Abstracts

Rationale: Thrombospondins (TSPs) are astrocyte-secreted extracellular matrix proteins that play key roles as regulators of synaptogenesis in the central nervous system. We previously showed that TSP1/2 are upregulated in the partial neocortical isolation model ( “undercut” or “UC” below) of posttraumatic epileptogenesis and may contribute to abnormal axonal sprouting, aberrant synaptogenesis and epileptiform discharges in the UC cortex. These results led to the current experiments to test the hypothesis that epileptiform activity would be reduced in TSP1/2 knockout (TSP^1/2KO) mice. Methods: Four groups of mice of either sex were studied: 1) FVB Wild type (WT) naïve; 2) WT + UC; 3) TSP^1/2KO naïve; 4) TSP^1/2KO + UC. We made UC lesions in at P21 and subsequent experiments were done ~14d later at P35. In vitro extracellular single or multi-electrode field potential recordings were obtained from layer V in cortical slices at P35 and in vivo video-EEGs of spontaneous epileptiform bursts were recorded to examine the effect of TSP1/2 deletion on epileptogenesis. Immunocytochemistry was used to assess the effect of TSP^1/2KO+UC on the number of putative excitatory synapses and the expression of TSP4 and Hevin, other astrocytic proteins known to up-regulate excitatory synapse formation. Results: Compared with WT+UC mice, TSP^1/2KO+UC mice showed significant increases in epileptogenesis including 1) increased incidence and more rapid propagation of evoked and spontaneous epileptiform discharges in UC neocortical slices (~5 vs. 20 m/s; n= 4-5 mice/each group, p1/2KO+UC versus WT+UC mice;  3) increased occurrence of spontaneous epileptiform discharges in vivo. There was an associated increase in density of VLUT1/PSD95-IR colocalizations (putative excitatory synapses) and significantly increased TSP4- and Hevin-IR in TSP^1/2KO+UC versus WT+UC mice.                                                                                        Conclusions: Results show that TSP1/2 deletion has unexpected epileptogenic effects in a model of neocortical injury, associated with compensatory upregulation of other synaptogenic astrocytic proteins, TSP4 and Hevin, that may contribute to the increase in density of excitatory synapses and resulting neural network hyperexcitability. The increased speed in propagation of epileptiform bursts are likely due to reductions in GABAergic inhibition, such as are known to occur in the UC and other models of epileptogenesis. Funding: Supported by NIH grant R01NS090076 (DAP) and the Pimley Research Fund.