Thalamocortical network dysfunction in a novel genetic model of GRIN2D developmental and epileptic encephalopathy

PROJECT SUMMARY Developmental and epileptic encephalopathy (DEE) is a collection of severe childhood seizure disorders, with a significant and diverse genetic component. DEE patients usually experience a significant seizure burden and suffer from cognitive and developmental impairments, as well as sleep and motor disturbances. The 50 or more genes harboring DEE-causing mutations include the ?GRIN? genes, which encode components of membrane protein complexes that are important for electrochemical signaling between neurons. Mutations in the GRIN2D subunit cause particularly severe and intractable DEE. We recently developed a mouse model of GRIN2D carrying a mutation that has recurred several times in GRIN2D DEE patients. These mice have very robust epileptic features, including both convulsive seizures and non-convulsive seizures known to be regulated by signaling between the cerebral cortex and the thalamus (i.e. thalamocortical network). This thalamocortical network is also associated with regulation of sleep, awareness, and the activity of many other brain regions. In preliminary findings, we also noticed that the protein product of Grin2D is expressed in excitatory and inhibitory cells of the cortex, while it is enriched in the inhibitory cells of the thalamus. Additionally, the primary sites of neuronal signaling ? the synapse ? have unusual structural features in the cortex, suggesting that they do not develop normally. Altogether, these preliminary data motivate an examination of how mutation in Grin2D alters the function of the cortex and thalamus. In this pilot program, we will apply electrophysiological approaches to measure the function of synapses in the GRIN2D DEE model to determine if structural changes correspond to functional impairments. We will also identify the neuronal connections that are most severely affected within the thalamocortical network in GRIN2D DEE. Aim1 will use slice electrophysiology methods to examine population-level responses within the cortex and thalamus to look for susceptibility of these regions to generate seizure-like activity. We will also adapt innovative approaches to dissect the component parts of the response in the cortex, which will allow us to efficiently pinpoint amongst the many connections in the cortex, which are most likely altered by the Grin2D mutation. These network-level effects will be further explored in Aim2 using intracellular whole-cell patch clamp recordings of synaptic currents. Aim2 will assess two distinct modes of GRIN2D signaling, synaptic and tonic, which are associated with different cellular consequences and are likely involved in the GRIN2D DEE pathogenic mechanism. Together these experiments will determine the consequence of the GRIN2D DEE mutation on the activity of neural circuits, which are implicated in the generation of seizures in these animals, and will determine, at the cellular level, the underlying mechanism of epileptogenesis. This study is an important first step for determining how GRIN2D shapes the development and maintenance of thalamocortical circuits (under normal conditions and in a DEE model), information which will guide future efforts to develop targeted corrective therapeutics.