The reaction of nitric oxide (NO) with superoxide yields the cytotoxic oxidant peroxynitrite, produced during inflammation and shock. A novel pathway of peroxynitrite cytotoxicity involves activation of the nuclear enzyme poly(ADP) ribosyltransferase, and concomitant ADP-ribosylation. NAD+ consumption and exhaustion of intracellular energy stores. In the present report we provide evidence that pre-exposure of J774 macrophages to heat shock reduces peroxynitrite-induced activation of poly(ADP) ribosyltransferase and protects against the peroxynitrite-induced suppression of mitochondrial respiration. The protection was significant at 8 h after heat shock, but was absent at 24 h after heat shock. Thus, the protection showed a temporal correlation with the expression of heat shock protein 70, the expression of which was maximal at 8 h. Exposure to heat shock did not alter the expression of poly(ADP) ribosyltransferase over 24 h. In summary, the heat shock phenotype or heat shock proteins may protect against peroxynitrite induced cytotoxicity.