Effects of the neuroprotectant lubeluzole on the cytotoxic actions of veratridine,barium, ouabain and 6-hydroxydopamine in chromaffin cells |
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Authors: | María F Cano-Abad Manuela G López Jesús M Hernández-Guijo Pedro Zapater Luis Gandía Pedro Sánchez-García Antonio G García |
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Affiliation: | 1.Departamento de Farmacología, Facultad de Medicina, Universidad Autónoma de Madrid, Arzobispo Morcillo 4; 28029 Madrid, Spain;2.Servicio de Farmacología Clínica e Instituto de Gerontología, Hospital de la Princesa, Diego de León, 62; 28006 Madrid, Spain |
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Abstract: | - Incubation of bovine adrenal chromaffin cells with veratridine (10–100 μM) during 24 h, caused a concentration-dependent release of the cytosolic lactate dehydrogenase (LDH) into the bathing medium, an indicator of cell death. Lubeluzole or its R(−) enantiomer, {"type":"entrez-nucleotide","attrs":{"text":"R91154","term_id":"958694","term_text":"R91154"}}R91154, did not enhance LDH release. Both lubeluzole and {"type":"entrez-nucleotide","attrs":{"text":"R91154","term_id":"958694","term_text":"R91154"}}R91154 (0.3–10 μM) decreased the veratridine-induced LDH release.
- Penfluridol did not increase LDH release at concentrations 0.003–1 μM; 3–10 μM increased LDH release to 50–60%, after 24 h exposure. Penfluridol (0.03–0.3 μM) did not protect against the cytotoxic effects of veratridine; at 1 μM, 15% protection was produced. Higher concentrations (3–10 μM) enhanced the cytotoxic effects of veratridine.
- Ba2+ ions caused a concentration-dependent increase of LDH release. This cytotoxic effect was partially prevented by 3 μM lubeluzole and fully counteracted by 1 μM penfluridol. {"type":"entrez-nucleotide","attrs":{"text":"R91154","term_id":"958694","term_text":"R91154"}}R91154 was less potent than lubeluzole and only protected against the lesion induced by 0.5 mM Ba2+.
- Ouabain (10 μM during 24 h) increased LDH release to about 30%. Both lubeluzole (0.3–10 μM) and the lower concentrations of penfluridol (0.003–0.3 μM) prevented the ouabain cytotoxic effects. At higher concentrations (3 μM), penfluridol increased drastically the ouabain cytotoxic effects.
- 6-Hydroxydopamine (6-OHDA) caused significant cytotoxic effects at 30 and 100 μM. Lubeluzole (3–10 μM) or penfluridol (0.03–0.3 μM) had no cytoprotective effects against 6-OHDA.
- Lubeluzole (3 μM), {"type":"entrez-nucleotide","attrs":{"text":"R91154","term_id":"958694","term_text":"R91154"}}R91154 (3 μM) and penfluridol (1 μM) blocked the current through Na+ channels in voltage-clamped chromaffin cells (INa) by around 20–30%. Ca2+ current through Ca2+ channels (ICa) was inhibited 57% by lubeluzole and {"type":"entrez-nucleotide","attrs":{"text":"R91154","term_id":"958694","term_text":"R91154"}}R91154 and 50% by penfluridol. The effects of penfluridol were not washed out, but those of lubeluzole and {"type":"entrez-nucleotide","attrs":{"text":"R91154","term_id":"958694","term_text":"R91154"}}R91154 were readily reversible.
- Lubeluzole (3 μM) induced reversible blockade of the oscillations of the cytosolic Ca2+, [Ca2+]i, in fura-2-loaded cells exposed to 30 or 100 μM veratridine. Penfluridol (1 μM) inhibited those oscillations in an irreversible manner.
- The results suggest that lubeluzole and its R-isomer caused cytoprotection against veratridine cell damage, by blocking the veratridine stimulated Na+ and Ca2+ entry, as well as the [Ca2+]i oscillations. The Ba2+ and ouabain cytotoxic effects were prevented more efficiently by penfluridol, likely by blocking the plasmalemmal Na+/Ca2+ exchanger. It remains dubious whether these findings are relevant to the reported neuroprotective action of lubeluzole in stroke; the doubt rests in the stereoselective protecting effects of lubeluzole in in vivo stroke models, as opposed to its lack of stereoselectivity in the in vitro model reported here.
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Keywords: | Cytotoxicity, cytoprotection, lubeluzole, __tag_381091753" class=" tag_hotlink" href=" /nuccore/R91154" ref=" /nuccore/R91154" >{" type" :" entrez-nucleotide" ," attrs" :{" text" :" R91154" ," term_id" :" 958694" ," term_text" :" R91154" }}R91154, penfluridol, veratridine, chromaffin cell, 6-hydroxydopamine, ouabain |
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