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The majority of retinal ganglion cells (RGCs) degenerate and die after transection of the optic nerve (ON) in the adult rat. This secondary cell death can primarily be ascribed to apoptosis. Recent work strongly suggests a decisive role for a family of cysteine proteases, termed caspases, as mediators of neuronal apoptosis. In this study, we investigated whether activation of caspases contributes to delayed death of RGCs after axotomy. Intraocular application of various caspase inhibitors rescued up to 34% of RGCs that would otherwise have died 14 d after ON transection. Using a modified affinity-labeling technique, we detected a 17 kDa protease subunit upregulated after axotomy. Upregulation was prevented by caspase inhibitor treatment. The 17 kDa protein was identified as a CPP32-like protease by Western blot analysis and affinity labeling with biotinylated acetyl-Asp-Glu-Val-Asp-aldehyde, which specifically inhibits CPP32-like caspases. In vivo application of the irreversible caspase inhibitor benzyloxycarbonyl-Asp-Glu-Val-Asp-chloromethylketone revealed CPP32-like proteases to be major mediators of caspase-induced apoptosis in axotomized RGCs, because this inhibitor showed an even higher neuroprotective potential than the irreversible wide-range inhibitor benzyloxycarbonyl-Val-Ala-DL-Asp-fluoromethylketone. In summary, the data presented here provide further insight into the mechanisms of injury-induced neuronal apoptosis and could give rise to more effective therapeutic intervention strategies in CNS trauma and neurodegenerative diseases.

Background and Purpose-Diagnosis of paroxysmal atrial fibrillation is difficult but highly relevant in patients presenting withcerebral ischemia yet free from atrial fibrillation on admission. Early initiation and prolongation of continuous Holter monitoring may improve diagnostic yield compared with the standard of care including a 24-hour Holter recording. Methods-In the observational Find-AF trial (ISRCTN 46104198), consecutive patients presenting with symptoms of cerebral ischemia were included. Patients free from atrial fibrillation at presentation received 7-day Holter monitoring. Results-Two hundred eighty-one patients were prospectively included. Forty-four (15.7%) had atrial fibrillation documented by routine electrocardiogram on admission. All remaining patients received Holter monitors at a median of 5.5 hours after presentation. In those 224 patients who received Holter monitors but had no previously known paroxysmal atrial fibrillation, the detection rate with early and prolonged (7 days) Holter monitoring (12.5%) was significantly higher than for any 24-hour (mean of 7 intervals: 4.8%, Pϭ0.015) or any 48-hour monitoring interval (mean of 6 intervals: 6.4%, Pϭ0.023). Of those 28 patients with new atrial fibrillation on Holter monitoring, 15 (6.7%) had been discharged without therapeutic anticoagulation after routine clinical care (ie, with data from 24-hour Holter monitoring only). Detection rates were 43.8% or 6.3% for short supraventricular runs of Ն10 beats or prolonged episodes (Ͼ5 hours) of atrial fibrillation, respectively. Diagnostic yield appeared to be only slightly and not significantly increased during the first 3 days after the index event. Conclusions-Prolongation of Holter monitoring in patients with symptoms of cerebral ischemic events increases the rate of detection of paroxysmal atrial fibrillation up to Day 7, leading to a relevant change in therapy in a substantial number of patients. Early initiation of monitoring does not appear to be crucial. Hence, prolonged Holter monitoring (Ն7 days) should be considered for all patients with unexplained cerebral ischemia. (Stroke. 2010;41:2884-2888.)

Recently we have shown that the majority of retinal ganglion cells (RGCs) dies via activation of caspase-3 after transection of the optic nerve (ON) in the adult rat. In the present study we investigated whether insulin-like growth factor-I (IGF-I), an important factor in retinal development, prevents secondary death of RGCs after axotomy. Moreover, we studied potential intracellular mechanisms of IGF-mediated neuroprotection in more detail. Our results indicate that intraocular application of IGF-I protects RGCs from death after ON transection in a dosedependent manner. We show reduced caspase-3 activity as one possible neuroprotective mechanism of IGF-I treatment in vivo. Caspase-3 mRNA expression remained unchanged. Because caspase inhibition can be mediated by Akt in vitro, we examined phosphorylation of Akt after axotomy and under IGF treatment. Western blot analysis revealed decreased Akt phos-phorylation after axotomy without treatment and an increased phosphorylation of Akt under treatment with IGF-I. This strong increase could be reduced by simultaneous injection of wortmannin (WM), a potent inhibitor of phosphatidylinositol 3-kinase (PI3-K). To prove the pathway suggested by these experiments as relevant for the in vivo situation, we assessed the number of RGCs 14 d after ON transection under a combined treatment strategy of IGF-I and WM. As expected, WM significantly reduced the neuroprotective effects of IGF-I. In summary, we show for the first time in vivo that IGF is neuroprotective via PI3-K-dependent Akt phosphorylation and by inhibition of caspase-3.

Recently, we have shown that inhibition of caspase-3-like caspases is the most effective treatment strategy to protect adult rat retinal ganglion cells from secondary death following optic nerve transection. In the present study, we localized active caspase-3 in axotomized retinal ganglion cells in vivo and demonstrated a co-localization of the active p20 fragment and TUNEL-staining in some of these cells. In line with this, we detected an enhanced cleavage and activity of caspase-3 protein in retinal tissue after lesion, while caspase-3 mRNA expression remained unchanged. These data suggest caspase-3 as an important mediator of secondary retinal ganglion cell death following axotomy in vivo.z 1999 Federation of European Biochemical Societies.

The neurotrophin brain-derived neurotrophic factor (BDNF) serves as a survival, mitogenic, and differentiation factor in both the developing and adult CNS and PNS. In an attempt to identify the molecular mechanisms underlying BDNF neuroprotection, we studied activation of two potentially neuroprotective signal transduction pathways by BDNF in a CNS trauma model. Transection of the optic nerve (ON) in the adult rat induces secondary death of retinal ganglion cells (RGCs). Repeated intraocular injections of BDNF prevent the degeneration of RGCs 14 d after ON lesion most likely by inhibition of apoptosis. Here, we report that BDNF activates both protein kinase B (PKB) via a phosphatidyl-inositol-3Ј-kinase (PI-3-K)-dependent mechanism and the mitogen-activated protein kinases extracellular signalregulated kinase 1 (ERK1) and ERK2. Furthermore, we provide evidence that BDNF suppresses cleavage and enzymatic activity of the neuronal cell death effector caspase-3. Distinct from our recent study in which inhibition of the PI-3-K/PKB pathway attenuated the survival-promoting action of insulin-like growth factor-I on axotomized RGCs (Kermer et al., 2000), it does not in the case of BDNF. Thus, we assume that BDNF does not depend on a single signal transduction pathway exerting its neuroprotective effects on lesioned CNS neurons.

Only higher CASPR2 serum antibody titres indicate anti-CASPR2 encephalitis, and diagnostic accuracy increases if MRI findings are considered. Anti-CASPR2 encephalitis has characteristic features and a favourable outcome with immunotherapy.