Ischemia-elicited Oxidative Modulation of Ca2+/Calmodulin-dependent Protein Kinase II

Pavan K Shetty,Freesia L Huang,Kuo-Ping Huang

doi:10.1074/jbc.m708479200

Abstract

Ca2+/calmodulin (CaM)-dependent protein kinase II (CaMKII) plays a critical role in neuronal signal transduction and synaptic plasticity. Here, we showed that this kinase was very susceptible to oxidative modulation. Treatment of mouse brain synaptosomes with H2O2, diamide, and sodium nitroprusside caused aggregation of CaMKII through formation of disulfide and non-disulfide linkages, and partial inhibition of the kinase activity. These CaMKII aggregates were found to associate with the post synaptic density. However, treatment of purified CaMKII with these oxidants did not replicate those effects observed in the synaptosomes. Using two previously identified potential mediators of oxidants in the brain, glutathione disulfide S-monoxide (GS-DSMO) and glutathione disulfide S-dioxide (GS-DSDO), we showed that they oxidized and inhibited CaMKII in a manner partly related to those of the oxidant-treated synaptosomes as well as the ischemia-elicited oxidative stress in the acutely prepared hippocampal slices. Interestingly, the autophosphorylated and activated CaMKII was relatively refractory to GS-DSMO- and GS-DSDO-mediated aggregation. Short term ischemia (10 min) caused a depression of basal synaptic response of the hippocampal slices, and re-oxygenation (after 10 min) reversed the depression. However, oxidation of CaMKII remained at above the pre-ischemic level throughout the treatment. Oxidation of CaMKII also prevented full recovery of CaMKII autophosphorylation after re-oxygenation. Subsequently, the high frequency stimulation-mediated synaptic potentiation in the hippocampal CA1 region was significantly reduced compared with the control without ischemia. Thus, ischemia-evoked oxidation of CaMKII, probably via the action of glutathione disulfide S-oxides or their analogues, may be involved in the suppression of synaptic plasticity.

Highlights

Ca2ϩ/calmodulin (CaM)2-dependent protein kinase II (CaMKII) is one of the major Ca2ϩ-sensing enzymes important in transducing neuronal, hormonal, and electrical signals in brain, heart, and other tissues
Using two previously identified potential mediators of oxidants in the brain, glutathione disulfide S-monoxide (GS-DSMO) and glutathione disulfide S-dioxide (GS-DSDO), we showed that they oxidized and inhibited CaMKII in a manner partly related to those of the oxidanttreated synaptosomes as well as the ischemia-elicited oxidative stress in the acutely prepared hippocampal slices
CaMKII holoenzyme is a dodecamer composed of two stacked hexameric rings, in which each catalytic/regulatory domain from the upper ring interacts with the equivalent catalytic/regulatory domain in the lower ring by an antiparallel coiled-coil, which resides in regulatory domains [2]

Summary

Introduction

Ca2ϩ/calmodulin (CaM)2-dependent protein kinase II (CaMKII) is one of the major Ca2ϩ-sensing enzymes important in transducing neuronal, hormonal, and electrical signals in brain, heart, and other tissues. Translocation and clustering of CaMKII have been implicated in the NMDA receptor-dependent enhancement of synaptic plasticity (6 –9) as well as in neurological disorders associated with ischemic injury (10 –12) and seizure [13]. Stimulation of NMDA receptors by excitatory amino acids increases postsynaptic Ca2ϩ influx, which facilitates the activation and autophosphorylation of CaMKII and its interaction with the NR2B subunit of NMDA receptors and other proteins associated with the postsynaptic density (16 –18). Stimulation of NMDA receptors causes an increase in intracellular Ca2ϩ and a slight acidification of cellular cytoplasm [21, 22], a condition conducive for self-association, it increases production of reactive oxygen species, such as nitric oxide [23] and superoxide [24]. The ischemia-evoked oxidation of CaMKII was not readily reversed by re-oxygenation, and this lingering oxidative modification suppressed the autophosphorylation of the kinase and synaptic potentiation post ischemia

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