Elsevier

Neuroscience

Volume 136, Issue 1, 2005, Pages 259-267
Neuroscience

Neuropharmacology
Corticosterone and dexamethasone potentiate cytotoxicity associated with oxygen-glucose deprivation in organotypic cerebellar slice cultures

https://doi.org/10.1016/j.neuroscience.2005.07.043Get rights and content

Abstract

Many patients display elevated levels of serum cortisol following acute ischemic stroke. Given that glucocorticoids may potentiate some forms of insult, these studies examined the effects of corticosterone or dexamethasone exposure on cytotoxicity following oxygen-glucose deprivation in the cerebellum, a brain region susceptible to stroke. In organotypic cerebellar slice cultures prepared from neonatal rat pups, 90-min of oxygen-glucose deprivation at 15 days in vitro resulted in significant cytotoxicity at 24-, 48-, and 72-h post-oxygen-glucose deprivation, as measured by uptake of propidium iodide. Exposure of cultures following oxygen-glucose deprivation to the antioxidant trolox (500μM), but not to the glucocorticoid receptor antagonist RU486 (10μM), completely blocked oxygen-glucose deprivation-induced cytotoxicity. Corticosterone (1μM) or dexamethasone (10μM) exposure alone did not significantly increase propidium iodide uptake above levels observed in control cultures. However, corticosterone or dexamethasone exposure after oxygen-glucose deprivation potentiated oxygen-glucose deprivation-mediated propidium iodide uptake at each time point. Trolox, as well as RU486, co-exposure of cultures to corticosterone or dexamethasone after oxygen-glucose deprivation abolished all cytotoxicity. In conclusion, these data demonstrated that glucocorticoid exposure modulated oxygen-glucose deprivation-mediated propidium iodide uptake, which likely involved glucocorticoid receptor activation and pro-oxidant effects.

Section snippets

Organotypic cerebellar slice culture preparation

Preparation of cerebellar cultures followed procedures described by Stoppini et al. (1991) with modifications as detailed below. Cerebella from 8-day old male and female Sprague–Dawley rat pups (Harlan, Indianapolis, IN, USA) were aseptically removed and placed into ice-cold dissecting medium (Minimum Essential Medium with 2mM l-glutamine plus 25mM HEPES and 50μM penicillin/streptomycin solutions). Using a McIllwain tissue chopper (Mickle Laboratory Engineering Co. Ltd., Gomshall, UK), each

OGD

Initial studies examined the effects of 90-min exposure to OGD on propidium iodide uptake. Exposure to OGD resulted in significant elevations in uptake of propidium iodide when compared with control cultures at 24-, 48-, and 72-h post-insult [F(1,162)=16.084, P<0.001, post hoc P<0.05; Fig. 1]. Representative images are presented as a part of Fig. 3. In addition, analysis failed to demonstrate a significant main effect for time, though propidium iodide uptake was reduced with each subsequent

Discussion

This report has demonstrated that 90-min exposure to OGD in organotypic cerebellar slice cultures resulted in significant cytotoxicity, at 24-, 48-, and 72-h post-insult. Propidium iodide fluorescence in OGD-treated cultures was lower at 72-h when compared with levels observed 24-h post-insult and varied between experiment. The reason for this is unknown; however, it may be related to microglia activation or exclusion of propidium iodide. Exposure to trolox, a potent antioxidant, completely

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    Present address: Medical University of South Carolina, Center for Drug and Alcohol Programs IOP4N, 67 President Street, Charleston, SC 29425, USA.

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