Serotonin transporter and receptor expression in osteocytic MLO-Y4 cells

M. Bliziotes, A. Eshleman, B. Burt-Pichat, X. W. Zhang, J. Hashimoto, Kristine Wiren, C. Chenu

Research output: Contribution to journalArticle

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Abstract

Neurotransmitter regulation of bone metabolism has been a subject of increasing interest and investigation. We reported previously that osteoblastic cells express a functional serotonin (5-HT) signal transduction system, with mechanisms for responding to and regulating uptake of 5-HT. The clonal murine osteocytic cell line, MLO-Y4, demonstrates expression of the serotonin transporter (5-HTT), and the 5-HT1A, and 5-HT2A receptors by real-time RT-PCR and immunoblot analysis. Immunohistochemistry using antibodies for the 5-HTT, and the 5-HT1A and 5-HT2A receptors reveals expression of all three proteins in both osteoblasts and osteocytes in rat tibia. 5-HTT binding sites were demonstrated in the MLO-Y4 cells with nanomolar affinity for the stable cocaine analog [125I]RTI-55. Imipramine and fluoxetine, antagonists with specificity for 5-HTT, show the highest potency to antagonize [125I]RTI-55 binding in the MLO-Y4 cells. GBR-12935, a relatively selective dopamine transporter antagonist, had a much lower potency, as did desipramine, a selective norepinephrine transporter antagonist. The maximal [3H]5-HT uptake rate in MLO-Y4 cells was 2.85 pmol/15 min/well, with a Km value of 290 nM. Imipramine and fluoxetine inhibited specific [3H]5-HT uptake with IC50 values in the nanomolar range. 5-HT rapidly stimulated PGE2 release from MLO-Y4 cells; the EC50 for 5-HT was 0.1 μM, with a 3-fold increase seen at 60 min. The rate-limiting enzyme for serotonin synthesis, tryptophan hydroxylase, is expressed in MLO-Y4 cells as well as osteoblastic MC3T3-E1 cells. Thus, osteocytes, as well as osteoblasts, are capable of 5-HT synthesis, and express functional receptor and transporter components of the 5-HT signal transduction system.

Original languageEnglish (US)
Pages (from-to)1313-1321
Number of pages9
JournalBone
Volume39
Issue number6
DOIs
StatePublished - Dec 2006

Fingerprint

Serotonin Plasma Membrane Transport Proteins
Serotonin Receptors
Serotonin
Receptor, Serotonin, 5-HT2A
Osteocytes
Imipramine
Fluoxetine
Osteoblasts
Signal Transduction
Norepinephrine Plasma Membrane Transport Proteins
Tryptophan Hydroxylase
Dopamine Plasma Membrane Transport Proteins
Desipramine
Dopamine Antagonists
Tibia
Cocaine
Dinoprostone
Inhibitory Concentration 50
Neurotransmitter Agents
Real-Time Polymerase Chain Reaction

Keywords

  • Bone
  • Neurotransmitter
  • Osteoblast
  • Osteocyte
  • Transporter

ASJC Scopus subject areas

  • Physiology
  • Hematology

Cite this

Bliziotes, M., Eshleman, A., Burt-Pichat, B., Zhang, X. W., Hashimoto, J., Wiren, K., & Chenu, C. (2006). Serotonin transporter and receptor expression in osteocytic MLO-Y4 cells. Bone, 39(6), 1313-1321. https://doi.org/10.1016/j.bone.2006.06.009

Serotonin transporter and receptor expression in osteocytic MLO-Y4 cells. / Bliziotes, M.; Eshleman, A.; Burt-Pichat, B.; Zhang, X. W.; Hashimoto, J.; Wiren, Kristine; Chenu, C.

In: Bone, Vol. 39, No. 6, 12.2006, p. 1313-1321.

Research output: Contribution to journalArticle

Bliziotes, M, Eshleman, A, Burt-Pichat, B, Zhang, XW, Hashimoto, J, Wiren, K & Chenu, C 2006, 'Serotonin transporter and receptor expression in osteocytic MLO-Y4 cells', Bone, vol. 39, no. 6, pp. 1313-1321. https://doi.org/10.1016/j.bone.2006.06.009
Bliziotes M, Eshleman A, Burt-Pichat B, Zhang XW, Hashimoto J, Wiren K et al. Serotonin transporter and receptor expression in osteocytic MLO-Y4 cells. Bone. 2006 Dec;39(6):1313-1321. https://doi.org/10.1016/j.bone.2006.06.009
Bliziotes, M. ; Eshleman, A. ; Burt-Pichat, B. ; Zhang, X. W. ; Hashimoto, J. ; Wiren, Kristine ; Chenu, C. / Serotonin transporter and receptor expression in osteocytic MLO-Y4 cells. In: Bone. 2006 ; Vol. 39, No. 6. pp. 1313-1321.
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AU - Wiren, Kristine

AU - Chenu, C.

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AB - Neurotransmitter regulation of bone metabolism has been a subject of increasing interest and investigation. We reported previously that osteoblastic cells express a functional serotonin (5-HT) signal transduction system, with mechanisms for responding to and regulating uptake of 5-HT. The clonal murine osteocytic cell line, MLO-Y4, demonstrates expression of the serotonin transporter (5-HTT), and the 5-HT1A, and 5-HT2A receptors by real-time RT-PCR and immunoblot analysis. Immunohistochemistry using antibodies for the 5-HTT, and the 5-HT1A and 5-HT2A receptors reveals expression of all three proteins in both osteoblasts and osteocytes in rat tibia. 5-HTT binding sites were demonstrated in the MLO-Y4 cells with nanomolar affinity for the stable cocaine analog [125I]RTI-55. Imipramine and fluoxetine, antagonists with specificity for 5-HTT, show the highest potency to antagonize [125I]RTI-55 binding in the MLO-Y4 cells. GBR-12935, a relatively selective dopamine transporter antagonist, had a much lower potency, as did desipramine, a selective norepinephrine transporter antagonist. The maximal [3H]5-HT uptake rate in MLO-Y4 cells was 2.85 pmol/15 min/well, with a Km value of 290 nM. Imipramine and fluoxetine inhibited specific [3H]5-HT uptake with IC50 values in the nanomolar range. 5-HT rapidly stimulated PGE2 release from MLO-Y4 cells; the EC50 for 5-HT was 0.1 μM, with a 3-fold increase seen at 60 min. The rate-limiting enzyme for serotonin synthesis, tryptophan hydroxylase, is expressed in MLO-Y4 cells as well as osteoblastic MC3T3-E1 cells. Thus, osteocytes, as well as osteoblasts, are capable of 5-HT synthesis, and express functional receptor and transporter components of the 5-HT signal transduction system.

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