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. 1993 Sep;469:653–671. doi: 10.1113/jphysiol.1993.sp019836

Ca2+ oscillations and Ca2+ influx in Xenopus oocytes expressing a novel 5-hydroxytryptamine receptor.

A B Parekh 1, M Foguet 1, H Lübbert 1, W Stühmer 1
PMCID: PMC1143893  PMID: 8271222

Abstract

1. We expressed a novel 5-hydroxytryptamine receptor (SRL) in Xenopus oocytes and monitored cytosolic Ca2+ through the endogenous Ca(2+)-dependent Cl- channel activity using the double electrode voltage-clamp technique. 2. 5-Hydroxytryptamine (5-HT; 200 nM) led to an initial rapid oscillatory current followed by a pronounced secondary one, which lasted long after 5-HT wash-out (20-40 min) and was not affected by the receptor antagonist yohimbine. 3. Both phases of the current were abolished by heparin demonstrating a key role for IP3-induced Ca2+ release. 4. Caffeine (10 mM) alone did not evoke a current but reduced both phases of the current evoked by 5-HT. Ryanodine had no effect. No evidence for Ca(2+)-induced Ca2+ release was found. 5. The secondary current activated by 5-HT was sensitive to changes in extracellular Ca2+, suggesting it was evoked by Ca2+ influx. Reducing external Na+ did not affect this current, demonstrating that it was rather specific for Ca2+. 6. The Ca2+ influx pathway was much more sensitive to Cd2+ than other divalent ions (Co2+, Mn2+, Sr2+, Ba2+). It was insensitive to verapamil. 7. Injection of D-myo-inositol 1,4,5-trisphosphate, 3-deoxy-3-fluoro (IP3-F; an analogue not metabolized to D-myo-inositol 1,3,4,5-tetrakisphosphate (IP4)), evoked either an oscillatory current or a rapid current followed by a sustained secondary one. The latter was sensitive to external Ca2+ and was blocked by Cd2+. Heparin dramatically reduced the IP3-F-evoked current. 8. Perfusion in Ca(2+)-free solution, once a secondary current had been generated, significantly decreased the amount of intracellular Ca2+ mobilized by 5-HT, indicating that the Ca2+ influx pathway plays an important role in pool refilling. 9. Block of Ca2+ influx by Cd2+ in cells that were oscillating transiently increased the amplitude and then either abolished the oscillations or made them irregular. This effect was also elicited by increasing external Ca2+. 10. These results demonstrate that 5-HT, acting via IP3, both releases Ca2+ from internal stores and evokes a pronounced Ca2+ influx. This last step is activated by pool depletion and is important for both refilling of the agonist-sensitive stores and modifying the oscillatory pattern.

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Selected References

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