive micro-test technology (NMT), the effects of extracellular ATP (eATP) on salt-altered flux profiles of K+, Na+, and H+ were investigated in salt-tolerant poplar species, Populus euphratica and salt-sensitive P. popularis. A short-term NaCl (100 mM NaCl, 12 h) resulted in a Na+ efflux and a correspondingly increased H+ influx in P. euphratica cells, but the effect was not seen in P. popularis. ATP (50 μM) enhanced exchange of Na+ with H+ in salt-stressed cells of two species (6, 12 h), especially in P. popularis. However, the ATP-stimulated Na+ efflux and H+ influx were significantly inhibited by amiloride (a Na+/H+ antiporter inhibitor) or sodium orthovanadate (a plasma membrane H+-ATPase inhibitor), indicating that the ATP induction of Na+ extrusion resulted from an active Na+/H+ antiport across the plasma membrane (PM). NaCl accelerated K+ efflux in the two species, with a more pronounced effect in the salt-sensitive poplar. The salt-induced K+ efflux was markedly restricted by the K+ channel blocker, tetraethylammonium chloride, indicating that the K+ efflux is mediated by depolarization-activated outward rectifying K+ channels and non-selective cation channels. ATP benifited poplar cells, especially the salt-sensitive P. popularis, in maintaining K+ homeostasis under external salinity. This was likely the result of activated H+ pump in the PM, which restricted the K+ efflux through the inhibition of depolarization-activated K+ channels in both species. Na+ and K+ flux recordings revealed that non-hydrolysing analogues of ATP, αβ-meATP (50 μM), and ATPγS (50 μM) produced an effect similar to that of the hydrolysable form but with a more pronounced effect. However, ADP- and AMP-stimulated cells (50 μM) exhibited behaviors different from those invoked by ATP, αβ-meATP, and ATPγS treatments. eATP signalling in K+ and Na+ homeostasis was blocked by the antagonists of animal P2 receptors, PPADS, and suramin. Moreover, ATP-stimulated Na+ extrusion and reduction of K+ loss in NaCl-stressed cells were inhibited by LaCl3 (an inhibitor of Ca2+-permeable channels) and DPI (an inhibitor of PM NADPH oxidase), indicating that ATP signalling was mediated via second messengers, H2O2 and Ca2+, in the two poplars differing in salt tolerance.