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Protein fingerprints of cultured CA3-CA1 hippocampal neurons: comparative analysis of the distribution of synaptosomal and cytosolic proteins.

ABSTRACT: BACKGROUND: All studies aimed at understanding complex molecular changes occurring at synapses face the problem of how a complete view of the synaptic proteome and of its changes can be efficiently met. This is highly desirable when synaptic plasticity processes are analyzed since the structure and the biochemistry of neurons and synapses get completely reshaped. Because most molecular studies of synapses are nowadays mainly or at least in part based on protein extracts from neuronal cultures, this is not a feasible option: these simplified versions of the brain tissue on one hand provide an homogeneous pure population of neurons but on the other yield only tiny amounts of proteins, many orders of magnitude smaller than conventional brain tissue. As a way to overcome this limitation and to find a simple way to screen for protein changes at cultured synapses, we have produced and characterized two dimensional electrophoresis (2DE) maps of the synaptic proteome of CA3-CA1 hippocampal neurons in culture. RESULTS: To obtain 2D maps, hippocampal cultures were mass produced and after synaptic maturation, proteins were extracted following subfractionation procedures and separated by 2D gel electrophoresis. Similar maps were obtained for the crude cytosol of cultured neurons and for synaptosomes purified from CA3-CA1 hippocampal tissue. To efficiently compare these different maps some clearly identifiable reference points were molecularly identified by mass spectrometry and immunolabeling methods. This information was used to run a differential analysis and establish homologies and dissimilarities in these 2D protein profiles. CONCLUSION: Because reproducible fingerprints of cultured synapses were clearly obtained, we believe that our mapping effort could represent a simple tool to screen for protein expression and/or protein localization changes in CA3-CA1 hippocampal neurons following plasticity.

SUBMITTER: Corti V

PROVIDER: S-EPMC2324106 | biostudies-other | 2008

REPOSITORIES: biostudies-other

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Protein fingerprints of cultured CA3-CA1 hippocampal neurons: comparative analysis of the distribution of synaptosomal and cytosolic proteins.

Corti Valeria V Sanchez-Ruiz Yovan Y Piccoli Giovanni G Bergamaschi Andrea A Cannistraci Carlo V CV Pattini Linda L Cerutti Sergio S Bachi Angela A Alessio Massimo M Malgaroli Antonio A

BMC neuroscience 20080410

<h4>Background</h4>All studies aimed at understanding complex molecular changes occurring at synapses face the problem of how a complete view of the synaptic proteome and of its changes can be efficiently met. This is highly desirable when synaptic plasticity processes are analyzed since the structure and the biochemistry of neurons and synapses get completely reshaped. Because most molecular studies of synapses are nowadays mainly or at least in part based on protein extracts from neuronal cult ...[more]

PMID: 18402664

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Project description:OBJECTIVE:Functions ascribed to the hippocampal sub-regions for encoding episodic memories include the separation of activity patterns propagated from the entorhinal cortex (EC) into the dentate gyrus (DG) and pattern completion in CA3 region. Since a direct assessment of these functions is lacking at the level of specific axonal inputs, our goal is to directly measure the separation and completion of distinct axonal inputs in engineered pairs of hippocampal sub-regional circuits. APPROACH:We co-cultured EC-DG, DG-CA3, CA3-CA1 or CA1-EC neurons in a two-chamber PDMS device over a micro-electrode array (MEA60), inter-connected via distinct axons that grow through the micro-tunnels between the compartments. Taking advantage of the axonal accessibility, we quantified pattern separation and completion of the evoked activity transmitted through the tunnels from source into target well. Since pattern separation can be inferred when inputs are more correlated than outputs, we first compared the correlations among axonal inputs with those of target somata outputs. We then compared, in an analog approach, the distributions of correlation distances between rate patterns of the axonal inputs inside the tunnels with those of the somata outputs evoked in the target well. Finally, in a digital approach, we measured the spatial population distances between binary patterns of the same axonal inputs and somata outputs. MAIN RESULTS:We found the strongest separation of the propagated axonal inputs when EC was axonally connected to DG, with a decline in separation to CA3 and to CA1 for both rate and digital approaches. Furthermore, the digital approach showed stronger pattern completion in CA3, then CA1 and EC. SIGNIFICANCE:To the best of our knowledge, these are the first direct measures of pattern separation and completion for axonal transmission to the somata target outputs at the rate and digital population levels in each of four stages of the EC-DG-CA3-CA1 circuit.

Dataset Information

Protein fingerprints of cultured CA3-CA1 hippocampal neurons: comparative analysis of the distribution of synaptosomal and cytosolic proteins.

Publications

Protein fingerprints of cultured CA3-CA1 hippocampal neurons: comparative analysis of the distribution of synaptosomal and cytosolic proteins.

Similar Datasets

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