There is a fundamental gap in understanding how an appropriate balance of excitatory and inhibitory (E/I) connectivity is achieved during development of cortical networks and adjusted through synaptic plasticity for normal functioning of the cerebral cortex. Until this gap is filled, understanding neuropsychiatric disorders with GABAergic inhibitory connection deficits, such as schizophrenia and autism, will remain a mystery. The long term goal is to identify the molecular mechanisms that establish E/I balance in the prefrontal cortex, which may identify new targets for disorders where this balance is altered. The objective is to define a novel mechanism for limiting inhibitory connections between basket interneurons and the perisomatic region of pyramidal neurons in developing prefrontal cortex. The central hypothesis is that neural cell adhesion molecule NCAM, tyrosine kinase EphA3, and ADAM10 metalloprotease comprise a presynaptic receptor complex for postsynaptic ephrinA5 that promotes elimination of perisomatic synapses critical for proper prefrontal network organization and functioning, such as in working memory.
Aim 1. To identify a novel molecular mechanism for limiting perisomatic basket cell innervation in the developing mouse prefrontal cortex through NCAM-dependent ephrinA5/EphA3 signaling We will identify NCAM/EphA3 binding sites, assess the ability of NCAM to stabilize EphA3 on the cell surface by inhibiting endocytosis and promoting ephrinA5-induced EphA3 kinase signaling, and define the developmental and activity-dependent regulation of ephrinA5 in mouse prefrontal cortex.
Aim 2. To define presynaptic and postsynaptic functions of NCAM, ephrinA5/EphA3, and ADAM10 metalloprotease in perisomatic inhibitory synapse regulation Analysis of new conditional NCAM and ADAM10 mutant mice and cell-specific expression in brain slices will distinguish pre- versus post-synaptic functions for NCAM, ephrinA5/EphA3, and ADAM10, and test causal roles for their interactions in perisomatic synapse regulation. Dynamics of inhibitory synapse elimination will be analyzed by time-lapse two-photon microscopy in cortical slice cultures.
Aim 3. To delineate the contribution of NCAM to prefrontal cortical network organization and function using optogenetic mapping and behavioral assessment of working memory. Optogenetic mapping will be performed in brain slices from NCAM null and conditional mutant mice expressing channelrhodopsin-2 from the VGAT promoter in interneurons. Working memory performance will be measured in live mice by the delayed non-match-to-sample T-maze task. The outcome of these studies is expected to have a sustained, positive impact, because it will illuminate novel molecular mechanisms of interneuronal connectivity that control cognitive function, while innovative optogenetic technology will elucidate cortical networks targeted in neurodevelopmental disorders.

Public Health Relevance

The proposed research is relevant to public health because it seeks to define the molecular mechanisms that regulate the development and refinement of inhibitory circuits in the mammalian cortex. The research is relevant to the mission of NIH in illuminating basic mechanisms of neurodevelopment important for understanding inherited brain disorders, and is especially relevant for disorders such as schizophrenia and autism spectrum disorders where GABAergic interneuron dysfunction and a known genetic association of NCAM and ephrinA/EphA may contribute to the etiology of disease.

Agency
National Institute of Health (NIH)
Institute
National Institute of Mental Health (NIMH)
Type
Research Project (R01)
Project #
5R01MH101605-02
Application #
8821673
Study Section
Special Emphasis Panel (ZRG1)
Program Officer
Panchision, David M
Project Start
2014-04-01
Project End
2019-03-31
Budget Start
2015-04-01
Budget End
2016-03-31
Support Year
2
Fiscal Year
2015
Total Cost
Indirect Cost
Name
University of North Carolina Chapel Hill
Department
Biochemistry
Type
Schools of Medicine
DUNS #
608195277
City
Chapel Hill
State
NC
Country
United States
Zip Code
27599
Zhang, Xuying; Sullivan, Chelsea S; Kratz, Megan B et al. (2017) NCAM Regulates Inhibition and Excitability in Layer 2/3 Pyramidal Cells of Anterior Cingulate Cortex. Front Neural Circuits 11:19
Sullivan, Chelsea S; Kümper, Maike; Temple, Brenda S et al. (2016) The Neural Cell Adhesion Molecule (NCAM) Promotes Clustering and Activation of EphA3 Receptors in GABAergic Interneurons to Induce Ras Homolog Gene Family, Member A (RhoA)/Rho-associated protein kinase (ROCK)-mediated Growth Cone Collapse. J Biol Chem 291:26262-26272
Barão, Soraia; Gärtner, Annette; Leyva-Díaz, Eduardo et al. (2015) Antagonistic Effects of BACE1 and APH1B-?-Secretase Control Axonal Guidance by Regulating Growth Cone Collapse. Cell Rep 12:1367-76
Hernandez-Enriquez, Berenice; Wu, Zhuhao; Martinez, Edward et al. (2015) Floor plate-derived neuropilin-2 functions as a secreted semaphorin sink to facilitate commissural axon midline crossing. Genes Dev 29:2617-32
Demyanenko, Galina P; Mohan, Vishwa; Zhang, Xuying et al. (2014) Neural cell adhesion molecule NrCAM regulates Semaphorin 3F-induced dendritic spine remodeling. J Neurosci 34:11274-87