Liver regeneration after partial hepatectomy (PHx) is orchestrated by multiple signals from cytokines and growth factors. We investigated whether increased energy demand on the remnant liver after PHx contributes to regenerative signals. Changes in the tissue's energy state were determined from adenine nucleotide levels. Adenosine triphosphate (ATP) levels in remnant livers decreased markedly and rapidly (to 48% of control by 30 seconds post-PHx) and remained significantly lower than those in sham-operated controls for 24 to 48 hours. The ATP decrease was not reflected in corresponding increases in adenosine diphosphate (ADP) and adenosine monophosphate (AMP), resulting in a marked decline in total adenine nucleotides (TAN). We found no evidence of mitochondrial damage or uncoupling of oxidative phosphorylation. Multiple lines of evidence indicated that the decline in TAN was not caused by increased energy demand, but by ATP release from the liver. The extent of ATP loss was identical after 30% or 70% PHx, whereas fasting or hyperglycemia, conditions that greatly alter energy demand for gluconeogenesis, affected the ATP/ADP decline but not the loss of TAN. Presurgical treatment with the α-adrenergic antagonist phentolamine completely prevented loss of TAN, although changes in ATP/ADP were still apparent. Importantly, phentolamine treatment inhibited early signaling events associated with the priming stages of liver regeneration and suppressed the expression of c-fos. Pretreatment with the purinergic receptor antagonist suramin also partly suppressed early regenerative signals and c-fos expression, but without preventing TAN loss. Conclusion: The rapid loss of adenine nucleotides after PHx generates early stress signals that contribute to the onset of liver regeneration. (HEPATOLOGY 2008.)