1. ¹Laboratory for Psychoneuroimmunology, University Medical Center Utrecht, Utrecht, The Netherlands
2. ²Department of Neonatology, University Medical Center Utrecht, Utrecht, The Netherlands
3. ³Perinatal Center, Sahlgrenska University Hospital, Göteborg, Sweden

Correspondence: Dr CJ Heijnen, Laboratory for Psychoneuroimmunology, University Medical Center Utrecht, Room KC 03.063.0, PO Box 85090, 3508 AB Utrecht, The Netherlands. E-mail: c.heijnen@umcutrecht.nl

Received 15 February 2006; Revised 18 April 2006; Accepted 28 April 2006; Published online 31 May 2006.

Abstract

We have shown earlier that 2-iminobiotin (2-IB) reduces hypoxia–ischemia (HI)-induced brain damage in neonatal rats, and presumed that inhibition of nitric oxide synthases (NOS) was the underlying mechanism. We now investigated the effect of 2-IB treatment in P7 rat pups to determine the role of gender and the neuroprotective mechanism. Pups were subjected to HI (occlusion of right carotid artery and 120 mins FiO₂ 0.08) and received subcutaneous (s.c.) 10 mg/kg 2-IB at 0, 12 and 24 h after hypoxia. After 6 weeks, neuronal damage was assessed histologically. We determined cerebral nitrite and nitrate (NO_x) and nitrotyrosine, heat-shock protein 70, cytosolic cytochrome c, cleaved caspase 3, nuclear translocation of apoptosis-inducing factor (AIF) and the effect of 2-IB on NOS activity in cultured cells. 2-Iminobiotin treatment reduced long-term brain damage in female but not male rats. Unexpectedly, 2-IB treatment did not reduce cerebral NO_x or nitrotyrosine levels, and did not inhibit NOS activity in vitro. The gender-dependent neuroprotective effect of 2-IB was reflected in inhibition of the HI-induced increase in cytosolic cytochrome c and cleaved caspase 3 in females only. Hypoxia–ischemia-induced activation of AIF was observed in males only and was not affected by 2-IB. Post-HI treatment with 2-IB provides gender-specific long- and short-term neuroprotection in female P7 rats via inhibition of the cytochrome c-caspase 3 neuronal death pathway. 2-Iminobiotin did not alter cerebral NO_x nor inhibited NOS in intact cells. Therefore, we conclude that it is highly unlikely that the neuroprotective effect of 2-IB involves NOS inhibition.