Ectopic HCN4 expression drives mTOR-dependent epilepsy in mice

Abstract

The causative link between focal cortical malformations (FCMs) and epilepsy is well accepted, especially among patients with focal cortical dysplasia type II (FCDII) and tuberous sclerosis complex (TSC). However, the mechanisms underlying seizures remain unclear. Using a mouse model of TSC- and FCDII-associated FCM, we showed that FCM neurons were responsible for seizure activity via their unexpected abnormal expression of the hyperpolarization-activated cyclic nucleotide–gated potassium channel isoform 4 (HCN4), which is normally not present in cortical pyramidal neurons after birth. Increasing intracellular cAMP concentrations, which preferentially affects HCN4 gating relative to the other isoforms, drove repetitive firing of FCM neurons but not control pyramidal neurons. Ectopic HCN4 expression was dependent on the mechanistic target of rapamycin (mTOR), preceded the onset of seizures, and was also found in diseased neurons in tissue resected from patients with TSC and FCDII. Last, blocking HCN4 channel activity in FCM neurons prevented epilepsy in the mouse model. These findings suggest that HCN4 play a main role in seizure and identify a cAMP-dependent seizure mechanism in TSC and FCDII. Furthermore, the unique expression of HCN4 exclusively in FCM neurons suggests that gene therapy targeting HCN4 might be effective in reducing seizures in FCDII or TSC.

Convergent and Divergent Mechanisms of Epileptogenesis in mTORopathies

“…Neuronal cytomegaly was also consistently reported for the evaluated gene variants. Increased neuron soma size was reported as early as E17.5 (Tarkowski et al, 2019) and persisted into adulthood (Lim et al, 2015(Lim et al, , 2017Park et al, 2018;Ribierre et al, 2018;Nguyen et al, 2019;Zhang et al, 2019;Dawson et al, 2020;Hsieh et al, 2020;Zhong et al, 2021). Furthermore, increasing mTORC1 activity levels by varying concentration of the Rheb S16H plasmid resulted in dose-dependent increases in neuron soma size, supporting a direct relationship between mTORC1 activity level and soma size (Nguyen et al, 2019).…”

“…These studies reported that dysmorphic, cytomegalic neurons within the tissue samples exhibit altered intrinsic excitability that may contribute to the generation of epileptic activity (Cepeda et al, 2003(Cepeda et al, , 2005(Cepeda et al, , 2012Wang et al, 2007). In addition, a recent IUE study in Rheb S16H mutant pyramidal neurons reported that decreasing neuronal excitability by expressing an inwardly-rectifying potassium channel (Kir2.1) significantly reduced seizures, supporting that mutant neurons are directly involved in seizure generation (Hsieh et al, 2020). In light of these findings supporting a cell autonomous mechanism of hyperexcitability, we review the electrophysiological data on neurons expressing the different PI3K-mTOR and GATOR1 gene variants.…”

“…Interestingly, no changes in R in or I/O curve were observed for neurons expressing the Pi3k E545K variant (Goz et al, 2020). Regarding the resting membrane potential (RMP), Rheb S16H mutant neurons in the mPFC displayed a depolarized RMP (Lin et al, 2016;Hsieh et al, 2020). In contrast, Pi3k E545K, Pten, Rheb P37L, and Depdc5 mutant neurons in the SSC displayed no changes in RMP (Chen et al, 2015;Hu et al, 2018;Goz et al, 2020;Onori et al, 2020).…”

“…In particular, increased capacitance was found in mutant neurons expressing Rheb P37L (Onori et al, 2020) and Depdc5 ( Ribierre et al, 2018) variants, which is consistent with increased cell size. Additionally, Pten, Tsc1, Rheb S16H, Rheb P37L, and Depdc5 mutant neurons all displayed decreased input resistance (R in ) (Chen et al, 2015;Hu et al, 2018;Ribierre et al, 2018;Goz et al, 2020;Hsieh et al, 2020;Onori et al, 2020). As a result of decreased R in , mutant neurons required a larger current injection to reach AP threshold [i.e., decreased input/output (I/O) or increased rheobase].…”

Mitochondria and metabolic transitions in cardiomyocytes: lessons from development for stem cell-derived cardiomyocytes

“…AMPK activation downregulates Kir2.1 [291], and we recently found that sustained activation can enhance maturation of contractile and metabolic features of PSC-CMs but also reduces expression of KCNJ2 [292]. In contrast, in neurons, mTOR signaling increases expression of HCN channels [293]. Dysregulation of AMPK and mTOR signaling may affect expression levels of Kir2.1 and HCN4 in cardiomyocytes, leading to cardiomyocyte membrane depolarization.…”