Glutamate transporter inhibitors influence osteoblast gene expression and activity

Brakspear, Karen Sarah, Parsons, P. and Mason, Deborah Jane ORCID: https://orcid.org/0000-0002-8666-6094 2009. Glutamate transporter inhibitors influence osteoblast gene expression and activity. International Journal of Experimental Pathology 90 (2) , A107-A108. 10.1111/j.1365-2613.2008.00644.x

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Abstract

Introduction Mechanical loading plays a key role in the physiology of bone, allowing functional adaption to its environment. A screen for genes associated with mechanical load-induced osteogenic signalling identified the glutamate transporter GLAST-1, implicating the excitatory amino acid glutamate in the mechanoresponse. Five high affinity Na+- dependant excitatory amino acid transporters (EAATs) terminate glutamatergic signalling. EAAT1 (GLAST-1) is expressed by osteocytes and bone-forming osteoblasts in vivo. Materials and Methods We inhibited glutamate transport in osteosarcoma cell lines and primary osteoblasts using small molecule inhibitors of EAATs 1-5 [L-trans-Pyrrolidine-2,4- dicarboxylic acid (t-PDC) and DL-threo-b-benzyloxyaspartic acid (TBOA)]. These inhibitors (0-1 mM) were toxic at high concentrations and had no effect on proliferation over 5 days, but did influence gene expression determined by quantitative RT-PCR and alkaline phosphatase activity. Results RT-PCR revealed mRNA expression of EAATs 1-3 and both splice variants of EAAT1 (EAAT1a and EAAT1exon9s kip) in MG-63 and SaOS-2 osteoblasts. To our knowledge, this is the first reported observation of expression of the EAAT1ex9skip variant in bone cells. In MG-63 osteoblasts, osteocalcin and osteonectin expression were significantly upregulated upon EAAT inhibition by t-PDC in the presence of glutamate and also by TBOA alone over 24 h; whereas no changes in expression were detected upon treatment with glutamate alone. In primary osteoblasts, long-term exposure to high levels of glutamate increased osteocalcin expression and down-regulated EAAT1. TBOA significantly upregulated alkaline phosphatase activity in SaOS-2 and primary osteoblasts. Discussion This data confirms previously published data that mechanically regulated glutamate transporters may be important in regulating bone homeostasis.

Item Type:	Article
Date Type:	Publication
Status:	Published
Schools:	Schools > Biosciences
Subjects:	Q Science > QH Natural history > QH426 Genetics Q Science > QR Microbiology
Publisher:	Wiley-Blackwell
ISSN:	0959-9673
Last Modified:	27 Oct 2022 10:08
URI:	https://orca.cardiff.ac.uk/id/eprint/69051

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