Our goal is to understand the mechanism of metabolically induced resistance to epileptic seizures. Epilepsy is extremely common, affecting on the order of 1% of the population, and about a third of people with epilesy are not well treated by existing medications. Dietary therapies such as the ketogenic diet can be very effective for such pharmacoresistant epilepsy, but the diets are often difficul for patients and providers. However, if we can understand the mechanism of the seizure resistance produced by these diets, it may be possible to find new medications to treat a wide range of currently poorly treated epilepsies. In the previous grant period, we found suppor for the hypothesis that ATP--sensitive potassium channels (KATP channels) are critical for metabolic seizure resistance. We observed that KATP channels in neurons could be activated synergistically by neuronal activity and by ketone bodies (which are an alternative fuel used by the brain on the ketogenic diet). We discovered a mouse genetic model for metabolic seizure resistance that does not require a change in diet and learned that in this model, KATP channels are required for the seizure resistance. We also engineered new fluorescent biosensors to detect intracellular changes of the key metabolic cofactors ATP and NADH, to enable a better understanding of cellular metabolism.
Our first aim for this grant renewal is to learn how brain cell metabolism responds o the energy demands of stimulation and how those responses depend on different cellular fuels (such as the ketone bodies that become available to the brain on a ketogenic diet). Our own biosensors and others will be used to monitor the responses, in individual cells, of specific key metabolic signals: ATP, NADH, glutathione redox, and AMP-- activated protein kinase activity. Each of these signals not only reports on metabolism but also has effects on downstream mechanisms that affect neuronal excitability. We will study these responses in cultured hippocampal neurons, for which we have optimal control of extracellular fuels. We will also study acute brain slices, which will allow us to examine distinctive behavior i excitatory neurons, inhibitory interneurons, and nearby astrocytes, in response to either synaptic or electrical stimulation. These studies should provide an unprecedented view of metabolic responses in brain cells, and give fundamental insights into how neurons and astrocytes respond to energy demands. These insights will be valuable not only for understanding the basis of metabolic seizure resistance, but also for how brain cells respond to metabolic challenges in disorders ranging from traumatic brain injury to neurodegeneration.
Our second aim i s to link metabolic changes to KATP channel activity by learning how KATP channels respond to energy levels in intact neurons. We will record the ope probability of KATP channels simultaneously with biosensor measurements of [ATP] or ATP:ADP ratio, to learn the actual dose--response relation for intact neurons, and the conditions for engaging these channels to produce seizure resistance.

Public Health Relevance

One of the best treatments for epilepsy (a seizure disorder affecting roughly 1% of the population) is a very- low-carbohydrate, high-fat ketogenic diet. Because the diet is unpalatable and difficult, it would be useful to understand how it acts on brain cells so that better drug therapies (or easier diets) can be designed. This project will stud how neuronal energy status responds when neurons are excited (during normal or seizure- like activity), how different cellular fuels (glucose versus the ketone bodies that supply the brain on ketogenic diet) affect the energy response, and how these energy responses translate into anti-seizure effects on the neuronal electrical response.

National Institute of Health (NIH)
National Institute of Neurological Disorders and Stroke (NINDS)
Research Project (R01)
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Clinical Neuroplasticity and Neurotransmitters Study Section (CNNT)
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Whittemore, Vicky R
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Harvard Medical School
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Martínez-François, Juan Ramón; Fernández-Agüera, María Carmen; Nathwani, Nidhi et al. (2018) BAD and KATP channels regulate neuron excitability and epileptiform activity. Elife 7:
Yellen, Gary (2018) Fueling thought: Management of glycolysis and oxidative phosphorylation in neuronal metabolism. J Cell Biol 217:2235-2246
Foley, Jeannine; Burnham, Veronica; Tedoldi, Meghan et al. (2018) BAD knockout provides metabolic seizure resistance in a genetic model of epilepsy with sudden unexplained death in epilepsy. Epilepsia 59:e1-e4
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Hung, Yin P; Teragawa, Carolyn; Kosaisawe, Nont et al. (2017) Akt regulation of glycolysis mediates bioenergetic stability in epithelial cells. Elife 6:
Lutas, Andrew; Lahmann, Carolina; Soumillon, Magali et al. (2016) The leak channel NALCN controls tonic firing and glycolytic sensitivity of substantia nigra pars reticulata neurons. Elife 5:
Mongeon, Rebecca; Venkatachalam, Veena; Yellen, Gary (2016) Cytosolic NADH-NAD(+) Redox Visualized in Brain Slices by Two-Photon Fluorescence Lifetime Biosensor Imaging. Antioxid Redox Signal 25:553-63
Yellen, Gary; Mongeon, Rebecca (2015) Quantitative two-photon imaging of fluorescent biosensors. Curr Opin Chem Biol 27:24-30
Hung, Yin Pun; Yellen, Gary (2014) Live-cell imaging of cytosolic NADH-NAD+ redox state using a genetically encoded fluorescent biosensor. Methods Mol Biol 1071:83-95
Lutas, Andrew; Birnbaumer, Lutz; Yellen, Gary (2014) Metabolism regulates the spontaneous firing of substantia nigra pars reticulata neurons via KATP and nonselective cation channels. J Neurosci 34:16336-47

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