Ž . Molecular Brain Research 59 1998 119–134 Research report Co-ordinated and cellular specific induction of the components of the IGFrIGFBP axis in the rat brain following hypoxic–ischemic injury Erica J. Beilharz a , Vincenzo C. Russo b , Gary Butler b , Naomi L. Baker b , Bronwyn Connor c , Ernest S. Sirimanne a , Mike Dragunow c , George A. Werther b , Peter D. Gluckman a , Chris E. Williams a, ) , Arjan Scheepens a a Research Centre for DeÕelopmental Medicine and Biology, School of Medicine, UniÕersity of Auckland, PriÕate Bag 92019, Auckland, New Zealand b Centre for Hormone Research RCH, Melbourne, Australia c Pharmacology Department, UniÕersity of Auckland, Auckland, New Zealand Accepted 5 May 1998 Abstract Ž . Ž . Insulin-like growth factor 1 IGF-1 is induced after hypoxic–ischemic HI brain injury, and therapeutic studies suggest that IGF-1 may restrict delayed neuronal and glial cell loss. We have used a well-characterised rat model of HI injury to extend our understanding of the modes of action of the IGF system after injury. The induction of the IGF system by injury was examined by in situ hybridization, Ž . immunohistochemistry, Northern blot analysis, RNase protection assay and reverse transcriptase-polymerase chain reaction RT-PCR . IGF-1 accumulated in blood vessels of the damaged hemisphere within 5 h after a severe injury. By 3 days, IGF-1 mRNA was expressed by reactive microglia in regions of delayed neuronal death, and immunoreactive IGF-1 was associated with these microglia and reactive astrocytes juxtaposed to surviving neurones surrounding the infarct. Total IGF-1 receptor mRNA was unchanged by the injury. IGFBP-2 mRNA was strongly induced in reactive astrocytes throughout the injured hemisphere, and IGFBP-3 and IGFBP-5 mRNA were moderately induced in reactive microglia and neurones of the injured hippocampus, respectively. IGFBP-6 mRNA was induced in the damaged hemisphere by 3 days and increased protein was seen on the choroid plexus, ependyma and reactive glia. In contrast, insulin II was not induced. These results indicate cell type-specific expression for IGF-1, IGFBP-2,3,5 and 6 after injury. Our findings suggest that the IGF-1 produced by microglia after injury is transferred to perineuronal reactive astrocytes expressing IGFBP-2. Thus, modulation of IGF-1 action by IGFBP-2 might represent a key mechanism that restricts neuronal cell loss following HI brain injury. q 1998 Elsevier Science B.V. All rights reserved. Keywords: IGF; IGFBP; Brain; Rat; Growth factor; Hypoxia–ischemia; Neurone; Glia Abbreviations: BBB, blood-brain barrier; BDHC, benzidine dihydrochloride; BDNF, brain-derived neurotrophic factor; BP, binding protein; CA, cornu ammonis; CNS, central nervous system; cRNA, complementary ribonucleic acid; CSF, cerebrospinal fluid; DAB, di-amino benzidine; DNA, deoxyribonu- Ž . cleic acid; DND, delayed neuronal death; GFAP, glial fibrillary acidic protein; GPE, glycine–proline–glutamate N-terminal tripeptide of IGF-1 ; HI, hypoxic–ischemic; IEGs, immediate early genes; ICV, intracerebroventricular; IGF, insulin-like growth factor; IGFBP, insulin-like growth factor binding protein; IGF-1R, insulin-like growth factor type I receptor; IGF-IIR, insulin-like growth factor type II receptor; IgG, immunoglobulin class G; IR, insulin receptor; IRR, insulin-related receptor; IP , inositol triphosphate; LDTN, laterodorsal thalamic nucleus; M, molar; MBP, myelin basic protein; mRNA, 3 messenger ribonucleic acid; NMDA, N-methyl-D-aspartate; PBS, phosphate-buffered saline; PCD, programmed cell death; PCR, polymerase chain reaction; PDGF, platelet-derived growth factor; rh, recombinant human; RNA, ribonucleic acid; Rnasin, RNase inhibitor; RPA, RNase protection assay; RT, room temperature; RT-PCR, reverse transcriptase-polymerase chain reaction; S.E.M., standard error of the mean ) Corresponding author. Fax: q64-9-3737-497; E-mail: ce.williams@auckland.ac.nz 0169-328Xr98r$19.00 q 1998 Elsevier Science B.V. All rights reserved. Ž . PII: S0169-328X 98 00122-3