Differential thapsigargin-sensitivities and interaction of Ca2+ stores in human SH-SY5Y neuroblastoma cells.

Abstract

In human SH-SY5Y neuroblastoma cells, two distinct intracellular Ca2+ stores, a KCl-/caffeine-sensitive and a carbachol-/IP3-sensitive store, were demonstrated previously. In this study, responses of these two intracellular Ca2+ stores to thapsigargin were characterized. Ca2+-release from these stores was evoked either by high K+ (100 mM KCl) or by 1 mM carbachol, and changes in the intracellular Ca2+ level were monitored using Fura-2 fluorimetry. A sequential stimulation protocol (KCl-->carbachol or vice versa) allowed evaluation of the individual contribution of different Ca2+ stores to the evoked intracellular Ca2+ ([Ca2+]i)-transients and the dynamic interaction between them. Thapsigargin (0.05 nM - 20 microM) alone induced a [Ca2+]i-transient. Both the carbachol- and the KCl-evoked [Ca2+]i-transients were inhibited by thapsigargin, but with very different sensitivities. Thapsigargin inhibited the carbachol-evoked [Ca2+]i-transients with (IC50 = 0.353 nM) or without (IC50 = 0.448 nM) a KCl-prestimulation, but an additional small component, with a much lower sensitivity (IC50=4814 nM), was observed in the absence of a KCl-prestimulation. In contrast, the KCl-evoked [Ca2+]i-transients displayed only one component with a very low sensitivity to thapsigargin in both absence (IC50=3343 nM) and presence (IC50=6858 nM) of a carbachol-prestimulation. These findings suggest that the sarco-/endoplasmic reticular Ca2+ ATPases associated with the KCl-/caffeine- and carbachol-/IP3-sensitive intracellular Ca2+ stores differ from each other, either in types or in their post-translational modification. Such difference might play important role in the regulation of neuronal Ca2+ homeostasis.

• Authors: Zayra V Garavito-Aguilar, Esperanza Recio-Pinto, Alexandra V Corrales, Jin Zhang, Thomas J J Blanck, Fang Xu
• Journal: Brain research (Brain Res) Vol. 1011 Issue 2 Pg. 177-86 (Jun 18 2004) ISSN: 0006-8993 [Print] Netherlands
• PMID: 15157804 (Publication Type: Comparative Study, Journal Article)
• Copyright: Copyright 2004 Elsevier B.V.

Chemical References

• Cholinergic Agonists
• Enzyme Inhibitors
• Muscarinic Antagonists
• Thapsigargin
• Atropine
• Carbachol
• Potassium
• Calcium
• Fura-2

Topics

• Atropine (pharmacology)
• Calcium (metabolism)
• Carbachol (pharmacology)
• Cell Line, Tumor
• Cholinergic Agonists (pharmacology)
• Dose-Response Relationship, Drug
• Drug Interactions
• Enzyme Inhibitors (pharmacology)
• Fura-2 (metabolism)
• Humans
• Inhibitory Concentration 50
• Intracellular Space (drug effects, metabolism)
• Muscarinic Antagonists (pharmacology)
• Neuroblastoma
• Potassium (pharmacology)
• Thapsigargin (pharmacology)
• Time Factors