3-Nitropropionic acid: an astrocyte-sparing neurotoxin in vitro.

3-Nitropropionic acid (NPA), an inhibitor of succinate dehydrogenase, is dietary neurotoxin. It is not known if neurons and astrocytes differ in their vulnerability to NPA, therefore, we investigated its toxicity in primary cultures of cerebellar granule cells and astrocytes. NPA inhibited succinate dehydrogenase and tricarboxylic acid cycle activity to the same degree in neurons and astrocytes. Even so NPA acid was 16 times more toxic to neurons than to astrocytes (LC50: 0.7 and 11 mM, respectively). The neurotoxicity of NPA was mediated by NMDA-receptor activation, calcium influx, and formation of reactive oxygen species, as revealed by the protective effect of NMDA-receptor blockade, the accumulation of 45Ca, and the protective effect of N-t-butyl-alpha-phenylnitron (PBN), a scavenger of reactive oxygen species. Cytotoxic concentrations of NPA caused a reduction in the intracellular level of glutathione, which probably contributed to the oxidative damage in both neurons and astrocytes. The relative resistance of astrocytes to NPA appeared to be related to their low tricarboxylic acid cycle activity (5%-10% of that in neurons) and to the inability of NPA to cause astrocytic calcium overload. We conclude that NPA acid predominantly is an astrocyte-sparing neurotoxin.

The osmotic fragility of red blood cells: a re-evaluation of technical conditions.

The osmotic fragility of red blood cells is influenced by even modest environmental changes. Consequently, the technical procedure must be strictly standardized. This implies that temperature equilibrium of the buffered salt solutions should be reached prior to the addition of blood. Furthermore, since erroneous statements concerning the composition of phosphate buffers regularly used to secure correct pH of the salt solutions have repeatedly appeared in the literature, pH control of such solutions prior to use becomes essential.

Neurotoxic effect of L-2-chloropropionic acid on primary cultures of rat cerebellar granule cells.

L-2-Chloropropionic acid (L-CPA), when administered orally to rats, produces selective necrosis to the granule cell layer of the rat cerebellum which is delayed in onset, not appearing until 36-48 h after exposure. The present study was conducted to characterise the toxic effect of L-CPA in primary cell cultures of rat cerebellar granule cells in vitro. Exposure to L-CPA produced a time and concentration dependent loss in cerebellar granule cell viability. Mean 50% effective concentration (EC50) values for L-CPA toxicity were 18.3 +/- 0.3, 7.4 +/- 0.1, and 3.5 +/- 0.1 mM for 24, 48 and 72 h exposure respectively. Exposure for 24 h followed by a return to L-CPA free medium for 24 h was more toxic than exposure for 24 h alone. Cells maintained in culture for a longer duration were more susceptible to L-CPA-induced toxicity. The toxic effects of L-CPA could be partially or fully prevented by concomitant exposure of the cells to putative neuroprotective compounds. The N-methyl-D-aspartate (NMDA) receptor antagonist, MK-801 (3 microM), afforded partial protection against L-CPA induced toxicity, whilst other glutamate receptor antagonists including, D(-)-2-amino-5-phosphopentanoic acid (D-AP5; 300 microM), D(-)-3-(2-carboxypiperazine-4-yl)-propyl-1-phosphonic acid (D-CPP; 300 microM), 5,7-dichlorokynurenic acid (10 microM) and 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX; 1 microM) were ineffective. The antioxidant, vitamin E (10 microM), afforded significant but incomplete protection from L-CPA toxicity. However when both MK-801 (3 microM) and vitamin E (10 microM) were present during L-CPA exposure, a greater degree of protection was observed than with either compound alone, although the combination failed to provide complete protection. Cyclosporin A, an inhibitor of the mitochondrial transition pore, also provided partial protection. By contrast, the free radical trapping agent, N-tert-butyl-alpha-(2-sulfophenyl)-nitrone (S-PBN) provided concentration (1-10 mM) dependent protection against the L-CPA-induced toxicity, which was complete at 10 mM. Our findings suggest that free radical production may be involved in the mechanism of L-CPA-induced toxicity.