Optical method for neuronal activity mapping during seizures in zebrafish

Conventionally, epileptic seizures are detected and characterized in zebrafish either as behavioral alterations [1, 2, 3] or by means of direct electrographic recordings [1, 2, 4]. To overcome the inherent limitations of these conventional approaches (namely limited sensitivity of behavioural assays and lack of spatial resolution of electrographic recordings), we implemented the real time optical mapping of neuronal activity during the onset and propagation of epileptic seizures. We employed a transgenic zebrafish line with pan-neuronal expression of the genetically encoded calcium indicator GCaMP6s5 to measure neuronal activity in zebrafish larvae during seizures induced by pentylenetetrazole (PTZ). With this approach, we simultaneously measured the neuronal activity in different regions of the larva brain, showing the high sensitivity of this method to detect different levels of alteration, as induced by increasing PTZ concentrations. We also performed simultaneous recording of brain and locomotor activity, demonstrating that GCaMP measurements can be much more sensitive than behavioural assays for the detection of subclinical epileptic seizures, thus enabling future investigations on hypomorphic human muta- tions. Moreover, taking advantage of the strong fluorescence signal of GCaMP reporter, we developed a novel and comprehensive high-throughput drug screening method able to measure neuronal activity and behavioural parameters on 60 individual larvae, at the same time. The methodological approach described here could provide a deeper comprehension of the mecha- nisms underlying epilepsy and improve the development of new anticonvulsant drugs. Further- more, this method can also be easily applied to the study of many human neuropathologies modelled in zebrafish, enabling a simple but detailed investigation of brain activity alterations associated with the pathological phenotype. References 1. Barabanetal.Pentylenetetrazoleinducedchangesinzebrafishbehavior,neuralactivityandc-FoSexpression.2005 Neuroscience 131, 759–768. 2. Afrikanova et al. Validation of the zebrafish pentylenetetrazol seizure model: locomotor versus electrographic responses to antiepileptic drugs. 2013 PloS one 8, e54166. 3. Winteretal.Validationofalarvalzebrafishlocomotorassayforassessingtheseizureliabilityofearly-stagedevelop- ment drugs. 2008 J Pharmacol Toxicol Methods 57, 176–187. 4. Zdebik et al. Epilepsy in kcnj10 morphant zebrafish assessed with a novel method for long-term EEG recordings. 2013 PloS one 8, e79765. 5. Vladimirov et al. Light-sheet functional imaging in fictively behaving zebrafish. 2014 N Meth 11, 883–884.