Watch Out for GATORs: Fasting, Seizures, and Nutrient Sensing

Abstract

DEPDC5-Dependent mTORC1 Signaling Mechanisms Are Critical for the Anti-Seizure Effects of Acute Fasting

Yuskaitis CJ, Modasia JB, Schrötter S, Rossitto LA, Groff KJ, Morici C, Mithal DS, Chakrabarty RP, Chandel NS, Manning BD, Sahin M. Cell Rep. 2022;40(9):111278. doi:10.1016/j.celrep.2022.111278

Caloric restriction and acute fasting are known to reduce seizures but through unclear mechanisms. mTOR signaling has been suggested as a potential mechanism for seizure protection from fasting. We demonstrate that brain mTORC1 signaling is reduced after acute fasting of mice and that neuronal mTORC1 integrates GATOR1 complex-mediated amino acid and tuberous sclerosis complex (TSC)-mediated growth factor signaling. Neuronal mTORC1 is most sensitive to withdrawal of leucine, arginine, and glutamine, which are dependent on DEPDC5, a component of the GATOR1 complex. Metabolomic analysis reveals that Depdc5 neuronal-specific knockout mice are resistant to sensing significant fluctuations in brain amino acid levels after fasting. Depdc5 neuronal-specific knockout mice are resistant to the protective effects of fasting on seizures or seizure-induced death. These results establish that acute fasting reduces seizure susceptibility in a DEPDC5-dependent manner. Modulation of nutrients upstream of GATOR1 and mTORC1 could offer a rational therapeutic strategy for epilepsy treatment.

Commentary

The benefits of fasting on epilepsy provide a rationale for dietary therapies, including the ketogenic diet. Although a metabolic switch between glucose-derived to free fatty acid-derived energy may underlie the potential for reduced hyperexcitability in the fasted state, the biochemical processes producing these changes are not fully understood. The regulatory protein mammalian (or mechanistic) target of rapamycin (mTOR) plays important roles in several neurochemical pathways important for seizure control. In the non-fasted state, mTOR promotes protein and lipid synthesis. In contrast, fasting is associated with decreased mTOR activity, inhibition of protein synthesis, and reparative processes that may reduce oxidative stress. ¹ mTOR inhibition is an important target for seizure reduction, as demonstrated by the approval of the mTOR antagonist everolimus to treat patients with tuberous sclerosis complex (TSC), a disease caused by TSC1/TSC2 mutations that increase mTOR activity. Therefore, mTOR is a key protein in neurophysiologic processes involved in fasting and seizure control.

mTOR complex 1 (mTORC1) is an important nutrient sensor and functions collectively with other proteins (e.g., Rag and Rheb GTPases). Over activation of mTORC1 produces seizures ² and can be partially reduced by mTOR inhibition. ³ Under caloric restriction mTORC1 activity is inhibited, but importantly the reasons for this are unknown. As demonstrated by Yuskaitis and colleagues, one component of the GATOR1 protein complex, DEP domain-containing protein 5 (DEPDC5), may be a key link between fasting and seizure reduction for mTORC1. ⁴ DEPDC5 deficiency is associated with seizures in humans and in animal models, and seizure reduction can be achieved with mTOR inhibitors. ^3,5 In a series of experiments, the interplay between DEPDC5 and mTOR in responding to nutrient signals is elucidated, revealing a potential mechanism whereby fasting reduces seizures.

The authors begin by confirming the anti-seizure effect of fasting using the pentylenetetrazole model, demonstrating that acute fasting reduces seizures in mice by approximately half. Under these conditions, mTOR activity is also decreased. Using a conditional knockout mouse model for DEPDC, the authors show a reversal of the effect of fasting on PTZ-induced seizures. These experiments thus highlight the potential interaction between DEPDC5 and mTORC1 in the fasted state.

To better elucidate the mechanisms by which amino acids interact with mTORC1, the authors used primary neuronal culture and the addition or exclusion of specific amino acids and growth factors to study mTORC1 signaling. Withdrawal of amino acids or growth factors led to decreased mTORC1 activity. Individual amino acids were further studied in this paradigm to better determine whether specific amino acids are responsible for the mTORC1 response. These studies revealed a key role for mTORC1 in sensing leucine, arginine, and glutamate.

To specifically determine whether nutrient sensing is GATOR1-dependent, the authors demonstrate that DEPDC5 knock down in neurons increases mTORC1 activity, which was partially reversed by nutrient withdrawal. While DEPDC5 is critical for sensing of key amino acids, there may be other regulatory elements at play contributing to responses of other nutrients. For example, the authors also determined that while TSC2 may not be involved in amino acid-sensing, this protein inhibits mTORC1 in response to growth factor withdrawal. Furthermore, a double knockdown of both DEPDC5 and TSC2 reveals a significant, although not complete, reduction in mTORC1.

Finally, the authors used metabolomic analysis to evaluate changes in various metabolites in DEPDC5-deficient mouse brain. Using fasted and fed states, the authors demonstrate that DEPDC5-deficiency produces a reduction in amino acids, independent of the potential benefits of fasting. This mutation is also associated with a high mortality rate, which may be important for sudden unexpected death in epilepsy. As serotonin has emerged as an important regulator of respiratory responses to seizures, ⁶ tryptophan and serotonin were also examined and found to be deficient in Depdc knockout mice, which was not rescued by fasting.

The beneficial effects of fasting on seizure control extend to dietary therapies including the ketogenic diet. Furthermore, the anti-seizure effects of fasting and the ketogenic diet may be due to distinct but overlapping mechanisms. Fasting increases neuroprotective proteins such as brain-derived neurotrophic factor, and decreases the expression of inflammatory cytokines, leading to improved overall bioenergetics and neuronal resilience. ⁷ Fasting also increases the expression of ketone bodies, most notably beta-hydroxybutyrate, which is also a principal player in the anti-seizure effects of the ketogenic diet. Beta-hydroxybutyrate not only acts as an energy source but also improves glucose consumption, increases GABA synthesis, and has direct anti-seizure effects. ⁸ These multiple modes of action suggest distinct potential signaling pathways contributing to the effect of fasting and the ketogenic diet on seizure control. The studies described by Yuskaitis et al ⁴ demonstrate a critical role for upstream regulation of mTORC1 in controlling seizures, where DEPDC5 responds to nutrient changes resulting from fasting. For many patients and caregivers, dietary therapies may be difficult to maintain and therefore there is a need for new treatments that duplicate the beneficial effects of fasting or dietary therapy. A greater understanding of signaling pathways intervening between caloric restriction and neuronal hyperexcitability may reveal novel targets for therapeutic intervention.

Cameron S. Metcalf, Ph.D
Department of Pharmacology and Toxicology, The University of Utah