D-Serine, a co-agonist at the NMDA receptor (NMDAR), is synthesized from l-serine by the enzyme serine racemase (SR), which is heavily expressed in the forebrain. Although SR was originally reported to be localized exclusively to astrocytes, recent conditional knock out results demonstrate that little SR is expressed in forebrain astrocytes. As a consequence, the cellular location of its product, D-serine, in the brain is also uncertain. Immunocytochemistry now indicates that SR is expressed primarily in forebrain glutamatergic neurons with the remainder in GABAergic interneurons. We utilized SR deficient (SR-/-) mice, which have <15 % of normal D-serine levels, to validate and optimize a D-serine immunohistochemical method. Nearly all of the D-serine in neocortex and hippocampus (HP) is found in neurons, with virtually no D-serine co-localizing with two astrocyte markers. Interestingly, only a subset of the D-serine positive neurons contained SR in the neocortex and HP. Greater than half of the D-serine positive neurons were GABAergic interneurons, with a majority of these neurons containing parvalbumin and/or somatostatin. Only ∼25-40 % of interneurons expressed SR in the neocortex and HP. Finally, we demonstrate in human post-mortem neocortex that SR is found in both excitatory and inhibitory neurons, but not in S100β-containing astrocytes. In sum, these findings conclusively demonstrate that the majority of D-serine is both synthesized and stored in neurons. It will be important to determine the functional significance for the separation of synthesis and storage of D-serine in neurons, as well as the presence of this NMDAR co-agonist in GABAergic interneurons.
Bibliographical noteFunding Information:
Formalin-fixed primary motor cortex (BA4) brain tissue from 3 male control cases (AN18870, AN10180, AN16218) was provided by the Harvard Brain Tissue Resource Center that is supported in part by PHS Grant
Acknowledgments We would like to thank Drs. Sabina Berretta, Sue Andersen, and Ole Isaacson for the generous use of their microscopes. We would also like to thank Harry Pantazopoulos, Rebecca Shivangi Misra, Anita Bechtholt, Jesse McLean, and John Muschamp for technical advice, and Jiamin Feng for animal colony maintenance and genotyping. We thank Francine Benes and the staff of the Harvard Brain Tissue Resource Center that is supported in part by PHS grant number R24-MH068855 for providing all the tissues, as well as the study subjects and their families. An Andrew P. Merrill Research Fellowship and Phyllis & Jerome Lyle Rappaport Mental Health Research Scholars Award (DTB), as well as the National Institutes of Health grants R01MH05190 and P50MH0G0450 (JTC) supported this work.
- NMDA receptor