SPECIFIC PROTEOME OF MAMMALIAN CORTEX INHIBITORY & EXCITATORY SYNAPSES

Rockefeller University, New York NY

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Abstract

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Four years ago, in collaboration with Nat Heintz (Rockefeller University), we initiated the study of the protein complement present at excitatory synapses in Purkinje cells. We used the Bacterial Artificial Chromosome (BAC) modification strategy to target the specific in vivo expression of GFP-fused GRID 2 to Purkinje cell's excitatory synapses. We performed dissections of mouse cerebella, and purified synapses bearing GFP-GRID2. Although challenging, our approach proved successful, as we isolated synapses and analyzed low-femtomol levels of proteins. During this last year, we continued our mass spectrometric analyses and identified ~70 synaptic proteins, confirming known excitatory proteins, the absence of inhibitory proteins, and identifying novel signatures of excitatory synapses. We are currently investigating their biological significance. Using a similar approach to the one described above, we have started the study of inhibitory synapses by isolating GABA receptors from specific cell populations in the Cortex. Together with Liz Heller in the laboratory of Nat Heintz, we have also studied several ribosomal protein interactions in the mammalian brain, such as the ribosomal L10 protein in the Cerebellum.