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Fluorescently-Labeled Estradiol Internalization and Membrane Trafficking in Live N-38 Neuronal Cells Visualized with Total Internal Reflection Fluorescence Microscopy

Citation

Kisler, K and Chow, RH and Dominguez, R, Fluorescently-Labeled Estradiol Internalization and Membrane Trafficking in Live N-38 Neuronal Cells Visualized with Total Internal Reflection Fluorescence Microscopy, Journal of steroids & hormonal science, Suppl 12 pp. 1-15. ISSN 2157-7536 (2013) [Refereed Article]


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DOI: doi:10.4172/2157-7536.S12-002

Abstract

Estradiol is a steroid hormone that binds and activates estradiol receptors. Activation of these receptors is known to modulate neuronal physiology and provide neuroprotection, but it is not completely understood how estradiol mediates these actions on the nervous system. Activation of a sub-population of estradiol receptor-α (ERα), originally identified as a nuclear protein, localizes to the plasma membrane and appears to be a critical step in neuroprotection against brain injury and disease. Previously we showed that estradiol stimulates the rapid and transient trafficking of plasma membrane ERα in primary hypothalamic neurons, and internalization of membraneimpermeant estradiol (E6BSA-FITC) into cortical neuron endosomes in vitro. These findings support the concept that estradiol activates and down-regulates plasma membrane ERα by triggering endocytosis. Here, we use TIRFM (total internal reflection fluorescence microscopy) to image the trafficking of E6BSA-FITC, and GFP-labeled ERα, in live cells in real time. We show that activation of plasma membrane ERs by E6BSA-FITC result in internalization of the fluorescent ligand in live N-38 neurons, an immortalized hypothalamic cell line. Pretreatment with ER antagonist ICI 182,780 decreased the number of E6BSA-FITC labeled puncta observed. We also observed in live N-38 neurons that E6BSA-FITC co-localized with FM4-64 and LysoTracker fluorescent dyes that label endosomes and lysosomes. Our results provide further evidence that plasma membrane ERα activation results in endocytosis of the receptor.

Item Details

Item Type:Refereed Article
Keywords:Live cell imaging, TIRF, trafficking, estrogen receptor
Research Division:Biological Sciences
Research Group:Zoology
Research Field:Animal Cell and Molecular Biology
Objective Division:Expanding Knowledge
Objective Group:Expanding Knowledge
Objective Field:Expanding Knowledge in the Biological Sciences
Author:Dominguez, R (Dr Reymundo Dominguez)
ID Code:122240
Year Published:2013
Deposited By:Medicine (Discipline)
Deposited On:2017-11-06
Last Modified:2017-11-10
Downloads:0

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