Functional expression of particular isoforms of excitatory amino acid transporters by rodent cartilage.
In the present study, we have attempted to demonstrate functional expression by the rodent cartilage of particular isoforms of excitatory amino acid transporters (EAATs) essentially required for central glutamatergic signal termination. Constitutive expression of mRNA was shown for the first time with the neuronal EAAT subtype excitatory amino acid carrier-1 (EAAC1), in addition to glial subtypes such as glutamate aspartate transporter (GLAST) and glutamate transporter-1 (GLT-1), in rat costal chondrocytes cultured for 7-21 days on reverse transcription polymerase chain reaction (RT-PCR). Western blotting analysis confirmed the expression of corresponding proteins for both GLAST and GLT-1 in cultured chondrocytes. The accumulation of [(3)H]glutamate (Glu) occurred in a temperature- and sodium-dependent manner with biochemical and pharmacological profiles similar to those seen for brain EAATs in chondrocytes cultured for 7 days, while [(3)H]Glu accumulation consisted of a single component with a K(m) of 39.1+/-2.3 microM and a V(max) of 1320+/-120pmol/mg protein/min, respectively. In organotypic cultured metatarsals isolated before vascularization from embryonic mice, where cells underwent maturational development from resting to proliferating, prehypertrophic, hypertrophic and calcified chondrocytes in a progressive order of cellular differentiation, moreover, mRNA expression was seen for GLAST, GLT-1 and EAAT4 but not for EAAC1 subtypes. Immunohistochemical analysis revealed distribution profiles different from each other with GLAST, GLT-1 and EAAT4 isoforms in sections of cultured metatarsals and isolated tibiae. These results suggest that extracellular Glu could be cleared up into intracellular locations through particular glial and/or neuronal EAAT isoforms functionally expressed by the rodent cartilage.
Hinoi
Eiichi
E
Laboratory of Molecular Pharmacology, Division of Pharmaceutical Sciences, Kanazawa University Graduate School of Natural Science and Technology, Kanazawa, Ishikawa 920-1192, Japan.
Wang
Liyang
L
Takemori
Akihiro
A
Yoneda
Yukio
Y
D027322:Amino Acid Transport System X-AG / Q000235:genetics; D000818:Animals; D002352:Carrier Proteins / Q000235:genetics*; D002478:Cells, Cultured; D019902:Chondrocytes / Q000378:metabolism*; D027341:Excitatory Amino Acid Transporter 1 / Q000235:genetics; D027342:Excitatory Amino Acid Transporter 2 / Q000235:genetics; D050580:Excitatory Amino Acid Transporter 3; D050581:Excitatory Amino Acid Transporter 4; D005260:Female; D050491:Glutamate Plasma Membrane Transport Proteins; D018698:Glutamic Acid / Q000378:metabolism*; D051379:Mice; D020033:Protein Isoforms; D012333:RNA, Messenger / Q000032:analysis; D051381:Rats; D017208:Rats, Wistar; D027981:Symporters / Q000235:genetics
D027322:Amino Acid Transport System X-AG; D002352:Carrier Proteins; D027341:Excitatory Amino Acid Transporter 1; D027342:Excitatory Amino Acid Transporter 2; D050580:Excitatory Amino Acid Transporter 3; D050581:Excitatory Amino Acid Transporter 4; D050491:Glutamate Plasma Membrane Transport Proteins; D020033:Protein Isoforms; D012333:RNA, Messenger; C493399:Slc1a1 protein, mouse; C493398:Slc1a1 protein, rat; C493393:Slc1a3 protein, mouse; C493392:Slc1a3 protein, rat; C493390:Slc1a6 protein, mouse; C493389:Slc1a6 protein, rat; D027981:Symporters; D018698:Glutamic Acid