Abstracts

Rationale: Sudden unexplained death in epilepsy (SUDEP) is a major concern for caregivers of patients with Dravet syndrome (DS). Fenfluramine has been shown to substantially reduce expected rates of SUDEP in DS patients (Cross JH et al, AES 2020), and a recent study found that fenfluramine reduces audiogenic seizure-induced respiratory arrest in the DBA/1 SUDEP mouse model (Tupal and Faingold, Epilepsia. 2019). Fenfluramine targets multiple molecular targets including serotonergic and sigma-1 receptor pathways (Martin et al, Int J Mol Sci. 2021); however definitive anti-SUDEP targets remain to be elucidated. Spreading depolarization (SD) is a leading pathophysiologic event linked to cardiorespiratory collapse in several mouse models of SUDEP; fenfluramine inhibits SD in rodent cortex in vivo (Cabral-Filho et al, Braz J Med Biol Res. 1995). Here we report that fenfluramine directly inhibits SD in ex vivo brain slices, and we explore its potential inhibitory mechanisms.

Methods: We examined the effects of fenfluramine, serotonin, and sigma-1 receptor agonists (dextromethorphan, carbetapentane) on SD evoked by KCl or oxygen glucose deprivation (OGD) in cortical brain slices of adult C57Bl/6 mice. We also measured the effects of these compounds on GABA_AR-mediated inhibitory postsynaptic currents (sIPSC) in cortical pyramidal neurons in vitro.

Results: Fenfluramine and sigma-1 receptor agonists increased the threshold for SD events at clinically relevant concentration ranges (~10 µM). In contrast, bath application of serotonin (10 and 100 µM) alone had little effect on cortical SD threshold in vitro, and coapplication of serotonin with fenfluramine did not show any additive effects. Despite its lack of effect on SD threshold, serotonin at 10 µM increased sIPSC frequency in pyramidal neurons, while fenfluramine showed a lesser effect on sIPSC frequency. Blocking GABA_ARs with gabazine did not fully occlude SD inhibition by fenfluramine, suggesting that GABA_AR potentiation is not involved in the inhibitory SD mechanism of fenfluramine.

Conclusions: Our ex vivo results demonstrate that in the clinically relevant concentration range, fenfluramine directly inhibits SD generation without acting via a neurovascular mechanism. Sigma-1 receptor agonists had a comparable effect. These results suggest SD inhibition could be a plausible mechanism responsible for the decrease in expected rates of SUDEP observed in patients treated with fenfluramine. Unlike previous reports in the DBA/1 mouse model, the effect of fenfluramine on SD does not appear to be driven solely by its serotonergic actions. Notably, fenfluramine exerts a positive allosteric effect at the sigma-1 receptor, and sigma-1 receptor agonists increase the threshold for evoked SD. Further experiments are warranted to better understand the molecular mechanisms by which fenfluramine inhibits SD and prevents SUDEP.

Funding: Please list any funding that was received in support of this abstract.: Sponsor: Zogenix, Inc.