Cellular contributions to ictal population signals

Abstract

ObjectiveThe amplitude of ictal activity is a defining feature of epileptic seizures, but the determinants of this amplitude have not been identified. Clinically, ictal amplitudes are measured electrographically (using e.g. EEG, ECoG, and depth electrodes), but these methods do not enable the assessment of the activity of individual neurons. To identify the cellular determinants of the ictal signal, we measured single cell and population electrical activity and neuronal calcium levels via optical imaging of the genetically encoded calcium indicator (GECI) GCaMP. MethodsSpontaneous seizure activity was assessed in an awake, behaving mouse model of focal cortical injury and in organotypic hippocampal slice cultures (OHSC), an in vitro preparation from which recurrent seizures can be readily captured. Single cell calcium signals were linked to a range of electrical activities by performing simultaneous GECI-based calcium imaging and whole-cell patch-clamp recordings in spontaneously seizing OHSCs. Neuronal resolution calcium imaging was then performed during spontaneous seizures in vivo and in vitro to quantify the cellular contributions to the population-level calcium signal. ResultsPopulation signal may increase from three potential sources: 1) increased synchrony, i.e. more co-active neurons, 2) altered active state, from bursts of action potentials and/or paroxysmal depolarizing shifts in membrane potential, and 3) altered subthreshold state, which includes all lower levels of activity. The largest contributor to the signal recorded at seizure onset was increased subthreshold activity, consistent with either barrages of excitatory postsynaptic potentials or sustained membrane depolarization. The relative contribution of synchrony increased as seizures progressed, but cell intrinsic alterations in both the subthreshold and active states remained the largest driver of the ictal signal. SignificanceWe introduce here a novel method for the quantification of the relative contributions of inter-versus intra-cellular changes to provide a critical link between single neuron activity and population measures of seizure activity. Key PointsO_LINeuronal calcium as measured by GCaMP reports a range of membrane depolarizations, from EPSPs to action potential firing and paroxysmal depolarizing shifts C_LIO_LIThe mean population calcium signal is highly correlated with the electrographic local field potential C_LIO_LIIncreased calcium signal during seizure onset is largely driven by increased subthreshold calcium within individual neurons C_LIO_LIRecruitment of newly active neurons is a minor contributor to the increasing population-level signal during the transition to frank seizure C_LI