NDLI: Role of membrane cholesterol in spontaneous exocytosis at frog neuromuscular synapses: reactive oxygen species

Content Provider	PubMed Central
Author	Petrov, Alexey M. Yakovleva, Anastasiya A. Zefirov, Andrey L.
Copyright Year	2014
Abstract	Using electrophysiological and optical techniques, we studied the mechanisms by which cholesterol depletion stimulates spontaneous transmitter release by exocytosis at the frog neuromuscular junction. We found that methyl-β-cyclodextrin (MCD, 10 mm)-mediated exhaustion of cholesterol resulted in the enhancement of reactive oxygen species (ROS) production, which was prevented by the antioxidant N-acetyl cysteine (NAC) and the NADPH oxidase inhibitor apocynin. An increase in ROS levels occurred both extra- and intracellularly, and it was associated with lipid peroxidation in synaptic regions. Cholesterol depletion provoked a rise in the intracellular Ca2+ concentration, which was diminished by NAC and transient receptor potential vanilloid (TRPV) channel blockers (ruthenium red and capsazepine). By contrast, the MCD-induced rise in [Ca2+]i remained unaffected if Ca2+ release from endoplasmic stores was blocked by TMB8 (8-(diethylamino)octyl-3,4,5-trimethoxybenzoate hydrochloride). The effects of cholesterol depletion on spontaneous release and exocytosis were significantly reduced by the antioxidant, intracellular Ca2+ chelation with BAPTA-AM and blockers of TRPV channels. Bath application of the calcineurin antagonist cyclosporine A blocked MCD-induced enhancement of spontaneous release/exocytosis, whereas okadaic acid, an inhibitor of phosphatases PP1 and PP2A, had no effect. Thus, our findings indicate that enhancement of spontaneous exocytosis induced by cholesterol depletion may depend on ROS generation, leading to an influx of Ca2+ via TRPV channels and, subsequently, activation of calcineurin.
Related Links	http://dx.doi.org/10.1113/jphysiol.2014.279695
Ending Page	5009
Page Count	15
Starting Page	4995
File Format	PDF
ISSN	14697793
e-ISSN	14697793
Journal	The Journal of Physiology
Issue Number	Pt 22
Volume Number	592
Language	English
Publisher	BlackWell Publishing Ltd
Publisher Date	2014-11-15
Access Restriction	Open
Rights Holder	BlackWell Publishing Ltd
Subject Keyword	Research in Higher Education
Content Type	Text
Resource Type	Article
Subject	Physiology Sports Science

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