Cooperativity between inhibition of cytosolic K+ efflux and AMPK activation during suppression of hypoxia-induced cellular apoptosis

Cellular potassium homeostasis has recently emerged as a critical regulator of apoptosis in response to variety of stimuli. However, functional hierarchy of this phenomenon in the apoptotic cascade and therefore, its significance as a pathway for intervention is not fully established. Chronic hypoxia, a known threat to cell survival, also modulates cellular potassium homeostasis. In this study, we tested if hypoxia-induced apoptosis in lymphocytes can be prevented by modulating cellular K+ homeostasis. We observed that chronic hypoxia accelerated the rate of apoptosis in resting murine splenocytes concomitant with cytosolic K+ efflux. We tested several modalities including elevated extracellular potassium besides various K+ channel inhibitors to curtail hypoxia-induced K+ efflux and interestingly, established that the supplementation of KCl in extracellular medium was most effective in preventing hypoxia-induced apoptosis in these cells. Subsequent mechanistic dissection of pathways underlying this phenomenon revealed that besides effectively inhibiting hypoxia-induced efflux of K+ ion and its downstream cell-physiological consequences; elevated extracellular KCl modulated steady state levels of cellular ATP and culminated in stabilization of AMPKα with pro-survival consequences. Also, interestingly, global gene expression profiling revealed that KCl supplementation down regulated a distinct p53-regulated cellular sub-network of genes involved in regulation of DNA replication. Additionally, we present experimental evidence for the functional role of AMPK and p53 activation during suppression of hypoxia-induced apoptosis. In conclusion, our study highlights a novel bimodal effect wherein cooperativity between restoration of K+ homeostasis and a sustainable 'metabolic quiescence' induced by AMPK activation appeared indispensible for curtailing hypoxia-induced apoptosis.