Heme oxygenase-1 activity after excitotoxic injury: Immunohistochemical localization of bilirubin in neurons and astrocytes and deleterious effects of heme oxygenase inhibition on neuronal survival after kainate treatment

Abstract

An increased expression of the inducible form of heme oxygenase (HO), HO-1, is found in the hippocampus after kainate injection, but thus far it is unclear whether the HO-1 is enzymatically active. The present study was carried out, using monoclonal antibodies to bilirubin and HO-1 and histochemical staining for iron, to compare the products of HO enzymatic activity, bilirubin and iron, with HO-1 expression in the kainate-lesioned hippocampus. There was a close correlation between bilirubin and HO-1 expression, and both bilirubin and HO-1 were observed in damaged neurons at early times, and astrocytes at later times (weeks), after kainate injection. These results indicate that the increased HO-1 in the hippocampus is enzymatically active. Too determine whether HO-1 activity after kainate could have a protective or, perhaps, destructive effect, kainate-injected rats were injected intraperitoneally with a blood-brain barrier-permeable inhibitor of HO, tin protoporphyrin (SnPP), and the effects of such treatment were compared with effects in rats that received kainate and saline injection. It was found that SnPP treatment did not improve neuronal survival. Instead, increased mortality was observed in rats treated with SnPP. Four SnPP-injected rats vs. one saline-injected rats died after kainate treatment. The surviving SnPP-treated rats showed significantly less hippocampal field that containing Nissl or MAP2 staining (an indicator of surviving neurons) compared with the saline-injected rats. These results indicate that HO-1 induction had a net protective effect on neurons in the kainate model of excitotoxic injury.