Rhabdomyolysis is characterised by acute kidney injury (AKI) resulting from skeletal muscle injury. Lipid peroxidation-mediated oxidant injury and pro-inflammatory cytokine-mediated inflammatory response play critical roles in the pathogenesis of rhabdomyolysis-induced AKI. The present study aimed to investigate the short-term effects of both lipid peroxidation and inflammatory responses on rhabdomyolysis-induced AKI in a rat model of glycerol-induced rhabdomyolysis. Rhabdomyolysis was induced by the intramuscular injection of 50% glycerol in saline (10 mL/kg) into the hind limbs of rats. Rats were killed 1 or 3 hours after glycerol injection. Time-dependent increases in serum biochemical parameters, including blood urea nitrogen, creatinine, lactate dehydrogenase and creatine phosphokinase levels, were observed 1 hour after glycerol injection. In kidneys, glycerol injection resulted in histopathological changes such as renal tubular injury and renal tubular myoglobin deposition. Levels of Nε-(hexanoyl)lysine-modified, 4-hydroxy-2-nonenal-modified, and nitrotyrosine-modified proteins in rat kidneys were unaltered at 1 hour after glycerol injection, but increased significantly at 3 hours. Increases in renal nitric oxide production and the expression levels of inducible nitric oxide synthase, interleukin-6 and tumour necrosis factor-α in the renal parenchyma were observed at 1 hour after glycerol injection and plateaued at 3 hours. Our findings suggest that the pro-inflammatory cytokine-mediated inflammatory response may cause rhabdomyolysis-induced AKI very shortly after glycerol injection, and lipid peroxidation-mediated oxidant injury may promote the development of these pathophysiological processes.