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      Neuronal medium that supports basic synaptic functions and activity of human neurons in vitro.

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          Abstract

          Human cell reprogramming technologies offer access to live human neurons from patients and provide a new alternative for modeling neurological disorders in vitro. Neural electrical activity is the essence of nervous system function in vivo. Therefore, we examined neuronal activity in media widely used to culture neurons. We found that classic basal media, as well as serum, impair action potential generation and synaptic communication. To overcome this problem, we designed a new neuronal medium (BrainPhys basal + serum-free supplements) in which we adjusted the concentrations of inorganic salts, neuroactive amino acids, and energetic substrates. We then tested that this medium adequately supports neuronal activity and survival of human neurons in culture. Long-term exposure to this physiological medium also improved the proportion of neurons that were synaptically active. The medium was designed to culture human neurons but also proved adequate for rodent neurons. The improvement in BrainPhys basal medium to support neurophysiological activity is an important step toward reducing the gap between brain physiological conditions in vivo and neuronal models in vitro.

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          Most cited references30

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          Direct conversion of fibroblasts to functional neurons by defined factors

          Cellular differentiation and lineage commitment are considered robust and irreversible processes during development. Recent work has shown that mouse and human fibroblasts can be reprogrammed to a pluripotent state with a combination of four transcription factors. This raised the question of whether transcription factors could directly induce other defined somatic cell fates, and not only an undifferentiated state. We hypothesized that combinatorial expression of neural lineage-specific transcription factors could directly convert fibroblasts into neurons. Starting from a pool of nineteen candidate genes, we identified a combination of only three factors, Ascl1, Brn2, and Myt1l, that suffice to rapidly and efficiently convert mouse embryonic and postnatal fibroblasts into functional neurons in vitro. These induced neuronal (iN) cells express multiple neuron-specific proteins, generate action potentials, and form functional synapses. Generation of iN cells from non-neural lineages could have important implications for studies of neural development, neurological disease modeling, and regenerative medicine.
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            Rapid single-step induction of functional neurons from human pluripotent stem cells.

            Available methods for differentiating human embryonic stem cells (ESCs) and induced pluripotent cells (iPSCs) into neurons are often cumbersome, slow, and variable. Alternatively, human fibroblasts can be directly converted into induced neuronal (iN) cells. However, with present techniques conversion is inefficient, synapse formation is limited, and only small amounts of neurons can be generated. Here, we show that human ESCs and iPSCs can be converted into functional iN cells with nearly 100% yield and purity in less than 2 weeks by forced expression of a single transcription factor. The resulting ES-iN or iPS-iN cells exhibit quantitatively reproducible properties independent of the cell line of origin, form mature pre- and postsynaptic specializations, and integrate into existing synaptic networks when transplanted into mouse brain. As illustrated by selected examples, our approach enables large-scale studies of human neurons for questions such as analyses of human diseases, examination of human-specific genes, and drug screening. Copyright © 2013 Elsevier Inc. All rights reserved.
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              Sugar for the brain: the role of glucose in physiological and pathological brain function.

              The mammalian brain depends upon glucose as its main source of energy, and tight regulation of glucose metabolism is critical for brain physiology. Consistent with its critical role for physiological brain function, disruption of normal glucose metabolism as well as its interdependence with cell death pathways forms the pathophysiological basis for many brain disorders. Here, we review recent advances in understanding how glucose metabolism sustains basic brain physiology. We synthesize these findings to form a comprehensive picture of the cooperation required between different systems and cell types, and the specific breakdowns in this cooperation that lead to disease. Copyright © 2013 Elsevier Ltd. All rights reserved.
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                Author and article information

                Journal
                Proc. Natl. Acad. Sci. U.S.A.
                Proceedings of the National Academy of Sciences of the United States of America
                1091-6490
                0027-8424
                May 19 2015
                : 112
                : 20
                Affiliations
                [1 ] Salk Institute for Biological Studies, Sanford Consortium for Regenerative Medicine, La Jolla, CA 92037; cbardy@salk.edu gage@salk.edu.
                [2 ] Salk Institute for Biological Studies, Sanford Consortium for Regenerative Medicine, La Jolla, CA 92037; Department of Psychiatry and Neuropsychology, Division of Translational Neuroscience, Maastricht University, 6200 MD, Maastricht, The Netherlands;
                [3 ] Salk Institute for Biological Studies, Sanford Consortium for Regenerative Medicine, La Jolla, CA 92037;
                [4 ] Salk Institute for Biological Studies, Sanford Consortium for Regenerative Medicine, La Jolla, CA 92037; School of Life Sciences, Ecole Polytechnique Fédérale de Lausanne, CH-1015 Lausanne, Switzerland; and.
                [5 ] Conrad Prebys Center for Chemical Genomics, Sanford-Burnham Medical Research Institute, La Jolla, CA 92037.
                Article
                1504393112
                10.1073/pnas.1504393112
                4443325
                25870293
                a15a20a2-ea0f-428e-8b96-eaf2d84b08ac
                History

                BrainPhys,induced pluripotent stem cells,neurobasal DMEM,neuromedium,tissue culture milieu

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