These studies have addressed the role of caspase-3 activation in neuronal death after cerebral ischemia in different animal models. The authors were unable to show activation of procaspase-3 measured as an induction of DEVDase (Asp-Glu-Val-Asp) activity after focal or transient forebrain ischemia in rats. DEVDase activity could not be induced in the cytosolic fraction of the brain tissue obtained from these animals by exogenous cytochrome c/dATP and Ca2+. However, the addition of granzyme B to these cytosolic fractions resulted in a significant activation of DEVDase, confirming that the conditions were permissive to analyze proteolytic cleavage of the DEVD-AMC (7-amino-4-methyl-coumarin) substrate. Consistent with these findings, zVal-Ala-Asp-fluoromethylketone administered after focal ischemia did not have a neuroprotective effect. In contrast to these findings, a large increase in DEVDase activity was detected in a model of hypoxic-ischemia in postnatal-day-7 rats. Furthermore, in postnatal-day-7 animals treated with MK-801, in which it has been suggested that excessive apoptosis is induced, the authors were unable to detect activation of DEVDase activity but were able to induce it in vitro by the addition of cytochrome c/dATP and Ca2+ to the cytosolic fraction. Analysis of cytochrome c distribution did not provide definitive evidence for selective cytochrome c release in the permanent focal ischemia model, whereas in the transient model a small but consistent amount of cytochrome c was found in the cytosolic fraction. However, in both models the majority of cytochrome c remained associated with the mitochondrial fraction. In conclusion, the authors were unable to substantiate a role of mitochondrially derived cytochrome c and procaspase-3 activation in ischemia-induced cell death in adult brain, but did see a clear induction of caspase-3 in neonatal hypoxia.

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Nerve growth factor (NGF) binds to two neurotrophin receptors: p75NTR and p140Trk (TrkA). Both receptors dimerize in response to NGF binding. TrkA homodimers and heteromers of TrkA and p75NTR promote cell survival whereas homodimers of p75NTR mediate apoptosis upon binding of NGF. The interaction between receptor and NGF can be inhibited either on the level of the ligand by altering NGF conformation so that NGF is no longer recognized by the receptor or on the level of the receptor by blocking the binding site of p75NTR or TrkA. The effect of altering NGF conformation on NGF signaling was investigated in two neuron‐like cell lines: in human SK‐N‐MC cells that express only p75NTR and in rat PC12 cells that express both p75NTR and TrkA. In the present study we demonstrate that Ro 08‐2750 binds to the NGF dimer thereby probably inducing a change in its conformation such that NGF cannot bind to p75NTR anymore. In SK‐N‐MC cells this leads to inhibition of NGF‐induced programmed cell death. In PC12 cells enhanced signaling through TrkA was observed. J. Neurosci. Res. 61:263–272, 2000. © 2000 Wiley‐Liss, Inc.

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