Synapses of the mammalian central nervous system (CNS) are very deeply diverse in both molecular and functional properties. At present, unfortunately, our understanding of this diversity is rudimentary, and quantitative data on the subject are very few. Left unfathomed, CNS synapse diversity poses formidable obstacles to better understanding of the development, function and disorders of the brain's synaptic circuitry. The major reason for the persistence of this distressing state of ignorance lies in the fact that tools for exploring synapse populations at the level of individual synapses are few and limited in their capabilities. To address the challenges synapse diversity poses to both basic and clinical neuroscience, this project aims to develop a superlative new proteometric imaging platform capable of analyzing very large synapse populations in situ with single- synapse resolution. Deployment and dissemination of this platform will facilitate study and treatment of the many neurodevelopmental, mental and neurodegenerative disorders linked to specific synapse subpopulations, as well as opening new perspectives on molecular mechanisms, circuit architectures and disorders of CNS memory encoding, storage and retrieval. The platform will be based on immunofluorescence array tomography (IAT) and involve development of novel antibody standardization protocols and novel image acquisition hardware and software. These innovations will improve the reproducibility, quantitative reliability, and speed of IAT by large margins and overcome limitations that have so far prevented proteometric analysis of large synapse populations at the single-synapse level. The platform would be capable of proteometric census of a million of more synapses per hour, at 50 or more markers per synapse, while maintaining precise neuroanatomical and molecular coordinates for each synapse. The new platform will be demonstrated by a 7-marker proteometric survey of an adult mouse cortex barrel column that would enumerate each of the tens of millions of synapses in the column and allow classification of each synapse based on neurotransmitter type and a set of neurons type markers. The results will be disseminated via methods publications, open-source

Public Health Relevance

Abnormalities of individual synapses and of the brain's large and diverse synapse populations are widely believed to account for many or most neurodevelopmental, neurodegenerative and substance-abuse-related brain disorders. This work will address major gaps in present knowledge of such abnormalities by developing superlative new tools for the very rapid and highly detailed quantitative survey of large synapse populations in both research animal and human brain specimens.

Agency
National Institute of Health (NIH)
Institute
National Institute of Mental Health (NIMH)
Type
Exploratory/Developmental Grants (R21)
Project #
5R21MH099797-02
Application #
8544503
Study Section
Special Emphasis Panel (ZMH1-ERB-S (05))
Program Officer
Asanuma, Chiiko
Project Start
2012-09-12
Project End
2014-07-31
Budget Start
2013-08-01
Budget End
2014-07-31
Support Year
2
Fiscal Year
2013
Total Cost
$188,400
Indirect Cost
$68,400
Name
Stanford University
Department
Biophysics
Type
Schools of Medicine
DUNS #
009214214
City
Stanford
State
CA
Country
United States
Zip Code
94305
Valenzuela, Ricardo A; Micheva, Kristina D; Kiraly, Marianna et al. (2016) Array tomography of physiologically-characterized CNS synapses. J Neurosci Methods 268:43-52
Ning, Lipeng; Setsompop, Kawin; Michailovich, Oleg et al. (2016) A joint compressed-sensing and super-resolution approach for very high-resolution diffusion imaging. Neuroimage 125:386-400
Micheva, Kristina D; Wolman, Dylan; Mensh, Brett D et al. (2016) A large fraction of neocortical myelin ensheathes axons of local inhibitory neurons. Elife 5:
Ning, Lipeng; Laun, Frederik; Gur, Yaniv et al. (2015) Sparse Reconstruction Challenge for diffusion MRI: Validation on a physical phantom to determine which acquisition scheme and analysis method to use? Med Image Anal 26:316-31
Burette, Alain; Collman, Forrest; Micheva, Kristina D et al. (2015) Knowing a synapse when you see one. Front Neuroanat 9:100
Collman, Forrest; Buchanan, JoAnn; Phend, Kristen D et al. (2015) Mapping synapses by conjugate light-electron array tomography. J Neurosci 35:5792-807
Ning, Lipeng; Setsompop, Kawin; Michailovich, Oleg et al. (2015) A Compressed-Sensing Approach for Super-Resolution Reconstruction of Diffusion MRI. Inf Process Med Imaging 24:57-68
PaÅŸca, Anca M; Sloan, Steven A; Clarke, Laura E et al. (2015) Functional cortical neurons and astrocytes from human pluripotent stem cells in 3D culture. Nat Methods 12:671-8
Ning, Lipeng; Westin, Carl-Fredrik; Rathi, Yogesh (2015) Estimating diffusion propagator and its moments using directional radial basis functions. IEEE Trans Med Imaging 34:2058-78
Weiler, Nicholas C; Collman, Forrest; Vogelstein, Joshua T et al. (2014) Synaptic molecular imaging in spared and deprived columns of mouse barrel cortex with array tomography. Sci Data 1:140046

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