Inhibition of activation has been reported when neutrophils are suspended in Na + -free media. We considered the possibility that impairment of cellular pH (pH i ) regulation due to elimination of Na + /H + exchange underlies this effect. In the absence of Na + , the phorbol ester-induced respiratory burst was partially inhibited and a concomitant cytoplasmic acidification recorded. Using nigericin/K + to clamp pH i we demonstrated that the acidification accounts for the inhibition of O 2 uptake. Moreover, in Na + -free media, relieving the acidification by means of ionophores restored maximal O 2 consumption. It was concluded that Na + is not directly involved in signal transduction during stimulation. Instead, omission of Na + affects neutrophile activation indirectly, by impairing pH i regulation.