Hippocampal mtDNA depletion promotes nicotine addiction by suppressing mitochondrial biogenesis through oxidative stress

Nicotine addiction is a complex neurobiological process in which the hippocampus plays a pivotal role in consolidating drug-context associative memories. Although mitochondrial dysfunction has been implicated in addiction, the specific role and molecular mechanisms underlying changes in mitochondrial DNA (mtDNA) copy number during nicotine exposure remain poorly defined. This study aimed to elucidate the relationship between hippocampal mtDNA copy number alterations and nicotine addiction behaviors, as well as the molecular mechanisms driving mtDNA depletion. Using a nicotine-induced conditioned place preference (CPP) mouse model, we observed a marked reduction in mtDNA copy number specifically in the hippocampus, but not in the nucleus accumbens (NAc) or ventral tegmental area (VTA). To determine causality, recombinant adeno-associated virus (rAAV)-mediated genetic manipulation was employed to selectively overexpress (TFAM-OE) or knock down (TFAM-KD) mitochondrial transcription factor A (TFAM) in the hippocampus. TFAM-OE, which significantly increased baseline mtDNA copy number, completely abolished the development of nicotine-induced CPP, establishing a direct causal link between hippocampal mtDNA copy number and nicotine reward behavior. Mechanistically, chronic nicotine exposure induced a time-dependent reduction in mtDNA copy number and pronounced mitochondrial ultrastructural damage in hippocampal neurons. This depletion was primarily driven by suppression of the mitochondrial biogenesis pathway-particularly the Sirt1-Nrf1-Tfam signaling axis-rather than activation of mitophagy. Notably, oxidative stress, evidenced by depleted glutathione (GSH) levels, was identified as the upstream trigger mediating this suppression. Pharmacological intervention with mitochondrial-targeted antioxidants (melatonin and mito-TEMPO) restored mitochondrial biogenesis, prevented mtDNA copy number loss, and normalized ATP production. Melatonin can further significantly inhibit the formation of nicotine CPP behavior. Collectively, these findings establish a causal link between hippocampal mtDNA depletion and nicotine reward memory, demonstrating that nicotine suppresses mitochondrial biogenesis via oxidative stress-mediated inhibition of the Sirt1-Nrf1-Tfam axis. This work identifies a previously unrecognized metabolic mechanism underlying nicotine addiction and may also advance our understanding of mitochondrial contributions to other substance use disorders.