The polyamines (spermine, putrescine, and spermidine) can have neurotoxic or neuroprotective properties in models of neurodegeneration. However, assessment in a model of hypoxia'ischemia (HI) has not been defined. Furthermore, the putative mechanisms of neuroprotection have not been elucidated. Therefore, the present study examined the effects of the polyamines in a rat pup model of HI and determined effects on key enzymes involved in inflammation, namely, nitric oxide synthase (NOS) and arginase. In addition, effects on mitochondrial function were investigated. The polyamines or saline were administered i.p. at 10mg/kg/day for 6 days post-HI. Histological assessment 7 days post-HI revealed that only spermine significantly (P<0.01) reduced infarct size from 46.14 ± 10.4mm3 (HI + saline) to 4.9 ± 2.7 mm3. NOS activity was significantly increased following spermine treatment in the left (ligated) hemisphere compared with nonintervention controls (P<0.01) and HI + saline (P<0.05). In contrast, spermine decreased arginase activity compared with HI + saline but was still significantly elevated in comparison to nonintervention controls (P<0.01). Assessment of mitochondrial function in the HI + saline group, revealed significant and extensive damage to complex-I (P<0.01) and IV (P<0.001) and loss of citrate synthase activity (P<0.05). No effect on complex II-III was observed. Spermine treatment significantly prevented all these effects. This study has therefore confirmed the neuroprotective effects of spermine in vivo. However, for the first time, we have shown that this effect may, in part, be due to increased NOS activity and preservation of mitochondrial function.