Modulation by calcineurin of 5-HT3 receptor function in NG108-15 neuroblastoma x glioma cells

HWGM Boddeke*, [No Value] Meigel, P Boeijinga, J Arbuckle, RJ Docherty

*Corresponding author for this work

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Abstract

1 We have investigated the mechanism of regulation of 5-HT3 receptor channel sensitivity in voltage-clamped (-80 mV) NG108-15 neuroblastoma cells.

2 The 5-HT-induced inward current activated rapidly. The fast onset was followed by a biphasic decay which was characterized by two time constants, tau(1) (1.1+/-0.21s) and tau(2) (8.9+/-1.6s), respectively. Brief applications of 5-HT, applied at 2 min intervals, induced a decrease in the amplitude of the 5-HT3 receptor-mediated peak inward currents.

3 Buffering of intracellular calcium with the calcium chelator BAPTA (10 mM) instead of EGTA (10 mM) attenuated the 5-HT-induced loss of responsiveness of 5-HT3 receptors. Omission of calcium from the extracellular medium yielded a similar attenuation of loss of responsiveness.

4 Inclusion of the protein kinase inhibitor, staurosporine (1 mu M) or of okadaic acid (1 mu M), an inhibitor of protein phosphatases 1 and 2A, in the intracellular buffer solution did not affect 5-HT3 receptor sensitivity.

5 Injection of cyclosporin A-cyclophilin A complex (20 nM), which potently inhibits calcineurin, did not affect the time constants of the biphasic decay of the 5-HT response tau(1) (1.4+/-0.28s) and tau(2) (11.3+/-1.7s). The complex, however, prevented the loss of 5-HT3 receptor responsiveness upon repeated application of 5-HT. A similar, but weaker effect was observed after intracellular application of the autoinhibitory peptide domain of calcineurin (1 mu M).

6 The recovery of desensitized 5-HT3 receptors upon a second application of 5-HT (1 mu M) showed a half-life time (tau 1/2 of 2.6+/-0.12 min in control cells which was reduced to 1.6+/-0.09 min in cells treated with cyclosporin A-cyclophilin A (20 nM) complex.

7 We conclude that calcineurin does not affect the fast decay of the 5-HT3 receptor response but may be involved in a slower process which regulates channel activity.

Original languageEnglish
Pages (from-to)1836-1840
Number of pages5
JournalBritish Journal of Pharmacology
Volume118
Issue number7
Publication statusPublished - Aug-1996

Keywords

  • 5-HT3 receptors
  • calcineurin
  • NG108-15 cells
  • receptor desensitization
  • DEPENDENT PROTEIN-KINASE
  • ROOT GANGLION NEURONS
  • DESENSITIZATION
  • CURRENTS
  • CHANNEL
  • PHOSPHORYLATION
  • PHOSPHATASE
  • ACID

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