PT - JOURNAL ARTICLE AU - Shobu Namura AU - Jinmin Zhu AU - Klaus Fink AU - Matthias Endres AU - Anu Srinivasan AU - Kevin J. Tomaselli AU - Junying Yuan AU - Michael A. Moskowitz TI - Activation and Cleavage of Caspase-3 in Apoptosis Induced by Experimental Cerebral Ischemia AID - 10.1523/JNEUROSCI.18-10-03659.1998 DP - 1998 May 15 TA - The Journal of Neuroscience PG - 3659--3668 VI - 18 IP - 10 4099 - http://www.jneurosci.org/content/18/10/3659.short 4100 - http://www.jneurosci.org/content/18/10/3659.full SO - J. Neurosci.1998 May 15; 18 AB - We examined the expression, activation, and cellular localization of caspase-3 (CPP32) using immunohistochemistry, immunoblots, and cleavage of the fluorogenic substrateN-benzyloxycarbonyl-Asp-Glu-Val-Asp-7-amino-4-trifluoromethyl coumarin (zDEVD-afc) in adult mouse brain after temporary (2 hr) middle cerebral artery occlusion produced by filament insertion into the carotid artery. Immunoreactive caspase-3p32 but not its cleavage product caspase-3p20 was constitutively expressed in neurons throughout brain and was most prominent in neuronal perikarya within piriform cortex. Caspase-like enzyme activity was elevated in brain homogenate 0–3 hr after reperfusion and reached a peak within 30 to 60 min. Caspase-3p20 immunoreactivity became prominent in neuronal perikarya within the middle cerebral artery territory at the time of reperfusion and on immunoblots 1–12 hr later. DNA laddering (agarose gels) and terminal deoxynucleotidyl transferase-mediated dUTP—biotin nick-end labeling (TUNEL)-stained cells were detected 6–24 hr after reperfusion. At 12–24 hr, immunoreactive p20 was visualized in TUNEL-positive cells, a finding also observed in apoptotic mouse cerebellar granule cells on postnatal day 5. Together, these observations suggest the existence of a time-dependent evolution of ischemic injury characterized by the close correspondence between caspase-like enzyme activation and an associated increase in immunoreactive product (caspase-3p20) beginning at or before reperfusion and followed several hours later by morphological and biochemical features of apoptosis.