Adriamycin (ADR) has been shown to produce free radicals in NADPH microsomal systems, to increase oxygen consumption of both hepatic microsomes and heart sarcosomes and to stimulate superoxide formation in cardiac, submitochondria particles. These reactive products could produce the cardiotoxicity of ADR by oxidizing various membrane structures, especially if the heart lacks sufficient protective reducing substances such as thiols. We examined 1) the effect of ADR on reduced glutathione (G-SH) levels in various tissues including heart, 2) the ability of the sulfhydryl (SH) donor, cysteamine, to alter soluble SH levels in heart tissue after ADR administration and 3) the effects of SH donors (cysteamine and N-acetyl cysteine and G-SH depletion by diethyl maleate on ADR-induced lethality in Swiss ICR-HA mice. A single injection of ADR (15 mg/kg i.p.) elicited a statistically significant fall in liver (P < .05), heart (P < .02) and erythrocyte (P < .01) G-SH levels. Treatment with cysteamine protected against the fall in soluble SH groups in heart tissue. Cysteamine (50 mg/kg, i.p., every 8 hr for 6 days) or N-acetylcysteine (100 mg/kg, i.p., 1 hr before and 7 hr after ADR) protected against ADR-induced lethality and decreased the appearance of microscopic myocardial lesions. When endogenous levels of G-SH were depleted by diethyl maleate (300 mg/kg i.p., every 8 hr for 4 days), ADR lethality was markedly potentiated. Diethyl maleate alone did not cause death. We conclude 1) ADR significantly lowers G-SH levels in erythrocytes, liver and heart tissue, 2) the lowering of cardiac SH groups by ADR can be prevented by cysteamine and 3) ADR toxicity can be potentiated by diethyl maleate, a G-SH depletor, and reduced by cysteamine or N-acetyl cysteine, SH donors. These results suggest that the G-SH system may be involved in the modulation of ADR-induced toxicity.