Investigation of glutamate accumulation and excitotoxic mechanisms during the acute phase of metabolic stress

During metabolic stress, such as ischemic stroke, insufficient energy supply leads to ATP deprivation, resulting in ion imbalances and depolarization of cells. This results in uncontrolled glutamate release and impaired uptake by astrocytes. However, the role of elevated extracellular glutamate concentrations in exacerbating metabolic stress remains unclear. To investigate glutamate dynamics in the extracellular space during the acute phase of energy deprivation, we utilized the green-fluorescent glutamate sensor SF-iGluSnFR1. Organotypic mouse brain slice cultures were transduced with synapsin-driven SF-iGluSnFR (A184V), and brief chemical ischemia was induced by depleting glucose and applying sodium azide and 2-deoxy-D-glucose. Wide-field imaging of organotypic brain slices showed synchronous network activity but also revealed spontaneous, local glutamate release events with plume-like properties. Plumes were relatively large in size, heterogenous and comparably long-lasting. They occurred independently of network activity but inhibition of glutamate transporters with TBOA caused a strong increase in plume frequency. Importantly, plumes were the main driver of glutamate accumulation during chemical ischemia, during which they increased in size, duration, and frequency. Current experiments aim at examining the role of subsequent excitotoxic effects that occur in the acute phase of metabolic stress. In addition to iGluRs, we will investigate the contributions of the calcium-binding transient receptor potential melastatin 4 (TRPM4) channel, which has been shown to be activated during ischemic events2.1Marvin et al., Nat. Methods (2018) 15: 135-1432Leiva-Salcedo et al., Channels (2017) 6: 624-635