Upon massive DNA damage, hyperactivation of the nuclear enzyme poly(ADP-ribose) polymerase (PARP)-1 causes severe depletion of intracellular NAD and ATP pools as well as mitochondrial dysfunction. Thus far, the molecular mechanisms contributing to PARP-1-dependent impairment of mitochondrial functioning have not been identified. We found that degradation of the PARP-1 product poly(ADP-ribose) through the concerted actions of poly(ADP-ribose) glycohydrolase and NUDIX (nucleoside diphosphate-X) hydrolases leads to accumulation of AMP. The latter, in turn, inhibits the ADP/ATP translocator, prompting mitochondrial energy failure. For the first time, our findings identify NUDIX hydrolases as key enzymes involved in energy derangement during PARP-1 hyperactivity. Also, these data disclose unanticipated AMP-dependent impairment of mitochondrial exchange of adenine nucleotides, which can be of relevance to organelle functioning and disease pathogenesis.