PGC-1α (peroxisome proliferator-activated receptor [PPARγ] coactivator-1α) activates PPARα and mitochondrial transcription factor A (Tfam), which regulate proteins, fatty acid and ATP metabolism (i.e., FAT/CD36, MCAD, and COX I). Recently we found that the salutary effects of estradiol (E2) on cardiac function following trauma-hemorrhage (T-H) are mediated via estrogen receptor (ER)β. In this study we tested the hypothesis that ERβ-mediated cardioprotection is induced via up-regulation of PGC-1α through PPARα or Tfam-dependent pathway. Male rats underwent T-H and received ERα agonist propylpyrazole-triol (PPT), ERβ agonist diarylpropionitrile (DPN), E2, or vehicle. Another group was treated with antisense PGC-1α oligonucleotides prior to administration of DPN. E2 and DPN treatments attenuated the decrease in cardiac mitochondrial ATP, abrogated the T-H-induced lipid accumulation, and normalized PGC-1α, PPARα, FAT/CD36, MCAD, Tfam, and COX I after T-H. In contrast, PPT administration did not abrogate lipid accumulation. Moreover, in PPT-treated animals mitochondrial ATP remained significantly lower than those observed in DPN- or E2-treated animals. Prior administration of antisense PGC-1α prevented DPN-mediated cardioprotection and increase in ATP levels and Tfam but not in PPARα following T-H. These findings suggest that the salutary effects of E2 on cardiac function following T-H are mediated via ERβ up-regulation of PGC-1α through Tfam-dependent pathway. © FASEB.