Epilepsy is caused by excessive neuronal excitability characterized by seizures, which are abnormal and uncontrolled discharges of action potentials. This common brain disorder is often caused by mutations in ion channels that form the basis for neuronal intrinsic excitability. Voltage-gated Kv7/KCNQ K+ channels potently inhibit repetitive and burst firing of action potentials. Kv7 activators are used as anti-epileptic drugs whereas mutations in Kv7.2 and Kv7.3 subunits cause epilepsy in humans. My work has shown that some epilepsy mutations impair polarized enrichment of Kv7 channels at the axonal surface, the subcellular domain governing action potential initiation and conduction, suggesting that their correct axonal targeting is critical for their physiologic function. However, the underlying mechanisms remain poorly understood. The goal of this proposal is to understand how axonal targeting of Kv7 channels is achieved and regulated by epilepsy mutations and neuronal activity, and how basal and regulated targeting impacts intrinsic excitability. This proposal is significant because dissecting these unexplored mechanisms will increase our understanding of the role of axonal Kv7 channels in regulating excitability, and foster the development of new therapeutic strategies for epilepsy that could enhance axonal surface density of Kv7. Given that chronic blockade of neuronal activity in hippocampus leads to temporal lobe epilepsy, characterizing the regulation of Kv7 axonal targeting by chronic activity blockade of hippocampal neurons may provide mechanistic insights into plasticity of intrinsic excitability and epileptogenesis. My work has shown that Kv7 enrichment at the axonal surface involves a region in the Kv7.2 C- terminal tail upstream of ankyrin-G binding domain. This axon-targeting domain contains phosphorylation sites and interacts with calmodulin, syntaxin 1A, AKAP79/150 and PIP2. In our preliminary studies, we discover that Kv7 axonal targeting is regulated by calmodulin-mediated exit from the endoplasmic reticulum, dynamin-mediated endocytosis, and phosphorylation of Kv7.2. We further show that homeostatic increase in excitability induced by chronic activity blockade accompanies reduction in Kv7 current, Kv7.3, and AKAP150. Based on these data, our hypothesis is that Kv7.2 interacting molecules and phosphorylation mediate basal and activity-regulated Kv7 axonal targeting by linking Kv7 to the core traffic machinery for polarized targeting. To test this hypothesis, Aim 1 will determine the traffic pathways that mediate Kv7 axonal targeting.
Aim 2 will identify Kv7.2 interacting proteins and phosphorylation that bind to the core traffic machinery for polarized targeting.
Aim 3 will characterize the regulation of Kv7 axonal targeting by chronic activity blockade. To execute these aims, we will use live imaging, immunostaining, binding assays, and electrophysiology in dissociated and organotypic hippocampal cultures, which preserve the functional and morphological polarity of neurons. We will also utilize epilepsy mutations, knock-out mice, and in vivo alterations of neuronal activity.
Enriched at the axonal surface, Kv7 channels potently inhibit repetitive and burst firing of action potentials, which is the hallmark for neuronal hyperexcitabilty leading to epilepsy. The proposed research will identify the molecular mechanisms of Kv7 axonal targeting and its epilepsy-causing mutations and determine the roles that this targeting plays in neuronal excitability and plasticity. This research will therefore foster the development of new therapeutic strategies that could enhance Kv7 axonal surface density to combat epilepsy.
|Liu, Dai-Chi; Seimetz, Joseph; Lee, Kwan Young et al. (2017) Mdm2 mediates FMRP- and Gp1 mGluR-dependent protein translation and neural network activity. Hum Mol Genet 26:3895-3908|
|Baculis, Brian C; Weiss, Amanda C; Pang, Weilun et al. (2017) Prolonged seizure activity causes caspase dependent cleavage and dysfunction of G-protein activated inwardly rectifying potassium channels. Sci Rep 7:12313|
|Zhu, Jiuhe; Lee, Kwan Young; Jewett, Kathryn A et al. (2017) Epilepsy-associated gene Nedd4-2 mediates neuronal activity and seizure susceptibility through AMPA receptors. PLoS Genet 13:e1006634|
|Jang, Sung-Soo; Jeong, Han Gil; Chung, Hee Jung (2017) Electroconvulsive Seizures in Rats and Fractionation of Their Hippocampi to Examine Seizure-induced Changes in Postsynaptic Density Proteins. J Vis Exp :|
|Jang, Sung-Soo; Chung, Hee Jung (2016) Emerging Link between Alzheimer's Disease and Homeostatic Synaptic Plasticity. Neural Plast 2016:7969272|
|Vega L, Johana C M; Lee, Min Kyung; Qin, Ellen C et al. (2016) Three Dimensional Conjugation of Recombinant N-Cadherin to a Hydrogel for In Vitro Anisotropic Neural Growth. J Mater Chem B 4:6803-6811|
|Jang, Sung-Soo; Royston, Sara E; Lee, Gunhee et al. (2016) Seizure-Induced Regulations of Amyloid-?, STEP61, and STEP61 Substrates Involved in Hippocampal Synaptic Plasticity. Neural Plast 2016:2123748|