Serotonin regulation of serotonin uptake in RN46A cells

Nina Koldzic-Zivanovic, Patricia K. Seitz, Kathryn Cunningham, Mary L. Thomas, Thomas K. Hughes

Research output: Contribution to journalArticle

6 Citations (Scopus)

Abstract

1. Aim: The role of the serotonin transporter (SERT) is to remove serotonin (5-HT) from the synaptic space. In vitro studies have shown that 5-HT uptake via SERT is influenced by the availability of its substrate, 5-HT. We used RN46A cells, a line that expresses SERT, to investigate 5-HT regulation of 5-HT uptake and the intracellular signaling pathways involved. RN46A cells also express mRNAs for 5-HT receptors (5-HT1A, 5-HT1B, 5-HT2A, and 5-HT2C) and as cAMP and intracellular Ca 2+ are modulated by different 5-HT receptors, we studied both pathways. 2. Methods: 5-HT uptake was determined as imipramine-inhibitable uptake of [3H]5-HT, intracellular cAMP was measured by RIA and intracellular Ca2+ changes were determined using the ratiometric method of intracellular Ca2+ imaging. 3. Results: For uptake experiments, cells were kept for 30 min either with or without 1 μM 5-HT in the medium before measuring uptake. Removal of 5-HT for 30 min significantly decreased [3H]5-HT uptake. The absence of 5-HT for 15 min failed to induce any changes in intracellular cAMP levels. Removal of 5-HT from the medium did not change intracellular Ca2+ levels either; however, adding 1 μM 5-HT after 5 min in 5-HT-free conditions rapidly increased intracellular Ca2+ levels in 50% of the cells. The remaining cells showed no changes in the intracellular Ca2+ levels. 4. Conclusions: We have shown that in RN46A cells, that endogenously express SERT and mRNAs for several 5-HT receptors, changes in 5-HT levels influence 5-HT uptake rate as well as induce changes in intracellular Ca2+ levels. This suggests that 5-HT may utilize intracellular Ca2+ to regulate 5-HT uptake.

Original languageEnglish (US)
Pages (from-to)979-987
Number of pages9
JournalCellular and Molecular Neurobiology
Volume26
Issue number4-6
DOIs
StatePublished - Jul 2006

Fingerprint

Serotonin
Serotonin Plasma Membrane Transport Proteins
Serotonin Receptors
Messenger RNA
Imipramine

Keywords

  • Calcium
  • RN46A cells
  • Serotonin
  • Serotonin transporter

ASJC Scopus subject areas

  • Cellular and Molecular Neuroscience
  • Cell Biology
  • Neuroscience(all)
  • Clinical Biochemistry
  • Genetics

Cite this

Koldzic-Zivanovic, N., Seitz, P. K., Cunningham, K., Thomas, M. L., & Hughes, T. K. (2006). Serotonin regulation of serotonin uptake in RN46A cells. Cellular and Molecular Neurobiology, 26(4-6), 979-987. https://doi.org/10.1007/s10571-006-9097-x

Serotonin regulation of serotonin uptake in RN46A cells. / Koldzic-Zivanovic, Nina; Seitz, Patricia K.; Cunningham, Kathryn; Thomas, Mary L.; Hughes, Thomas K.

In: Cellular and Molecular Neurobiology, Vol. 26, No. 4-6, 07.2006, p. 979-987.

Research output: Contribution to journalArticle

Koldzic-Zivanovic, N, Seitz, PK, Cunningham, K, Thomas, ML & Hughes, TK 2006, 'Serotonin regulation of serotonin uptake in RN46A cells', Cellular and Molecular Neurobiology, vol. 26, no. 4-6, pp. 979-987. https://doi.org/10.1007/s10571-006-9097-x
Koldzic-Zivanovic, Nina ; Seitz, Patricia K. ; Cunningham, Kathryn ; Thomas, Mary L. ; Hughes, Thomas K. / Serotonin regulation of serotonin uptake in RN46A cells. In: Cellular and Molecular Neurobiology. 2006 ; Vol. 26, No. 4-6. pp. 979-987.
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AB - 1. Aim: The role of the serotonin transporter (SERT) is to remove serotonin (5-HT) from the synaptic space. In vitro studies have shown that 5-HT uptake via SERT is influenced by the availability of its substrate, 5-HT. We used RN46A cells, a line that expresses SERT, to investigate 5-HT regulation of 5-HT uptake and the intracellular signaling pathways involved. RN46A cells also express mRNAs for 5-HT receptors (5-HT1A, 5-HT1B, 5-HT2A, and 5-HT2C) and as cAMP and intracellular Ca 2+ are modulated by different 5-HT receptors, we studied both pathways. 2. Methods: 5-HT uptake was determined as imipramine-inhibitable uptake of [3H]5-HT, intracellular cAMP was measured by RIA and intracellular Ca2+ changes were determined using the ratiometric method of intracellular Ca2+ imaging. 3. Results: For uptake experiments, cells were kept for 30 min either with or without 1 μM 5-HT in the medium before measuring uptake. Removal of 5-HT for 30 min significantly decreased [3H]5-HT uptake. The absence of 5-HT for 15 min failed to induce any changes in intracellular cAMP levels. Removal of 5-HT from the medium did not change intracellular Ca2+ levels either; however, adding 1 μM 5-HT after 5 min in 5-HT-free conditions rapidly increased intracellular Ca2+ levels in 50% of the cells. The remaining cells showed no changes in the intracellular Ca2+ levels. 4. Conclusions: We have shown that in RN46A cells, that endogenously express SERT and mRNAs for several 5-HT receptors, changes in 5-HT levels influence 5-HT uptake rate as well as induce changes in intracellular Ca2+ levels. This suggests that 5-HT may utilize intracellular Ca2+ to regulate 5-HT uptake.

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