Activation of ERK1/2 after neonatal rat cerebral hypoxia–ischaemia Xiaoyang Wang,* , à Changlian Zhu,* , à Lin Qiu,* , à Henrik Hagberg,* , § Mats Sandberg¶ and Klas Blomgren* ,   Perinatal Center, Departments of *Physiology and  Pediatrics, Go ¨teborg University, Go ¨teborg, Sweden àDepartment of Pediatrics, The Third Affiliated Hospital of Zhengzhou University, Zhengzhou, People’s Republic of China §Perinatal Center, Department of Obstetrics and Gynecology, Sahlgrenska University Hospital/O ¨ stra, Go ¨teborg, Sweden ¶Department of Medical Biophysics, Go ¨teborg University, Go ¨teborg, Sweden Abstract Activation of extracellular signal-related kinases (ERK1/2), also known as p42/44 mitogen-activated protein kinase (MAPK), is considered important for neuronal survival, cell proliferation and apoptosis. In the present study, activation (phosphorylation) of ERK1/2 (P-ERK) was investigated in brains of 7-day-oldrats after hypoxia-ischaemia (HI). In damaged areas, P-ERK-positive neurons appeared imme- diately after HI and the staining remained for at least 8 h. At later time points, 24 and 72 h post-HI, P-ERK-positive neurons were found in the core of the infarct and in the border zone to undamaged tissue. These cells also showed signs of DNA damage and calpain-induced fodrin breakdown, indicat- ive of injury. At 72 h post-HI, P-ERK was also observed in microglia in the border zone to the damaged area and in astrocytes and oligodendrocytes in white matter of both hemispheres. P-ERK was strongly expressed in the subven- tricular zone in both hemispheres after HI at most time points, although the staining in the ipsilateral (damaged hemisphere) was stronger than in the contralateral (non-damaged hemi- sphere). In summary, ERK1/2 activation occurred early in neurons after HI in the neonatal brain, and mainly in cells displaying signs of damage. Keywords: cell death, calpain, DNA damage, MAP kinase, subventricular zone. J. Neurochem. (2003) 86, 351–362. The mitogen-activated protein kinases (MAPKs) regulate a diverse array of functions, such as neuronal survival, cell growth, proliferation (Boulton et al. 1991; Marshall 1995; Segal and Greenberg 1996) and apoptosis (Bading and Greenberg1991;Oppenheim1991;Alessandrini et al.1999). Members of the MAPK family include extracellular signal- regulated kinases (ERKs), c-jun NH2-terminal protein kinase (JNK) and p38 MAPK. JNK and p38 MAPK are strongly activatedbystressstimuli,bacteriallipopolysaccharide(LPS) and cytokines, and have been suggested to contribute to cell death (Oppenheim 1991). ERK, on the other hand, is activated by mitogenic stimuli, subsequently modulating the activity of many transcriptional factors, leading to prolifer- ationanddifferentiation(Boulton et al.1991;Marshall1995; Segal and Greenberg 1996). In general, ERK1/2 activation has been coupled to protective mechanisms. ERK1/2 are activated by dual phosphorylation at threonine and tyrosine residues by the MAPK kinases MEK1/2. ERK-1 and ERK-2 arecloselyrelated,predominantlyactivatedbygrowthfactors and both isoforms are highly expressed in the brain (Hetman et al. 1999; Singer et al. 1999). Neuronal ERK1/2 have been Received November 25, 2002; revised manuscript received March 23, 2003; accepted April 3, 2003. Address correspondence and reprint requests to Klas Blomgren, Perinatal Center, Department of Physiology, Go ¨teborg University, Medicinaregatan 13, SE 413 90 Go ¨teborg, Sweden. E-mail: klas.blomgren@fysiologi.gu.se Abbreviations used: BNDF, brain-derived neurotrophic factor; CNPase, 2¢,3¢-cyclic nucleotide 3¢-phosphodiesterase; CX, cortex; DAB, 3,3-diaminobenzidine tetrahydrochloride; DG, dentate gyrus; DTT, dithiothreitol; ERK, extracellular signal-regulated kinase; FBDP, fodrin breakdown product; GFAP, glial fibrillary acidic protein; HI, hypoxia–ischaemia; HPP, hairpin probe; HTH, hypothalamus; JNK, c-jun NH 2 -terminal protein kinase; LPS, lipopolysaccharide; MAPK, mitogen-activated protein kinase; NH, nucleus habenularis; PBS, phosphate-buffered saline; P-ERK, phosphorylated ERK; PI3-kinase, phosphatdylinositol 3-kinase; SDS–PAGE, sodium dodecyl sulphate polyacrylamide gel electrophoresis; SVZ, subventricular zone; TH, thalamus. Journal of Neurochemistry , 2003, 86, 351–362 doi:10.1046/j.1471-4159.2003.01838.x Ó 2003 International Society for Neurochemistry, J. Neurochem. (2003) 86, 351–362 351