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| A ryanodine fluorescent derivative reveals the presence of high-affinity ryanodine binding sites in the Golgi complex of rat sympathetic neurons, with |
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Cifuentes, F; Gonzalez, CE; Fiordelisio, T; Guerrero, G; Lai, FA; Arturo Hernández Cruz; |
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Publication date:
2001 |
| Journal:
CELL SIGNAL |
| Volume:
13 |
| Issue:
5 |
| Pages:
353-362 |
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| The plant alkaloid ryanodine (Ry) is a high-affinity modulator of ryanodine receptor (RyR) Ca2+ release channels. Although these channels are present in a variety of cell types, their functional role in nerve cells is still puzzling. Here, a monosubstituted fluorescent Ry analogue, B-FL-X Ry, was used to reveal the distribution of RyRs in cultured rat sympathetic neurons. B-FL-X Ry competitively inhibited the binding of [H-3]Ry to rabbit skeletal muscle SR membranes, with an IC50, of 150 nM, compared to 7 nM of unlabeled Ry. Binding of B-FL-X Ry to the cytoplasm of sympathetic neurons is saturable, reversible and of high affinity. The pharmacology of B-FL-X Ry showed marked differences with unlabeled Ry, which are partially explained by its lower affinity: (1) use-dependent reversible inhibition of caffeine-induced intracellular Ca2+ release; (2) diminished voltage-gated Ca2+ influx, due to a positive shift in the activation of voltage gated Ca currents. B-FL-X Ry-stained sympathetic neurons, viewed under confocal microscopy. showed conspicuous labeling of crescent-shaped structures pertaining to the Golgi complex, a conclusion supported by experiments showing co-localization with Golgi-specific fluorescent probes and the breaking up of crescent-shaped staining after treatment with drugs that disassemble Golgi complex. The presence of RyRs to the Golgi could be confirmed with specific anti-RyR(2) antibodies, but evidence of caffeine-induced Ca2+ release from this organelle could not be obtained using fast confocal microscopy. Rather, an apparent decrease of the cytosolic Ca2+ signal was detected close to this organelle. In spite of that, short-term incubation with brefeldin A (BFA) suppressed the fast component of caffeine-induced Ca2+ release, and the Ca2+ release process lasted longer and appeared less organized. These observations, which suggest a possible role of the Golgi complex in Ca homeostasis and signaling in nerve cells, could be relevant to reports involving derangement of the Golgi complex as a probable cause of some forms of progressive neuronal degeneration, such as Alzheimer's disease and amyotrophic lateral sclerosis. (C) 2001 Elsevier Science Inc. All rights reserved. |
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| Keywords:
ryanodine Golgi complex Ca2+ release channels caffeine Ca2+ stores |
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| Times cited: 19 |
| Journal impact: 3.40 |
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