Epilepsy is a common disorder and approximately 30% of patients are refractory to treatments. There has been growing attention to metabolic dysfunction as a causative factor in epilepsy. Recent studies have identified mutations in the SLC13A5 gene that codes for the NaCT Na+/citrate transporter in children with early onset epileptic encephalopathy. The NaCT transporter is located on the plasma membrane and carries citrate and succinate into the cell. The effect of a loss in transport of exogenous citrate, and possibly succinate, in neurons and astrocytes and the mechanism of development of epilepsy in these patients is completely unexplored. This exploratory proposal is to define the effect of loss of NaCT expression on transport of citrate and succinate in cultured neurons and astrocytes, and to understand the metabolic and bioenergetic consequences of this loss. We hypothesize that exogenous citrate, or possibly succinate, has an important role in neuronal bioenergetics and susceptibility to excitotoxicity. In addition, citrate is the starting material for lipid/membrane and cholesterol synthesis and can act as a divalent cation chelator. The potential roles for NaCT transport will be explored in two Specific Aims: 1. To define NaCT-mediated citrate or succinate transport in cultured neurons and astrocytes. NaCT expression will be manipulated by genetic means or coexpression of epilepsy-related NaCT mutants. We will also manipulate NaCT activity with novel high affinity, high specificity pharmacological inhibitors. 2. To determine the metabolic consequence of inhibiting or overexpressing NaCT in cultured neurons and astrocytes. Cellular bioenergetics will be assessed as will the metabolic fate of exogenous citrate and succinate. This multi-PI project will merge expertise in neuronal metabolism and in NaCT transport physiology. These experiments will be the first examination of diminished NaCT transporter to neuronal or astrocyte function. Groundwork will be laid for future, more in-depth studies of the mechanisms by which citrate and/or succinate homeostasis affects neuronal excitability and the development of epilepsy. We anticipate our findings to identify potential therapeutic strategies that relate not only to epilepsies resulting from the NaCT mutations but to other medically refractory forms as well.
Genetic mutations that inactivate the Na+-dependent citrate transporter (NaCT) on neurons are associated with early onset epilepsy that does not respond to most current therapies, yet the consequences of NaCT loss to neuronal function have not been investigated. The NaCT transports a number of metabolic intermediates including citrate and succinate. Citrate participates in fatty acid and cholesterol biosynthesis, energy sensing and overall metabolic regulation, and succinate is a critical participant in mitochondrial metabolism and ATP synthesis. The proposed experiments may reveal new strategies to treat this as well as other forms of epilepsy and neurodegenerative disease.
