Bimodal dynamics of granular organelles in primary renin-expressing cells revealed using TIRF microscopy

Buckley, Charlotte and Dun, Alison R. and Peter, Audrey and Bellamy, Christopher and Gross, Kenneth W. and Duncan, Rory R. and Mullins, John J. (2017) Bimodal dynamics of granular organelles in primary renin-expressing cells revealed using TIRF microscopy. American Journal of Physiology - Renal Physiology, 312 (1). F200-F209. ISSN 1931-857X (https://doi.org/10.1152/ajprenal.00384.2016)

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Abstract

Renin is the initiator and rate-limiting factor in the renin-angiotensin blood pressure regulation system. Although renin is not exclusively produced in the kidney, in nonmurine species the synthesis and secretion of the active circulatory enzyme is confined almost exclusively to the dense core granules of juxtaglomerular (JG) cells, where prorenin is processed and stored for release via a regulated pathway. Despite its importance, the structural organization and regulation of granules within these cells is not well understood, in part due to the difficulty in culturing primary JG cells in vitro and the lack of appropriate cell lines. We have streamlined the isolation and culture of primary renin-expressing cells suitable for high-speed, high-resolution live imaging using a Percoll gradient-based procedure to purify cells from RenGFP+ transgenic mice. Fibronectin-coated glass coverslips proved optimal for the adhesion of renin-expressing cells and facilitated live cell imaging at the plasma membrane of primary renin cells using total internal reflection fluorescence microscopy (TIRFM). To obtain quantitative data on intracellular function, we stained mixed granule and lysosome populations with Lysotracker Red and stimulated cells using 100 nM isoproterenol. Analysis of membrane-proximal acidic granular organelle dynamics and behavior within renin-expressing cells revealed the existence of two populations of granular organelles with distinct functional responses following isoproterenol stimulation. The application of high-resolution techniques for imaging JG and other specialized kidney cells provides new opportunities for investigating renal cell biology.

ORCID iDs

Buckley, Charlotte ORCID logoORCID: https://orcid.org/0000-0002-7961-4544, Dun, Alison R., Peter, Audrey, Bellamy, Christopher, Gross, Kenneth W., Duncan, Rory R. and Mullins, John J.;
  • Item type:Article
    ID code:78549
    Dates:
    Date
    Event
    9 January 2017
    Published
    1 January 2017
    Published Online
    4 November 2016
    Accepted
    Subjects:Medicine
    Science > Microbiology
    Department:UNSPECIFIED
    Depositing user: Pure Administrator
    Date deposited:12 Nov 2021 11:28
    Last modified:11 Nov 2024 13:17
    Related URLs:
    URI:https://strathprints.strath.ac.uk/id/eprint/78549
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