title: Functional characterization of N-methyl-D-aspartic acid-gated channels in bone cells.
abstract: Our recent identification of glutamate receptors in bone cells suggested a novel means of paracrine communication in the skeleton. To determine whether these receptors are functional, we investigated the effects of the excitatory amino acid, glutamate, and the pharmacological ligand, N-methyl-D-aspartic acid (NMDA), on glutamate-like receptors in the human osteoblastic cell lines MG63 and SaOS-2. Glutamate binds to osteoblasts, with a Kd of approximately 10(-4) mol/L and the NMDA receptor antagonist, D(L)-2-amino-5-phosphonovaleric acid (D-APV), inhibits binding. Using the patch-clamp technique, we measured whole-cell currents before and after addition of L-glutamate or NMDA and investigated the effects of the NMDA channel blockers, dizolcipine maleate (MK801), and Mg2+, and the competitive NMDA receptor antagonist, 3-((R)-2-carboxypiperazin-4-yl)-propyl-1-phosphoric acid (R-CPP), on agonist-induced currents. Both glutamate and NMDA induced significant increases in membrane currents. Application of Mg2+ (200 micromol/L) and MK801 (100 micromol/L) caused a significant decrease in inward currents elicited in response to agonist stimulation. The competitive NMDA receptor antagonist, R-CPP (100 micromol/L), also partially blocked the NMDA-induced currents in MG63 cells. This effect was reversed by addition of further NMDA (100 micromol/L). In Fura-2-loaded osteoblasts, glutamate induced elevation of intracellular free calcium, which was blocked by MK801. These results support the hypothesis that glutamate plays a role in bone cell signaling and suggest a possible role for glutamate agonists/antagonists in the treatment of bone diseases.
authors.affiliation: Department of Biology, University of York, UK.
authors.forename: I
authors.initials: I
authors.lastname: Laketić-Ljubojević
authors.forename: L J
authors.initials: LJ
authors.lastname: Suva
authors.forename: F J
authors.initials: FJ
authors.lastname: Maathuis
authors.forename: D
authors.initials: D
authors.lastname: Sanders
authors.forename: T M
authors.initials: TM
authors.lastname: Skerry
mesh_terms: D001665:Binding Sites; D001842:Bone and Bones / Q000166:cytology / Q000378:metabolism / Q000502:physiology*; D002118:Calcium / Q000378:metabolism; D002478:Cells, Cultured; D018691:Excitatory Amino Acid Antagonists / Q000494:pharmacology; D018698:Glutamic Acid / Q000378:metabolism; D006801:Humans; D007424:Intracellular Fluid / Q000378:metabolism; D015640:Ion Channel Gating / Q000502:physiology*; D007473:Ion Channels / Q000378:metabolism / Q000502:physiology*; D007700:Kinetics; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D016202:N-Methylaspartate / Q000378:metabolism* / Q000494:pharmacology; D010006:Osteoblasts / Q000378:metabolism / Q000502:physiology; D018408:Patch-Clamp Techniques; D016194:Receptors, N-Methyl-D-Aspartate / Q000819:agonists / Q000037:antagonists & inhibitors / Q000378:metabolism / Q000502:physiology*
chemical_list: D018691:Excitatory Amino Acid Antagonists; D007473:Ion Channels; D016194:Receptors, N-Methyl-D-Aspartate; D018698:Glutamic Acid; D016202:N-Methylaspartate; D002118:Calcium