ABSTRACT
When a single neuron is grown on a small island of glial cells, the neuron forms synapses onto itself. The so-called autaptic culture systems have proven extremely valuable in elucidating basic mechanisms of synaptic transmission, as they allow application of technical approaches that cannot be used in slice preparations. However, this method has been almost exclusively used for pyramidal cells and interneurons. In this study, we generated autaptic cultures from granule cells isolated from the dentate gyrus of rodent hippocampi. Our subsequent morphological and functional characterisation of these cells confirms that this culture model is suitable for investigating basic mechanisms of granule cell synaptic transmission. Importantly, the autosynaptic connectivity allows recordings of pure mossy fibre miniature EPSCs, which are not possible in slice preparations. Further, by fast application of hypertonic sucrose solutions it is possible to directly measure the readily releasable pool and to calculate the probability of vesicular release.
Subject(s)
Hippocampus/cytology , Hippocampus/physiology , Neurogenesis/physiology , Neuroglia/cytology , Neuroglia/physiology , Neurons/cytology , Neurons/physiology , Animals , Cells, Cultured , Culture Techniques , Excitatory Postsynaptic Potentials/physiology , Mice , Mice, Inbred C57BL , Rats , Rats, Sprague-Dawley , Synapses/physiologyABSTRACT
Protein-protein interaction maps provide a valuable framework for a better understanding of the functional organization of the proteome. To detect interacting pairs of human proteins systematically, a protein matrix of 4456 baits and 5632 preys was screened by automated yeast two-hybrid (Y2H) interaction mating. We identified 3186 mostly novel interactions among 1705 proteins, resulting in a large, highly connected network. Independent pull-down and co-immunoprecipitation assays validated the overall quality of the Y2H interactions. Using topological and GO criteria, a scoring system was developed to define 911 high-confidence interactions among 401 proteins. Furthermore, the network was searched for interactions linking uncharacterized gene products and human disease proteins to regulatory cellular pathways. Two novel Axin-1 interactions were validated experimentally, characterizing ANP32A and CRMP1 as modulators of Wnt signaling. Systematic human protein interaction screens can lead to a more comprehensive understanding of protein function and cellular processes.
