The development, plasticity, and stability of dendrites and dendritic spines are defective in autism, mental retardation, stroke, and psychiatric diseases. Mutations or reduced levels of heterotrimeric laminin extracellular matrix proteins are associated with these human brain disorders. We provide evidence that neuron-specific ablation of the laminin alpha5 subunit in mice increases spine densities, destabilizes dendrite branches, and compromises normal synaptic transmission and animal behavior. We propose to elucidate the mechanisms by which laminin alpha5 and a new putative laminin alpha5 receptor we have discovered regulate dendrite and dendritic spine development and function. We will use complementary in vivo imaging, electrophysiological, biochemical, and genetic approaches to achieve the following aims:
Aim 1. Determine how laminin alpha5 regulates development, plasticity, and function of dendrites, dendritic spines, and synapses. Our data strongly suggest that laminin alpha5 controls dendrite branch and dendritic spine dynamics. We will use transcranial two-photon microscopy of dendrites in the somatosensory cortex, alone and in combination with sensory input manipulation, to reveal how the loss of laminin alpha5 impacts branch and spine dynamics during development and activity-driven plasticity. We will also use electron microscopy and whole cell recording to test the hypothesis that laminin alpha5 regulates synaptic transmission by controlling the structure, transmission properties, and plasticity of individual synapses.
Aim 2. Elucidate the composition, origin, and timing of function of alpha5-containing laminins in dendrite and spine development. We do not know which laminin beta and gamma chains partner with laminin alpha5, where they are produced, or when they act. We will use biochemical and genetic knockout approaches to identify laminin beta and gamma chains that associate with laminin alpha5 in neurons to regulate dendrite and spine development. We will also inactivate laminin alpha5 in specific cell types using inducible Cre transgenes to determine where and when laminin alpha5 is required to regulate dendrite and dendritic spine development.
Aim 3. Characterize SIRPalpha function in laminin alpha5-mediated dendrite and dendritic spine development. We have shown that the integrin alpha3beta1 receptor for laminin alpha5 mediates dendrite branch stability, but our genetic analysis indicates that other receptors are essential to mediate the effects of laminin alpha5 on dendritic spine development. Our data strongly suggest that the Signal Regulatory Protein alpha (SIRPalpha) transmembrane receptor serves as a novel laminin alpha5 receptor in the control of spine development. We will use cell adhesion assays and in vitro binding assays with purified proteins to identify which domains in SIRPalpha and alpha5-laminins mediate these interactions. We will test how excitatory neuron-specific ablation of SIRPalpha function alone or in combination with integrin alpha3beta1 affects dendrite and spine development and synaptic function and plasticity.

Public Health Relevance

Connections between nerve cells do not form or operate properly in the brains of individuals with autism and mental retardation and become destabilized in psychiatric diseases and following stroke. We have discovered that a protein in the gel-like substance that surrounds and supports nerve cells in the brain plays a critical role in regulating the formation and function of connections between nerve cells. We will determine specifically how this protein, laminin alpha5, regulates nerve cell development and fosters the proper formation of connections between nerve cells. The findings of our study could lead to new diagnostic approaches and treatment strategies for these brain disorders.

Agency
National Institute of Health (NIH)
Institute
National Institute of Neurological Disorders and Stroke (NINDS)
Type
Research Project (R01)
Project #
1R01NS089662-01
Application #
8796341
Study Section
Special Emphasis Panel (ZRG1-MDCN-T (02))
Program Officer
Mamounas, Laura
Project Start
2014-06-15
Project End
2019-04-30
Budget Start
2014-06-15
Budget End
2015-04-30
Support Year
1
Fiscal Year
2014
Total Cost
$350,347
Indirect Cost
$131,597
Name
Yale University
Department
Biochemistry
Type
Schools of Medicine
DUNS #
043207562
City
New Haven
State
CT
Country
United States
Zip Code
06520