Rhabdomyolysis is associated with acute renal failure. The following study first characterized myoglobin in vitro toxicity using renal cortical slices isolated from male Fischer 344 rats. This model provided interaction between various cells within the nephron and provides myoglobin access predominantly through the basolateral membrane. Second, this study examined the effect of deferoxamine (DFX) and glutathione on myoglobin toxicity to determine the role of radicals and iron. Renal cortical slices were incubated for 30-120 min with 0, 4, 10 or 12 mg/ml myoglobin. Myoglobin was pretreated with 4 mM ascorbic acid prior to addition to the slices to ensure that myoglobin was in its reduced state. In other experiments tissues were pretreated for 15 min with 0.1 mM of the iron chelator DFX or 30 min with 1 mM glutathione prior to co-incubation with myoglobin. Finally, slices were pretreated with 1 mM glutathione for 30 min, rinsed and incubated only with myoglobin. Early event changes occurred within a 60 min exposure and included a decline in pyruvate-stimulated gluconeogenesis, increased lipid peroxidation levels and decreased glutathione levels. Loss of ATP levels and increased lactate dehydrogenase (LDH) release required a 120 min exposure to myoglobin. DFX reduced myoglobin induced effects on LDH leakage but had no effect on gluconeogenesis suggesting that myoglobin toxicity had an iron dependent (LDH) and independent (gluconeogenesis) pathway. Pretreatment with glutathione provided complete protection and was mediated by intracellular events.