Migration of naive CD4+ T lymphocytes into lymphoid tissue is essential for their activation and subsequent roles in adaptive immunity. The adhesion molecule L-selectin (CD62L), critical for this process, is highly expressed on naive CD4+ T lymphocytes and is downregulated upon T lymphocyte activation. We demonstrate protein expression of P2X7R on naive CD4+ T lymphocytes and show functional channel activity in whole-cell patch clamp recordings. CD62L downregulation occurs rapidly in response to extracellular ATP, a process that is blocked by selective antagonists of P2X7R. This loss of surface CD62L expression was not associated with externalization of phosphatidylserine. While investigating the mechanisms for this process, we revealed that pharmacological modulation of mitochondrial complex I or III, but not inhibition of NADPH oxidase, enhanced P2X7Rdependent CD62L downregulation by increasing ATP potency. Enhanced superoxide generation in the mitochondria of rotenoneand antimycin A-treated cells was observed and may contribute to the enhanced sensitivity of P2X7R to ATP. P2X7R-dependent exposure of phosphatidylserine was also revealed by preincubation with mitochondrial uncouplers prior to ATP treatment. This may present a novel mechanism whereby P2X7R-dependent phosphatidylserine exposure occurs only when cells have enhanced mitochondrial reactive oxygen species generation. The clearance of apoptotic cells may therefore be enhanced by this mechanism which requires functional P2X7R expression.