Long-term, repeated dose in vitro neurotoxicity of the glutamate receptor antagonist L-AP3, demonstrated in rat hippocampal slice cultures by using continuous propidium iodide incubation

Bjarne W Kristensen, Morten Blaabjerg, Jens Noraberg, Jens Zimmer

Publikation: Bidrag til tidsskriftTidsskriftartikelForskningpeer review

3 Citationer (Scopus)

Abstract

Most in vitro models are only used to assess short-term effects of test compounds. However, as demonstrated here, hippocampal slice cultures can be used for long-term studies. The test compound used was the metabotropic glutamate receptor antagonist, L(+)-2-amino-3-phosphonopropionic acid (L-AP3), which is known to be toxic in vivo after subchronic, but not acute, administration. Degenerative effects were monitored by measuring the cellular uptake of propidium iodide (PI; continuously present in the medium) and lactate dehydrogenase (LDH) leakage, and by using a panel of histological stains. Hippocampal slices, derived from 2-3 day old rats and grown for 3 weeks, were subsequently exposed for the next 3 weeks to 0, 10 or 100microM L-AP3, with PI (2microM) in the culture medium. Exposure to 100microM L-AP3 induced severe toxicity after 4-6 days, shown by massive PI uptake, LDH leakage, changes in MAP2 and GFAP immunostaining, and in Nissl and Timm staining. In contrast, 10microM L-AP3 did not induce detectable neuronal degeneration. Treatment with the NMDA receptor antagonist, MK-801, or the AMPA/KA receptor antagonist NBQX, together with 100microM L-AP3, reduced neurodegeneration down to close to control values. It is concluded that continuous incubation of hippocampal slice cultures with PI is technically feasible for use in studies of inducible neuronal degeneration over time.

OriginalsprogEngelsk
TidsskriftATLA Alternatives to Laboratory Animals
Vol/bind35
Udgave nummer2
Sider (fra-til)211-21
Antal sider11
ISSN0261-1929
DOI
StatusUdgivet - maj 2007
Udgivet eksterntJa

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