Publication:
Isolation of large dense-core vesicles from bovine adrenal medulla for functional studies

Thumbnail Image

School / College / Institute

Program

KU Authors

Co-Authors

Preobraschenski, Julia
Ganzella, Marcelo
Jahn, Reinhard

Publication Date

Language

Embargo Status

NO

Journal Title

Journal ISSN

Volume Title

Alternative Title

Abstract

Large dense-core vesicles (LDCVs) contain a variety of neurotransmitters, proteins, and hormones such as biogenic amines and peptides, together with microRNAs (miRNAs). Isolation of LDCVs is essential for functional studies including vesicle fusion, vesicle acidification, monoamine transport, and the miRNAs stored in LDCVs. Although several methods were reported for purifying LDCVs, the final fractions are significantly contaminated by other organelles, compromising biochemical characterization. Here we isolated LDCVs (chromaffin granules) with high yield and purity from bovine adrenal medulla. The fractionation protocol combines differential and continuous sucrose gradient centrifugation, allowing for reducing major contaminants such as mitochondria. Purified LDCVs show robust acidification by the endogenous V-ATPase and undergo SNARE-mediated fusion with artificial membranes. Interestingly, LDCVs contain specific miRNAs such as miR-375 and miR-375 is stabilized by protein complex against RNase A. This protocol can be useful in research on the biological functions of LDCVs.

Source

Publisher

Nature Publishing Group (NPG)

Subject

Science and technology

Citation

Has Part

Source

Scientific Reports

Book Series Title

Edition

DOI

10.1038/s41598-020-64486-3

item.page.datauri

Link

Rights

Copyrights Note

Endorsement

Review

Supplemented By

Referenced By

3

Views

3

Downloads

View PlumX Details