To investigate the mechanism by which the combination of Penis et Testis Cervi and Testudinis Carapax et Plastrum alleviates physical fatigue in kidney yin-deficient mice through the adenosine 5'-monophosphate(AMP)-activated protein kinase(AMPK)/peroxisome proliferator-activated receptor-γ coactivator 1α(PGC1α) signaling pathway.Methods:Male Kunming mice were administered dexamethasone acetate via gavage to establish a kidney yin deficiency model and were divided into seven groups：control，model，positive control(Zuogui Pills)，low-dose drug pair，high-dose drug pair，Penis et Testis Cervi alone，and Testudinis Carapax et Plastrum alone.Body weight changes and exhaustive weight-loaded swimming time were recorded.Serum levels of blood lactic acid(BLA)，lactate dehydrogenase(LDH)，creatinine(Cr)，creatine kinase(CK)，cyclic adenosine monophosphate(cAMP)，and cyclic guanosine monophosphate(cGMP) were measured.Additionally，skeletal muscle glycogen content，Na+-K+-ATPase and Ca2+-Mg2+-ATPase activities were assessed.Western blot analysis was used to detect phosphorylated AMPK(p-AMPK)，total AMPK，and PGC1α protein expression in skeletal muscle tissue.Results:Compared with the model group，all treatment groups showed increased body weight and improved yin deficiency symptoms.The exhaustive swimming time was prolonged，while BLA，CK，and Cr levels decreased.LDH，cGMP，and muscle glycogen levels increased，whereas the cAMP/cGMP ratio decreased.Na+-K+-ATPase and Ca2+-Mg2+-ATPase activities were reduced.Western blot analysis showed a decrease in p-AMPK/AMPK and PGC1α protein expression in skeletal muscle.Conclusion:The combination of Penis et Testis Cervi and Testudinis Carapax et Plastrum enhances glycogen storage and metabolic waste clearance.It may regulate glucose transport and oxidative capacity in skeletal muscle through the AMPK/PGC1α signaling pathway，helping to mitigate the excessive energy metabolism associated with yin deficiency，thereby improving exercise endurance and promoting fatigue recovery.