Abstracts

Rationale: The process by which imbalances arise during ictogenesis in the interactions within and between excitatory and inhibitory neuronal populations remains unknown. The zebrafish model system, where neuronal dynamics over a population can be measured and manipulated at the single cell level, provides an attractive setting where the processes underlying ictogenesis can be explored and understood. Here we combined local field potential recordings with in vivo two-photon calcium imaging to monitor activity over the entire brain with high spatial resolution in awake larval zebrafish after induction of seizure-like states. Methods:  Zebrafish larvae (Tg(HuC:h2b-GCaMP6f) x Tg(Vglut2a:dsRed)) were immobilized in agarose and placed in embryo medium. Glass electrodes (1-3 Mohm impedance) were inserted through the skin to record epileptiform activity in the brain. A custom built two photon microscope was used for imaging calcium activity (2-4Hz). Epileptiform activity was induced with bath application of the convulsant pentylenetetrazol (PTZ, 15 mM). Results: Dual electrode recordings of forebrain, midbrain, and hindbrain regional pairs indicated that the set of epileptiform events detected with midbrain recordings encompassed those observed at forebrain and hindbrain locations. Epileptiform events were clustered into two groups: putative ictal events that lasted 2-10 seconds, and putative interictal events that lasted less than 2 seconds. The ictal events showed higher power around 50 Hz than interictal events. Volumetric imaging at 4-6 planes revealed that the largest variation in calcium dynamics occurred along the rostro-caudal axis. Repeated imaging at a depth where a large fraction of forebrain, midbrain, and hindbrain neuronal population were visible revealed that many ictal events initiated in the optic tectum at caudal locations near the midbrain/hindbrain border. Conclusions: Ongoing work is geared towards further isolating the site(s) of initiation and determining the contributions of excitatory and inhibitory populations. This will set the stage for us to develop a detailed map of neuronal dynamics in the brain capturing the transition from interictal to ictal states and develop targeted optophysiological therapies. Funding: -