Introduction The Na + , K + -ATPase (Na + , K + -pump) acts as an electrogenic ion transporter in the plasma membrane of all mammalian cells, each cycle of Na + , K + -pump activity extrudes three Na + ions from the cell, moves two K + ions into the cell and uses 1 ATP (Rakowski et al., 1989). The primary role of the Na + , K + -pump is therefore to maintain high intracellular K + and low intracellular Na + . Failure of the Na + , K + -pump results in depletion of intracellular K + , accumulation of intracellular Na + , and, consequently, leads to membrane depolarization and increases in intracellular free Ca 2+ ([Ca 2+ ] i ) due to activation of voltage-gated Ca 2+ channels and a reversed operation of the Na + -Ca 2+ exchanger (Archibald and White, 1974; Lijnen et al., 1986; DiPolo and Beauge, 1991; Xiao et al., 2002). A central role for the Na + , K + -pump in pathogenesis has been widely implicated, particularly in heart ischemia (Ziegelhoffer et al., 2000). Blocking the Na + , K + -pump can induce apoptosis (Olej et al., 1998; Chueh et al., 2001) or a ‘hybrid death’ containing both apoptotic and necrotic components in individual neurons (Xiao et al., 2002). Conversely, a recent study showed that apoptotic thymocytes had decreased protein levels of the Na + , K + -ATPase (Mann et al., 2001). Whether the apoptotic process may alter the Na + , K + -pump activity, however, is still an open question and has so far not been directly investigated. Excessive K + efflux and intracellular K + depletion are thought to be critical steps in cell body shrinkage and apoptotic death (Yu et al., 1997; Dallaporta et al., 1998; Bortner and Cidlowski, 1999). A significant reduction in intracellular K + concentration may be a prerequisite for key apoptotic events including caspase-3 cleavage and endonuclease activation (Dallaporta et al., 1998; Bortner and Cidlowski, 1999). The pro-apoptotic K + efflux may be mediated by voltage-gated K + channels in neurons (Yu et al., 1997; Colom et al., 1998; Nadeau et al., 2000); and other cells (Nietsch et al., 2000; Wang et al., 1999; Diem et al., 2001; Krick et al., 2001). In addition, K + loss may occur through NMDA or AMPA/kainate receptor channels (Yu et al., 1999a; Xiao et al., 2001). Theoretically, K + homeostasis may not be altered if K + efflux can be balanced by sufficient K + uptake. Since the Na + , K + -pump is the only major mechanism for K + uptake, we hypothesized that cells undergoing apoptosis, in addition to an enhanced K + efflux, might additionally suffer from dysfunction of the Na + , K + -pump. To test this hypothesis, we identified the membrane currents associated with the Na + , K + -pump activity in cortical neurons, examined the effects of several apoptotic insults on the Na + , K + -pump current and modeled the putative role of the Na + , K + -pump in neuronal apoptosis. This work was partly presented in an abstract (Wang et al., 2001). 2099 The Na + , K + -ATPase (Na + , K + -pump) plays critical roles in maintaining ion homeostasis. Blocking the Na + , K + -pump may lead to apoptosis. By contrast, whether an apoptotic insult may affect the Na + , K + -pump activity is largely undefined. In cultured cortical neurons, the Na + , K + -pump activity measured as a membrane current I pump was time- dependently suppressed by apoptotic insults including serum deprivation, staurosporine, and C 2 -ceramide, concomitant with depletion of intracellular ATP and production of reactive oxygen species. Signifying a putative relationship among these events, I pump was highly sensitive to changes in ATP and reactive oxygen species levels. Moreover, the apoptosis-associated Na + , K + -pump failure and serum deprivation-induced neuronal death were antagonized by pyruvate and succinate in ATP- and reactive-oxygen-species-dependent manners. We suggest that failure of the Na + , K + -pump as a result of a combination of energy deficiency and production of reactive oxygen species is a common event in the apoptotic cascade; preserving the pump activity provides a neuroprotective strategy in certain pathological conditions. Key words: Na + , K + -ATPase, Apoptosis, Potassium homeostasis, Neuron Summary Apoptotic insults impair Na + , K + -ATPase activity as a mechanism of neuronal death mediated by concurrent ATP deficiency and oxidant stress Xue Qing Wang, Ai Ying Xiao, Christian Sheline, Krzystztof Hyrc, Aizhen Yang, Mark P. Goldberg, Dennis W. Choi* and Shan Ping Yu ‡,§ Center for the Study of Nervous System Injury and Department of Neurology, Washington University School of Medicine, St Louis, MO 63110, USA *Merck Research Labs, West Point, PA 19486, USA ‡ Department of Pharmaceutical Sciences, School of Pharmacy, Medical University of South Carolina, 280 Calhoun Street, Charleston, SC 29425 USA § Author for correspondence (e-mail: yusp@musc.edu) Accepted 29 January 2003 Journal of Cell Science 116, 2099-2110 © 2003 The Company of Biologists Ltd doi:10.1242/jcs.00420 Research Article JCS ePress online publication date 1 April 2003