Functional and immunocytochemical characterization of D-serine transporters in cortical neuron and astrocyte cultures.
D-serine is an endogenous coagonist of N-methyl-D-aspartate (NMDA) receptors that plays an important role in synaptic function, neuronal development, and excitotoxicity. Mechanisms of D-serine transport are important in regulation of extracellular D-serine concentration and therefore of these critical processes. D-serine can be transported with low affinity through the Na(+)-dependent amino acid transporter termed ASCT2, whereas high-affinity D-serine uptake has been reported through the Na(+)-independent transporter termed asc-1. We investigated immunoreactivity for ASCT2 and asc-1 and D-serine transport kinetics in cultured cortical neurons and astrocytes to gain insight into how D-serine transporters regulate CNS D-serine levels. Both neurons and astrocytes exhibited low-affinity Na(+)-dependent D-serine uptake (K(T) > 1 mM) with broad substrate selectivity that was consistent with uptake through ASCT2. Both neurons and astrocytes also stained positively for ASCT2 in immunocytochemistry studies. Neurons but not astrocytes stained positively for the high-affinity D-serine transporter asc-1, but no evidence of functional asc-1 could be detected in neurons with conditions that produced such activity in cortical synaptosomes. These data support ASCT2 function in both neuron and astrocyte cultures and identify a discrepancy between observed asc-1 immunoreactivity and lack of functional asc-1 activity in neuron cultures. Together these findings further our knowledge of the processes that govern D-serine regulation.
Shao
Zongjun
Z
Department of Pharmacology and Therapeutics, University of Manitoba, and Division of Neurodegenerative Disorders, St. Boniface Hospital Research Center, Manitoba, Canada.
Kamboj
Amit
A
Anderson
Christopher M
CM
D026962:Amino Acid Transport System ASC / Q000378:metabolism*; D000704:Analysis of Variance; D000818:Animals; D001253:Astrocytes / Q000378:metabolism*; D015153:Blotting, Western; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000378:metabolism*; D007150:Immunohistochemistry; D007700:Kinetics; D008297:Male; D051379:Mice; D015778:Minor Histocompatibility Antigens; D009474:Neurons / Q000378:metabolism*; D012694:Serine / Q000378:metabolism*; D013574:Synaptosomes / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism*
D026962:Amino Acid Transport System ASC; D015778:Minor Histocompatibility Antigens; C498850:Slc1a5 protein, mouse; D014157:Transcription Factors; C459902:Trip4 protein, mouse; D012694:Serine