Rapid purification and metabolomic profiling of synaptic vesicles from mammalian brain

  1. Lynne Chantranupong
  2. Jessica L Saulnier
  3. Wengang Wang
  4. Drew R Jones
  5. Michael E Pacold
  6. Bernardo L Sabatini  Is a corresponding author
  1. Harvard Medical School, United States
  2. NYU Langone Health, United States
  3. Howard Hughes Medical Institute, Harvard Medical School, United States

Abstract

Neurons communicate by the activity-dependent release of small-molecule neurotransmitters packaged into synaptic vesicles (SVs). Although many molecules have been identified as neurotransmitters, technical limitations have precluded a full metabolomic analysis of synaptic vesicle content. Here, we present a workflow to rapidly isolate SVs and to interrogate their metabolic contents at high-resolution using mass spectrometry. We validated the enrichment of glutamate in SVs of primary cortical neurons using targeted polar metabolomics. Unbiased and extensive global profiling of SVs isolated from these neurons revealed that the only detectable polar metabolites they contain are the established neurotransmitters glutamate and GABA. In addition, we adapted the approach to enable quick capture of SVs directly from brain tissue and determined the neurotransmitter profiles of diverse brain regions in a cell-type specific manner. The speed, robustness, and precision of this method to interrogate SV contents will facilitate novel insights into the chemical basis of neurotransmission.

Data availability

All data generated or analysed during this study are included in the manuscript and supporting files. Source data files have been provided for all figures.

Article and author information

Author details

  1. Lynne Chantranupong

    Neurobiology, Harvard Medical School, Boston, United States
    Competing interests
    The authors declare that no competing interests exist.
  2. Jessica L Saulnier

    Neurobiology, Harvard Medical School, Boston, United States
    Competing interests
    The authors declare that no competing interests exist.
  3. Wengang Wang

    Neurobiology, Harvard Medical School, Boston, United States
    Competing interests
    The authors declare that no competing interests exist.
  4. Drew R Jones

    Metabolomics Core Resource Laboratory, NYU Langone Health, New York, United States
    Competing interests
    The authors declare that no competing interests exist.
  5. Michael E Pacold

    Radiation Oncology, NYU Langone Health, New York, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-3688-2378
  6. Bernardo L Sabatini

    Department of Neurobiology, Howard Hughes Medical Institute, Harvard Medical School, Boston, United States
    For correspondence
    bsabatini@hms.harvard.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-0095-9177

Funding

Howard Hughes Medical Institute (Investigator)

  • Bernardo L Sabatini

National Institute of Neurological Disorders and Stroke (R37NS046579)

  • Bernardo L Sabatini

Howard Hughes Medical Institute (Hanna Gray Fellowship)

  • Lynne Chantranupong

Mary Kay Foundation (Cancer Research Grant 017-032)

  • Michael E Pacold

Hearst Foundation (V Foundation V Scholar Grant (V2017-004))

  • Michael E Pacold

National Cancer Institute (K22 Career Transition Award (1K22CA212059))

  • Michael E Pacold

NIH (R01 NS108151-01)

  • Drew R Jones

FNIH (RFA 2018-PACT001)

  • Drew R Jones

NIH (HHS-NIH-NIAD-BAA2018)

  • Drew R Jones

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Reviewing Editor

  1. Axel T Brunger, Stanford University, United States

Ethics

Animal experimentation: All experimental manipulations were performed in accordance with protocols (#03551) approved by the Harvard Standing Committee on Animal Care following guidelines described in the US National Institutes of Health Guide for the Care and Use of Laboratory Animals.

Version history

  1. Received: June 5, 2020
  2. Accepted: October 11, 2020
  3. Accepted Manuscript published: October 12, 2020 (version 1)
  4. Version of Record published: October 20, 2020 (version 2)

Copyright

© 2020, Chantranupong et al.

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

Metrics

  • 6,061
    views
  • 785
    downloads
  • 24
    citations

Views, downloads and citations are aggregated across all versions of this paper published by eLife.

Download links

A two-part list of links to download the article, or parts of the article, in various formats.

Downloads (link to download the article as PDF)

Open citations (links to open the citations from this article in various online reference manager services)

Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)

  1. Lynne Chantranupong
  2. Jessica L Saulnier
  3. Wengang Wang
  4. Drew R Jones
  5. Michael E Pacold
  6. Bernardo L Sabatini
(2020)
Rapid purification and metabolomic profiling of synaptic vesicles from mammalian brain
eLife 9:e59699.
https://doi.org/10.7554/eLife.59699

Share this article

https://doi.org/10.7554/eLife.59699

Further reading

    1. Neuroscience
    Songyao Zhang, Tuo Zhang ... Tianming Liu
    Research Article

    Cortical folding is an important feature of primate brains that plays a crucial role in various cognitive and behavioral processes. Extensive research has revealed both similarities and differences in folding morphology and brain function among primates including macaque and human. The folding morphology is the basis of brain function, making cross-species studies on folding morphology important for understanding brain function and species evolution. However, prior studies on cross-species folding morphology mainly focused on partial regions of the cortex instead of the entire brain. Previously, our research defined a whole-brain landmark based on folding morphology: the gyral peak. It was found to exist stably across individuals and ages in both human and macaque brains. Shared and unique gyral peaks in human and macaque are identified in this study, and their similarities and differences in spatial distribution, anatomical morphology, and functional connectivity were also dicussed.

    1. Neuroscience
    Avani Koparkar, Timothy L Warren ... Lena Veit
    Research Article

    Complex skills like speech and dance are composed of ordered sequences of simpler elements, but the neuronal basis for the syntactic ordering of actions is poorly understood. Birdsong is a learned vocal behavior composed of syntactically ordered syllables, controlled in part by the songbird premotor nucleus HVC (proper name). Here, we test whether one of HVC’s recurrent inputs, mMAN (medial magnocellular nucleus of the anterior nidopallium), contributes to sequencing in adult male Bengalese finches (Lonchura striata domestica). Bengalese finch song includes several patterns: (1) chunks, comprising stereotyped syllable sequences; (2) branch points, where a given syllable can be followed probabilistically by multiple syllables; and (3) repeat phrases, where individual syllables are repeated variable numbers of times. We found that following bilateral lesions of mMAN, acoustic structure of syllables remained largely intact, but sequencing became more variable, as evidenced by ‘breaks’ in previously stereotyped chunks, increased uncertainty at branch points, and increased variability in repeat numbers. Our results show that mMAN contributes to the variable sequencing of vocal elements in Bengalese finch song and demonstrate the influence of recurrent projections to HVC. Furthermore, they highlight the utility of species with complex syntax in investigating neuronal control of ordered sequences.