NMDA glutamate receptors are expressed by osteoclast precursors and involved in the regulation of osteoclastogenesis

Blandine Merle, Cecile Itzstein, Pierre D Delmas, Chantal Chenu

Research output: Contribution to journalArticle

46 Citations (Scopus)

Abstract

We previously identified functional N-methyl-D-aspartate (NMDA) glutamate receptors in mature osteoclasts and demonstrated that they are involved in bone resorption in vitro. In the present work, we studied the expression of NMDA receptors (NMDAR) by osteoclast precursors and their role in osteoclastogenesis using two in vitro models, the murine myelomonocytic RAW 264.7 cell line and mouse bone marrow cells, both of which differentiate into osteoclasts in the presence of macrophage colony-stimulating factor (M-CSF) and Rank ligand (RankL). Using RT-PCR analysis with specific probes, we showed that RAW 264.7 cells and mouse bone marrow cells express mRNA of NMDAR subunits NMDA receptor 1 (NR1) and NMDA receptor 2 (NR2) A, B, and D. These subunits are expressed all along the differentiation sequence from undifferentiated precursors to mature resorbing osteoclasts. Semi-quantitative PCR analysis showed no regulation of the expression of these subunits during the differentiation process. Two specific non competitive antagonists of NMDAR, MK801 and DEP, dose-dependently inhibited osteoclast formation in both models, indicating that osteoclastogenesis requires the activation of NMDAR expressed by osteoclast precursors. MK801 had no effect when added only during the first 2 days of culture, suggesting that NMDAR are rather involved in the late stages of osteoclast formation. Finally, we demonstrated using Western-blotting and immunofluorescence that activation of NMDAR in RAW 264.7 cells by specific agonists induces nuclear translocation of NF-kappa B, a factor required for osteoclast formation. Altogether, our results indicate that osteoclast precursors express NMDAR that are involved in the osteoclast differentiation process through activation of the NF-kappa B pathway.
Original languageEnglish
Pages (from-to)424-36
Number of pages13
JournalJournal of Cellular Biochemistry
Volume90
Issue number2
DOIs
Publication statusPublished - 1 Oct 2003

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Glutamate Receptors
Osteoclasts
N-Methylaspartate
N-Methyl-D-Aspartate Receptors
Osteogenesis
Bone
NF-kappa B
Chemical activation
Cells
Bone Marrow Cells
N-methylglutamate
Polymerase Chain Reaction
Macrophage Colony-Stimulating Factor
Bone Resorption
Fluorescent Antibody Technique
Western Blotting
Ligands
Messenger RNA

Keywords

  • Animals
  • Bone Marrow Cells
  • Brain
  • Carrier Proteins
  • Cell Differentiation
  • Cell Nucleus
  • Dizocilpine Maleate
  • Excitatory Amino Acid Antagonists
  • Gene Expression Regulation
  • Macrophage Colony-Stimulating Factor
  • Male
  • Membrane Glycoproteins
  • Mice
  • Monocytes
  • Myeloid Cells
  • NF-kappa B
  • Osteoclasts
  • Protein Subunits
  • Protein Transport
  • Pyrrolidines
  • RANK Ligand
  • Receptor Activator of Nuclear Factor-kappa B
  • Receptors, N-Methyl-D-Aspartate
  • Reverse Transcriptase Polymerase Chain Reaction

Cite this

NMDA glutamate receptors are expressed by osteoclast precursors and involved in the regulation of osteoclastogenesis. / Merle, Blandine; Itzstein, Cecile; Delmas, Pierre D; Chenu, Chantal.

In: Journal of Cellular Biochemistry, Vol. 90, No. 2, 01.10.2003, p. 424-36.

Research output: Contribution to journalArticle

Merle, Blandine ; Itzstein, Cecile ; Delmas, Pierre D ; Chenu, Chantal. / NMDA glutamate receptors are expressed by osteoclast precursors and involved in the regulation of osteoclastogenesis. In: Journal of Cellular Biochemistry. 2003 ; Vol. 90, No. 2. pp. 424-36.
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T1 - NMDA glutamate receptors are expressed by osteoclast precursors and involved in the regulation of osteoclastogenesis

AU - Merle, Blandine

AU - Itzstein, Cecile

AU - Delmas, Pierre D

AU - Chenu, Chantal

N1 - Copyright 2003 Wiley-Liss, Inc.

