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Published on: May 3, 2017
Neurosteroids and glutamate toxicity in fibroblasts expressing human NMDA receptors
M Scott1, J J Tanguay, R J Beninger
1Department of Pharmacology and Toxicology, Queen's University, Kingston, Ont., Canada K7L 3N6. boegmanr@post.queensu.ca
Abstract:
We characterized glutamate receptor-mediated toxicity in mouse fibroblasts expressing the human NR1a/2A or NR1a/2B NMDA receptor. After induction of NMDA receptors, cells in both lines died over a 24 h time period. This toxicity was associated with a progressive increase in the glutamate content of the media. Cell death could be prevented by including either the non-competitive NMDA receptor antagonist ketamine or the competitive antagonist D,L-AP5. Cells expressing NR1a/2A receptors were maximally protected by 0.5 mM D,L -AP-5, while those expressing NR1a/2B receptors required 2 mM D,L -AP-5 for maximal protection. The neurosteroid pregnenolone sulfate, which negatively modulates NMDA receptor function, partially protected fibroblasts containing NR1a/2A or NR1a/2B NMDA receptor constructs. However, the neurosteroid pregnenolone sulfate, which has been reported to act as a positive allosteric modulator of the NMDA receptor, had no effect on the toxicity caused by endogenous glutamate. Our results on cells expressing human NMDA receptors suggest that certain neurosteroids may protect against NMDA induced toxicity while having low neurotoxic liabilities of their own.
Insights
NMDA receptor activation causes cell death, which can be blocked by antagonists. Neurosteroids partially protect cells from NMDA receptor toxicity, suggesting potential therapeutic benefits with minimal self-toxicity.
Area of Science:
- Neuroscience
- Molecular Biology
- Pharmacology
Background:
- Glutamate receptors, particularly NMDA receptors, play critical roles in neuronal function and excitotoxicity.
- Understanding NMDA receptor-mediated toxicity is crucial for developing therapeutic strategies against neurological disorders.
Purpose of the Study:
- To characterize glutamate receptor-mediated toxicity in mouse fibroblasts expressing human NMDA receptors (NR1a/2A and NR1a/2B).
- To investigate the protective effects of NMDA receptor antagonists and neurosteroids against this toxicity.
Main Methods:
- Utilized mouse fibroblast cell lines engineered to express specific human NMDA receptor subunits (NR1a/2A or NR1a/2B).
- Induced NMDA receptor activity and monitored cell viability over 24 hours.
- Assessed the effects of ketamine (non-competitive antagonist), D,L-AP5 (competitive antagonist), and pregnenolone sulfate on cell death and glutamate levels.
Main Results:
- NMDA receptor activation led to cell death, correlated with increased media glutamate levels.
- Ketamine and D,L-AP5 effectively prevented cell death, with varying concentrations required for maximal protection depending on the NMDA receptor subtype.
- Pregnenolone sulfate partially protected cells against NMDA receptor-induced toxicity, but did not affect toxicity from endogenous glutamate.
Conclusions:
- NMDA receptor antagonists are effective in preventing NMDA receptor-mediated excitotoxicity.
- Certain neurosteroids, like pregnenolone sulfate, may offer partial protection against NMDA receptor toxicity with a favorable safety profile.
