Abstracts

Rationale: Nearly one-third of epilepsy patients suffer ongoing seizures despite multiple medication trials. Most studies describe patterns of increased activity and neuronal degeneration at seizure sites, but lack a description of the underlying circuit mechanism. In humans, the hippocampus (HC) and the entorhinal cortex (EC) are central to the pathophysiology of temporal lobe epilepsy, the most common drug-resistant epilepsy. However, most of what we know about HC and EC is from rodent studies. Our study fills this gap by characterizing the structure and function of human cortico-hippocampal circuit. We test whether the increased excitability in this circuit results from an increase in intrinsic excitability of neurons or from changes in circuit components and connections.

Methods: We use freshly resected human brain tissue from patients undergoing surgery for refractory epilepsy. Further, as shifts in the excitation-inhibition balance are considered key to the underlying pathophysiology of seizure generation, using electrophysiology and histology we characterize excitatory vs inhibitory neuron populations in the epileptogenic human brain, compared to non-epileptic human tissue and control mice. To obtain high resolution morphology and cell-type characterization of human neurons we use fluorescence immunohistochemistry combined with confocal and light sheet microscopy. We also corroborate our single neuron and circuit level physiology with patient-specific pathology of seizures vis a vis developmental timeline, focal area, spread and severity.

Results: At the level of individual neurons, in human epileptic brain slices we observed a selective increase in intrinsic neuronal excitability restricted to CA3 and not observed in CA1 or EC. Additionally, in mouse, CA3 is the least excitable as compared to CA1 and EC. However, in human epileptic tissue, we found CA3 to be the most excitable when compared to the other two regions. At the circuit level, we observed that while neurons in superficial layers of EC demonstrate a high frequency of spontaneous subthreshold events, deeper layers do not show much baseline activity. Moreover, a comparison of evoked electrical responses revealed that the EC-evoked responses recorded in human epileptic CA1 neurons are much larger than those recorded in the control mouse CA1. Whereas, the CA3 evoked responses in CA1 were comparable between the two species. Our preliminary data indicate an increase in the excitability and degradation of individual pyramidal neurons in HC, and disparity in the activity and strength of different layers and signal pathways in EC.

Conclusions: Acute slice electrophysiology from resected human tissue not only allows us to examine synaptic and circuit pathophysiology at various stages of epilepsy in humans and correlate it with clinical symptoms but also draw cross species comparisons in relatively intact circuits. Our study depicts the fundamental principles of synaptic transmission, input-output transformation and short-term plasticity dynamics in cortico-hippocampal circuit in the human brain and how these key processes may predispose a brain area to be more excitable and prone to seizures.

Funding: FACES 2022, CTSI 2021