PY - 2003/10/1

Y1 - 2003/10/1

N2 - We previously identified functional N-methyl-D-aspartate (NMDA) glutamate receptors in mature osteoclasts and demonstrated that they are involved in bone resorption in vitro. In the present work, we studied the expression of NMDA receptors (NMDAR) by osteoclast precursors and their role in osteoclastogenesis using two in vitro models, the murine myelomonocytic RAW 264.7 cell line and mouse bone marrow cells, both of which differentiate into osteoclasts in the presence of macrophage colony-stimulating factor (M-CSF) and Rank ligand (RankL). Using RT-PCR analysis with specific probes, we showed that RAW 264.7 cells and mouse bone marrow cells express mRNA of NMDAR subunits NMDA receptor 1 (NR1) and NMDA receptor 2 (NR2) A, B, and D. These subunits are expressed all along the differentiation sequence from undifferentiated precursors to mature resorbing osteoclasts. Semi-quantitative PCR analysis showed no regulation of the expression of these subunits during the differentiation process. Two specific non competitive antagonists of NMDAR, MK801 and DEP, dose-dependently inhibited osteoclast formation in both models, indicating that osteoclastogenesis requires the activation of NMDAR expressed by osteoclast precursors. MK801 had no effect when added only during the first 2 days of culture, suggesting that NMDAR are rather involved in the late stages of osteoclast formation. Finally, we demonstrated using Western-blotting and immunofluorescence that activation of NMDAR in RAW 264.7 cells by specific agonists induces nuclear translocation of NF-kappa B, a factor required for osteoclast formation. Altogether, our results indicate that osteoclast precursors express NMDAR that are involved in the osteoclast differentiation process through activation of the NF-kappa B pathway.

AB - We previously identified functional N-methyl-D-aspartate (NMDA) glutamate receptors in mature osteoclasts and demonstrated that they are involved in bone resorption in vitro. In the present work, we studied the expression of NMDA receptors (NMDAR) by osteoclast precursors and their role in osteoclastogenesis using two in vitro models, the murine myelomonocytic RAW 264.7 cell line and mouse bone marrow cells, both of which differentiate into osteoclasts in the presence of macrophage colony-stimulating factor (M-CSF) and Rank ligand (RankL). Using RT-PCR analysis with specific probes, we showed that RAW 264.7 cells and mouse bone marrow cells express mRNA of NMDAR subunits NMDA receptor 1 (NR1) and NMDA receptor 2 (NR2) A, B, and D. These subunits are expressed all along the differentiation sequence from undifferentiated precursors to mature resorbing osteoclasts. Semi-quantitative PCR analysis showed no regulation of the expression of these subunits during the differentiation process. Two specific non competitive antagonists of NMDAR, MK801 and DEP, dose-dependently inhibited osteoclast formation in both models, indicating that osteoclastogenesis requires the activation of NMDAR expressed by osteoclast precursors. MK801 had no effect when added only during the first 2 days of culture, suggesting that NMDAR are rather involved in the late stages of osteoclast formation. Finally, we demonstrated using Western-blotting and immunofluorescence that activation of NMDAR in RAW 264.7 cells by specific agonists induces nuclear translocation of NF-kappa B, a factor required for osteoclast formation. Altogether, our results indicate that osteoclast precursors express NMDAR that are involved in the osteoclast differentiation process through activation of the NF-kappa B pathway.

KW - Animals

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KW - Brain

KW - Carrier Proteins

KW - Cell Differentiation

KW - Cell Nucleus

KW - Dizocilpine Maleate

KW - Excitatory Amino Acid Antagonists

KW - Gene Expression Regulation

KW - Macrophage Colony-Stimulating Factor

KW - Male

KW - Membrane Glycoproteins

KW - Mice

KW - Monocytes

KW - Myeloid Cells

KW - NF-kappa B

KW - Osteoclasts

KW - Protein Subunits

KW - Protein Transport

KW - Pyrrolidines

KW - RANK Ligand

KW - Receptor Activator of Nuclear Factor-kappa B

KW - Receptors, N-Methyl-D-Aspartate

KW - Reverse Transcriptase Polymerase Chain Reaction

U2 - 10.1002/jcb.10625

DO - 10.1002/jcb.10625

M3 - Article

C2 - 14505357

VL - 90

SP - 424

EP - 436

JO - Journal of Cellular Biochemistry

JF - Journal of Cellular Biochemistry

SN - 0730-2312

IS - 2

ER -