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A MECHANISM FOR QUINOLINIC ACID INDUCED CYTOTOXICITY IN HUMAN PRIMARY ASTROCYTES

Nady Braidy 1 Gilles Guillemin 1,2 Ross Grant 1,3

1 School of Medical Sciences, Faculty of Medicine, University of New South Wales,
2 Centre for Immunology, St Vincent's Hospital,
3 Australasian Research Institute, Sydney Adventist Hospital.

Purpose

Quinolinic acid (QUIN) causes excitotoxicity in neurons through overactivation of the N-methyl D-Aspartate (NMDA) receptor. Recent evidence suggests that astrocytes are also susceptible to QUIN cytotoxicity by an unknown mechanism. This study investigated the putative role of NMDA receptor activation and oxidative stress in QUIN induced cytotoxicity in astrocytes.

Methods

Cultures of primary human foetal astrocytes and neurons were exposed to QUIN at concentrations between 0 and 1200nM for 24 hours. Intracellular NAD levels and poly(ADP-ribose) polymerase (PARP) activity were quantitated spectrophotometrically on culture homogenates. Cell death was assessed by quantitation of lactate dehydrogenase (LDH) activity in culture supernatants. All experiments were done in at least triplicate.

Results

Our results showed that QUIN increased intracellular concentrations of the essential pyridine nucleotide nicotinamide adenine dinucleotide (NAD+) at low, physiological, concentrations (<50 nM) in cultures of both neurons and astrocytes (n=4). However at concentrations above 150 nM, QUIN was shown to activate the NAD+ - dependent DNA repair enzyme PARP resulting in a dose dependant decrease in NAD+ and increased LDH activity in both astrocytes and neurons, (n=4).Treatment of astrocyte and neuron cultures, (n=3), with the NMDA ion channel blockers, MK-801 and memantine, and the nitric oxide (NO.) synthase inhibitor, N (G)-nitro-L- arginine methyl ester, L-NAME, prevented PARP activation and NAD+ depletion. Treatment with the competitive NMDA receptor antagonist AP-5 also ameliorated NAD+ depletion and PARP activation, but required much greater concentrations.

Conclusion

These results indicate for the first time that the astrocytes are susceptible to QUIN induced cytotoxicity via an apparent NMDA receptor mediated process, analogous to that previously identified for neurons.