To study the influence of voltage-gated sodium channels on calcium oscillations, 100 nM tetrodotoxin was added acutely to cultures after 200 frames. To study glutamatergic and GABAergic contributions to calcium signals and neuronal activity, a combination of 20 μM 6-Cyano-7-nitroquinoxaline-2,3-dione (CNQX, competitive AMPA/kainate receptor antagonist, Sigma) and 50 μM D-(-)-2-Amino-5-phosphonopentanoic acid (D-AP5, competitive NMDA receptor antagonist, Abcam), or 50 μM picrotoxin (GABAA receptor inhibitor, Tocris Bioscience) respectively was added acutely to the cultures after 200 frames. Calcium analysis of these recordings was done by splitting the analysis in two time stretches (before and after the pharmacological intervention), after which calcium parameters from both stretches were compared.
