Abstracts

Rationale: Epilepsy can develop following different precipitating events sharing blood brain barrier dysfunction as the common denominator, including traumatic brain injury. We previously demonstrated that the uptake of albumin into astrocytes is critical in the generation of perilesional hyperxcitability and seizures following blood-brain barrier compromise, and identified TGFβ signaling pathway activation as a critical step (Cacheaux et al, 2009, J Neurosci, 29: 8927-35). However, the exact mechanims by which the astrocytic reaction to serum albumin leads to sustained hyperexcitability is largely unknown. Recent studies have shown that astrocyte-secreted factors, including thrombospondins (TSP), can control the strength and number of both excitatory and inhibitory synapses (Christopherson et al, 2005, Cell, 120: 421-433; Hughes et al, 2010, Mol Cell Neurosci, 43: 136-145). Therefore, we investigated altered excitatory synaptogenesis in primary hippocampal cultures following albumin treatment, and its dependence on TGFβ signaling and astrocyte-secreted factors. Methods: The effects of albumin exposure on synaptic connectivity were assessed using primary cultured hippocampal CNS cells. Synaptogenesis was assessed after albumin exposure using immunocytochemical labeling of pre and postsynaptic markers, Synapsin 1 and PSD-95, respectively. Gene expression levels were determined of using qRT-PCR. Results: We exposed cultures of mixed neurons/glia hippocampal cells to albumin (0.2 mM), or TGFβ1 (10 ng/mL) and followed the development of excitatory synapses. The number of co-localized puncta of pre- and post-synaptic protein markers was increased 72 hours post treatment with either albumin (169% of control, p = 0.042) or TGFβ1 (189% of control, p = 0.006). Interestingly, there was no change in the number of synases following the treatments in pure neuronal cultures, suggesting that the albumin/TGFβ-induced increase in excitatory synapse number requires an astrocytic process. Furthermore, this albumin/ TGFβ-induced increase was blocked in the presence of the TGFβ blocker SJN. Conclusions: Our data suggest that the pathological exposure of brain cells to the serum protein albumin induces an increase in the number of excitatory synapses. This increase is dependent on the activation of TGFβ signaling and the presence of astrocytes. Overall, this aberrant synaptogenesis can lead to an excitatory/inhibitory imbalance and potentially link the proximal molecular events following the blood brain barrier failure to the delayed development of posttraumatic epilepsy.