Tracy S. Gertler, Ph.D. - US grants

[unreadable] DESCRIPTION (provided by applicant): Schizophrenia affects 1% of the population worldwide; neuroleptic (antipsychotic) medications are essential for treatment despite an incomplete understanding of the mechanisms through which palliative symptomatic remediation is exerted. Though different in the degree of antagonism for the dopamine 2 receptor (D2R) and extradopaminergic effects, both 'typical1 neuroleptics (e.g. haloperidol) and 'atypical1 neuroleptics such as clozapine are effective at long-term remodeling of the corticostriatal synapse, a primary site of disease pathophysiology and pharmacotherapeutic activity as demonstrated by in vivo imaging and postmortem tissue analyses. Previous study has been impeded by an inability to experimentally separate a heterogeneous cell population in the striatum, and by inadequate resolution from the soma of dendritic processes. BAG transgenic mice expressing eGFP under the dopamine 1 receptor (D1R) and D2R promoters will be used to distinguish dichotomous principal neuron populations in the striatum, in combination with 2-photon laser scanning microscopy (2PLSM) to gain insight into dendritic function. As adaptations to chronic neuroleptic administration may be different in normal versus diseased states, an RGS4 knockout mouse will be employed as a schizophrenic-like model in parallel to wildtype strains; RGS4 has been implicated as both a schizophrenia susceptibility gene in cases of demonstrable hereditary transmission and a gene sensitive to altered dopamine transmission in the striatum of Parkinsonian animal models. In summary, this proposal aims to characterize the compound effect by which pharmacotherapeutic improvement in clinical symptomatology is effected. Whole-cell patch clamp electrophysiology, combined with molecular transgenic, pharmacologic, and 2-photon imaging modalities will be used to study the remodeling of neuronal subpopulations in the dorsal striatum of wildtype and RGS4 knockout mice following subchronic typical (i.e. haloperidol) and atypical (i.e. clozapine) neuroleptic administration. Specific Aim 1 will examine the intrinsic, morphologic, and dendritic properties of medium spiny neurons of the dorsal striatum. To assess functional connectivity between prefrontal cortical and striatal neurons, Specific Aim 2 will target presynaptic and postsynaptic adjustments in the corticostriatal synapse. [unreadable] [unreadable] [unreadable]

Project summary Epilepsy is among the most common childhood neurologic disorders, affecting 40 children per 100,000 in the US alone. Children with seizure onset before age one have a six-fold increase in early mortality, due in part to a disproportionately poor response to conventional anticonvulsants. The availability of genetic testing has dramatically improved etiologic diagnoses of early-onset epilepsy, as ~26% of early-life epilepsy is now associated with pathogenic genetic mutations in a variety of genes such as ion channels. Yet, precisely- targeted therapeutic options remain limited. Missense pathogenic variants in KCNT1, a gene encoding a sodium-activated potassium channel, are causative for ~ 40% of cases of a severe infantile-onset epilepsy called epilepsy of infancy with migrating focal seizures (EIMFS), suggestive of a strong genotype-phenotype relationship. As a hallmark of EIMFS is medically-refractory seizures, targeting its pathogenic mechanism is an opportunity for novel anticonvulsant intervention. The goal of the proposed studies is to define the pathophysiologic mechanisms that lead to seizures in EIMFS so that anticonvulsant therapies can be rationally chosen and applied early in the disease course. Aim 1 delineates the cellular mechanisms governing a de novo KCNT1 gain-of-function variant in human neurons differentiated from patient-derived induced pluripotent stem cells (iPSCs). We hypothesize that altered KCNT1 channel kinetics result in increased persistent potassium current, impairing high-frequency firing of inhibitory neurons. Aim 2 combines detailed phenotyping of a mouse model of KCNT1-associated epilepsy with acute slice electrophysiology of labeled interneuron subpopulations. We hypothesize that hippocampal interneurons will be differentially affected by a gain-of- function Kcnt1 knock-in variant, evidenced by decreased action potential firing, with resultant decreased pre- synaptic GABA release and excessive excitatory neuron bursting. Taken together, these studies will broaden our understanding of the cellular mechanisms by which KCNT1 mutations contribute to the pathogenesis of severe childhood epilepsy, laying the groundwork for development of precise pharmacotherapies for EIMFS. This application is for a K08 Career Development Award for Tracy Gertler, M.D., Ph.D., Child Neurology Instructor at Lurie Children?s Hospital. To become an independent physician-scientist in the fields of ion channel physiology and neurogenetics, Dr. Gertler will commit the majority of her post-medical training to research in genetic epilepsy due to ion channelopathies. The division of pediatric neurology within the pediatrics department has an unwavering commitment to the career development of the candidate as she takes advantage of her neurophysiology background and adds training in applied stem cell biology and gene- editing and phenotyping of animal models of epilepsy under the mentorship of Drs. Alfred L. George, Jr. and Jennifer Kearney.