Live Imaging of Organelle Motility in RPE Flatmounts

Ankita Umapathy; David S Williams

doi:10.1007/978-3-030-27378-1_64

Live Imaging of Organelle Motility in RPE Flatmounts

Adv Exp Med Biol. 2019:1185:389-393. doi: 10.1007/978-3-030-27378-1_64.

Authors

Ankita Umapathy¹, David S Williams^{2

3

4

5}

Affiliations

¹ Department of Ophthalmology and Stein Eye Institute, University of California, Los Angeles (UCLA),, Los Angeles, CA, USA.
² Department of Ophthalmology and Stein Eye Institute, University of California, Los Angeles (UCLA), Los Angeles, CA, USA. dswilliams@ucla.edu.
³ Department of Neurobiology, David Geffen School of Medicine, University of California, Los Angeles (UCLA), Los Angeles, CA, USA. dswilliams@ucla.edu.
⁴ Molecular Biology Institute, David Geffen School of Medicine, University of California, Los Angeles (UCLA), Los Angeles, CA, USA. dswilliams@ucla.edu.
⁵ Brain Research Institute, David Geffen School of Medicine, University of California, Los Angeles (UCLA), Los Angeles, CA, USA. dswilliams@ucla.edu.

PMID: 31884643
DOI: 10.1007/978-3-030-27378-1_64

Abstract

The retinal pigment epithelium (RPE) performs several functions that are crucial for normal retinal function and vision, including the daily phagocytosis of photoreceptor outer segment (POS) membranes. Defects in the motility and degradation of POS phagosomes may be associated with some inherited and age-related retinal degenerations. Given the apical to basal translocation of phagosomes during maturation and degradation, studies of the underlying mechanisms require analyses of the dynamics in 3-D. In this chapter, we report a method for investigating the 3-D motility of POS phagosomes and lysosomes, utilizing high-speed, spinning disk confocal microscopy of live RPE flatmounts.

Keywords: Flatmount; Live imaging; LysoTracker Red; Lysosome; Phagosome; RHO-EGFP mice; RPE; Spinning disk confocal microscopy.

MeSH terms

Humans
Lysosomes / physiology*
Microscopy, Confocal
Phagocytosis
Phagosomes / physiology*
Retinal Photoreceptor Cell Outer Segment / physiology*
Retinal Pigment Epithelium / cytology
Retinal Pigment Epithelium / diagnostic imaging*