Russell J. Ferland, PhD - US grants

[unreadable] DESCRIPTION (provided by applicant): Developmental malformations of the brain are collectively recognized as an increasingly important cause of mental retardation, epilepsy, and perhaps autism. These disorders have deleterious effects on both the psychological and physical well-being of the affected individual. However, identification of causative genes and elucidation of their functions in the context of these developmental brain abnormalities provide valuable insights into human brain development. Joubert syndrome (JS) is an autosomal recessive developmental brain condition, which is characterized anatomically by cerebellar and brainstem malformations, and clinically by hypotonia, breathing abnormalities, atypical eye movements, cognitive problems, and autistic features. The goals of this proposal are to identify one of the causative genes for JS and to study the function of this gene in brain development. Our preliminary work mapped a JS locus to chromosome 6q. Our further studies identified a causative gene for JS, which is a novel gene encoding a putative adaptor protein called AHI1. Specific Aim 1 will test the hypothesis that AHI1 is a causative gene for JS by further analyzing the genetic and clinical aspects of JS in our patients, performing mutational analyses of AHI1 in our JS families, and analyzing the genomic structure of AHI1. Specific Aim 2 will characterize the temporal and spatial expression of AHI1 at both the mRNA and protein levels in addition to generating and characterizing mice with a targeted deletion of Ahi1. Specific Aim 3 will identify proteins that interact with AHI1 by yeast two-hybrid and confirm these interactions by co-immunoprecipitation analyses. The applicant has a PhD. He earned his doctoral degree in Neuroscience studying the role of the perirhinal cortex, the ventromedial nucleus of the hypothalamus, and hippocampal neurogenesis in animal models of epilepsy. His mentor is Christopher A. Walsh, MD, PhD, whose research interests center on genetic approaches toward understanding basic mechanisms governing the development of the brain. This research proposal focuses on training the candidate in methods of 1) genetic mapping, linkage analysis, and genetic analyses, and 2) neurogenetics and molecular biological approaches to brain development. It is the candidate's intention to combine both his previous training in neuroanatomy and behavior with these new molecular and genetic approaches in order to pursue an academic career in Neuroscience. [unreadable] [unreadable]

DESCRIPTION (provided by applicant): Epilepsy is one of the most common neurological diseases in the world. This makes epilepsy a serious issue not only for those affected individuals and their families, but also for society and the medical profession. While many of the current treatments for epilepsy involve blocking the symptoms of the epilepsy, namely the seizures, there are few strategies that can ameliorate the ongoing reorganizational processes that are occurring in the epileptic brain. Therefore, elucidating the mechanisms responsible for this reorganization is vital for a better understanding of this process and for developing therapies that can ultimately prevent epilepsy. We have refined a rodent model of seizures using the chemoconvulsant, flurothyl. In this model, initial myoclonic jerk threshold, initial generalized seizure threshold, and two reorganizational/epileptogenic processes can be studied, 1) decreases in seizure threshold over 8 seizure trials and 2) alterations in seizure behavior, both of which occur over time. Our data suggest the hypothesis that there is significant genetic control of the processes underlying these reorganizational events observed in the repeated-flurothyl model as exemplified by differences observed between C57BL/6J and DBA/2J mice. Moreover, these traits segregate independently in C57BL/6J x DBA/2J (BXD) recombinant inbred (RI) lines. Thus, the goals of this proposal are to map quantitative trait loci (QTLs) in BXD animals that control each aspect of the flurothyl seizure characteristics observed, and determine the genetic and molecular pathways that mediate the reorganizational processes in the brain. We also will determine the neuroanatomical underpinnings of the reorganizational processes, which occur in the flurothyl model, through analysis of detailed neuroactivity maps in the brain, changes in brain circuitry following repeated seizures, and the role of BDNF in these processes. Lastly, we will utilize the extensive brain expression dataset in the genenetwork.org database to correlate differential gene expression in BXD brains with seizure phenotypes in an effort to identify potential biomarkers to predict the likelihood of seizure occurrence. The repeated-flurothyl model will allow for greater insight into the genetic control of the generalized seizure threshold and brain reorganization, and can also lead to the identification of QTLs (and the underlying genes) that are directly responsible for these processes, and move beyond studies that examine generalized seizure threshold solely. As a whole, these studies will provide greater insight into the process of brain reorganization from the genetic to the systems level. PUBLIC HEALTH RELEVANCE: Epilepsy is one of the most common neurological diseases, affecting approximately 1 - 4% of the total population by 80 years of age. While many of the current treatments for epilepsy involve blocking the symptoms of the epilepsy through seizures suppression, no current therapies target the ongoing epileptogenesis/reorganization that occurs in the epileptic brain. Our project is designed to better understand processes that are involved in the reorganization of the brain that accounts for the differences in seizure traits in different strains of mice. Our goal, using the repeated flurothyl model in mice, is to identify gene(s) and molecular processes that are altered following repeated seizures, accounting for the brain reorganization that can occur in the epileptic brain.

? DESCRIPTION (provided by applicant): Apical abscission (AA) describes a recently observed process whereby neuroepithelal cells detach from the ventricular lining of the cortex, leaving an abscissed fragment at the apical membrane and releasing a newly formed migratory neuron/ intermediate progenitor to migrate into the cortical plate. This step is required for neuronal proliferation, initial migration from the ventricular zone, and maintenance of tissue architecture along the neuroependyma. Disruption of this process contributes to periventricular heterotopia (PH), a malformation of cortical development characterized by a smaller brain (impaired proliferation), abnormal neurons clustered deep in the brain along the lateral ventricles (failed migration) and loss in neuroependymal integrity (lost tissue architecture). The mechanisms that govern AA are not known. Given the shared features between AA in normal development and PH in disease, we hypothesize that genes causal for PH regulate AA. We have identified several genes causal for PH, including the actin binding Filamin A (FLNA) and vesicle trafficking associated ADP-ribosylation factor guanine exchange factor 2 (ARFGEF2) genes. In our preliminary data, we have identified a novel FlnA binding protein, Fmn2, which has been implicated in both actin regulation and endosomal trafficking. Disruption of Fmn2 causes the same defects in neocortical brain development in mice, similar to that seen with loss of FlnA and Big2. We have engineered Fmn2 knockout mice and Fmn2-FlnA double knockout mice, as well as developed techniques for transient over-expression/ inhibition of these various genes by in utero electroporation. In Aim1, we will determine what steps in AA (timing, cell polarity, cell fat, cell adhesion, actin) are affected by disruption of PH genes using wide field timed lapse microscopy on ex-vivo embryo slice cultures from knockout mice and following in utero electroporation. In Aim2, we will investigate step-wise interactions between the various PH genes in regulation of AA, through in utero electroporation of various PH gene constructs in constitutively active/inactive states or following over- expression/ inhibition. In Aim3, we will examine whether the proliferative changes seen following disruption of PH associated genes is a result of impaired AA.