Proceedings of The Physiological Society

University College Dublin (2009) Proc Physiol Soc 15, PC204

Poster Communications

The role of SUMOylation in Kainate Receptor trafficking during Oxygen Glucose Deprivation.

S. Dennis1, J. Mellor1

1. University of Bristol, Bristol, United Kingdom.


Ischaemic brain damage is thought to be largely caused by excitotoxicity due to over-activation of glutamate receptors. The roles of NMDA and AMPA receptors in mediating this excitotoxic damage are well documented but the role of kainate receptors (KARs) is less known. A number of studies have now begun to reveal a role for KARs in mediating ischaemic damage (1). Small Ubiquitin like Modifier (SUMO) is greatly upregulated during ischaemia and has been suggested that this may be a neuroprotective mechanism (2,3). We discovered that the KAR subunit GluR6a is SUMOylated by SUMO1 and that this controls KAR trafficking causing removal of KARs from the postsynaptic membrane (4). We hypothesised that neurones may be protected from ischaemic damage by increasing SUMOylation of KARs causing their removal from the cell surface and thereby reducing glutamate-mediated excitotoxicity. We exposed hippocampal slices taken from P13-15 rats to 15 minutes of oxygen glucose deprivation (OGD, oxygen and glucose replaced with nitrogen and sucrose respectively), an in vitro model of ischaemia, and measured synaptic KAR-EPSCs using whole-cell electrophysiological recordings. We manipulated the degree of SUMOylation within the cell by including SUMO1, SUMO2 or the deSUMOylating enzyme SENP1 in the patch pipette. We replicated previous results demonstrating that inclusion of SUMO1 (4.2µM) reduced KAR-EPSCs and that SENP1 (100nM) increased KAR-EPSCs (4). Inclusion of SUMO2 had no effect on the amplitude of KAR-EPSCs. This suggests that SUMO1 and SUMO2 play different roles in the regulation of KAR trafficking. OGD decreased the amplitude of KAR-EPSCs (measured 20-25 minutes after OGD) by 60.5±10% (n=8). The reduction in KAR-EPSC amplitude suggests internalisation of KARs from the membrane since it was not due to changes in pre-synaptic fibre volley. SUMO1, SUMO2 and SENP1 and corresponding inactive control proteins were infused and KAR-EPSC amplitude stabilised before OGD application. OGD caused a 69.4±10% (n=12) decrease in KAR-EPSC after SUMO1 infusion that was significantly greater than the decrease induced in interleaved controls using infusion of the inactive version of SUMO1, SUMO1ΔGG (47.1±10%, n=10, p=0.047, unpaired t-test). In contrast, OGD after infusion of SUMO2 caused significantly less reduction in the KAR-EPSC compared to SUMO2ΔGG (40.9±10% and 63.4±10% respectively, n=7, p=0.048, unpaired t-test). Infusion of SENP1 had no effect on the recovery of the KAR-EPSC after OGD compared to the inactive version of SENP1, SENP1-C603S (p=0.049, unpaired t-test). The results presented suggest that SUMOylation can alter the trafficking of KARs after OGD. However, SUMOylation does not appear to play a major role in the loss of KARs during OGD; as the KAR-EPSC never fully recovered after OGD. All errors given are standard errors of the mean.

Where applicable, experiments conform with Society ethical requirements