Selective Gene Expression by Postnatal Electroporation during Olfactory Interneuron Neurogenesis

Department of Biological Sciences, Columbia University, New York, New York, USA.
PLoS ONE (Impact Factor: 3.23). 02/2008; 3(1):e1517. DOI: 10.1371/journal.pone.0001517
Source: PubMed


Neurogenesis persists in the olfactory system throughout life. The mechanisms of how new neurons are generated, how they integrate into circuits, and their role in coding remain mysteries. Here we report a technique that will greatly facilitate research into these questions. We found that electroporation can be used to robustly and selectively label progenitors in the Subventicular Zone. The approach was performed postnatally, without surgery, and with near 100% success rates. Labeling was found in all classes of interneurons in the olfactory bulb, persisted to adulthood and had no adverse effects. The broad utility of electroporation was demonstrated by encoding a calcium sensor and markers of intracellular organelles. The approach was found to be effective in wildtype and transgenic mice as well as rats. Given its versatility, robustness, and both time and cost effectiveness, this method offers a powerful new way to use genetic manipulation to understand adult neurogenesis.

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Available from: Jessica H Brann, Jan 19, 2014
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    • "In utero electroporation method has been widely used to clarify molecular mechanisms of the embryonic cortical (Borrell et al., 2005; Tabata and Nakajima, 2008; Nishimura et al., 2012) and hippocampal (Tomita et al., 2011; Choe et al., 2013) development. Electroporation has also been applied to the analysis of postnatal development of olfactory neurons (Boutin et al., 2008; Chesler et al., 2008; Fernandez et al., 2011). "
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    ABSTRACT: Electroporation-mediated gene transfer has been developed for the analysis of mammalian brain development in vivo. Indeed, in utero electroporation method is widely used for the investigation of the mouse embryonic cortical development while in vivo electroporation using neonatal mouse brain is employed for the analysis of the rostral migratory stream (RMS) and postnatal olfactory neurogenesis. In the present study, we established a stable gene-transfer method to dentate gyrus (DG) neurons by carefully determining the in vivo electroporation conditions, such as position and direction of electrode, voltage for electric pulses, and interval between electroporation and sample preparation. Consequently, GFP-positive cells in DG were observed to extend branched dendrites and long axons into the molecular layer and the hilus, respectively, 21 days after electrporation. They were morphologically identified as dentate granule neurons with many protrusions on dendrites, and some of them had wide head and thin neck that resembled matured mushroom spines. Expression of GFP in dentate neurons sustained for at least 9 months after electroporation under our experimental conditions. Taken together, the method developed here could be a powerful new tool for the analysis of the postnatal DG development. © 2014 Wiley Periodicals, Inc.
    Hippocampus 12/2014; 24(12). DOI:10.1002/hipo.22325 · 4.16 Impact Factor
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    • "Initially, it was used for in utero manipulation of radial glia surrounding the lateral ventricles of the mouse embryo [13]. More recently, this technique has been extended for use in postnatal and adult brains [14-16]. However, except for one report in the hippocampus [14], most uses of EP to study neural stem cells involve injection of plasmid DNA into the brain ventricles. "
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    ABSTRACT: Recent findings have indicated the presence of a progenitor domain at the marginal zone/layer 1 of the cerebral cortex, and it has been suggested that these progenitors have neurogenic and gliogenic potential. However, their contribution to the histogenesis of the cortex remains poorly understood due to difficulties associated with genetically manipulating these unique cells in a population-specific manner. We have adapted the electroporation technique to target pial surface cells for rapid genetic manipulation at postnatal day 2. In vivo data show that most of these cells proliferate and progressively differentiate into both neuronal and glial subtypes. Furthermore, these cells localize to the superficial layers of the optic tectum and cerebral cortex prior to migration away from the surface. We provide a foundation upon which future studies can begin to elucidate the molecular controls governing neural progenitor fate, migration, differentiation, and contribution to cortical and tectal histogenesis. Furthermore, specific genetic targeting of such neural progenitor populations will likely be of future clinical interest.
    Neural Development 07/2012; 7(1):26. DOI:10.1186/1749-8104-7-26 · 3.45 Impact Factor
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    • "Implementing a highly selective in vivo genetic targeting strategy, we investigated the synaptic patterns of connectivity that are made between postnatal-born granule cells and their presynaptic targets using engineered rabies virus for monosynaptic circuit tracing. Our experimental design enabled us to selectively target granule cells for rabies infection and propagation via in vivo electroporation [53], allowing direct determination of the cell types presynaptic to targeted granule cells. Using this approach, we observed discrete synaptic networks associated with granule cell microcircuits. "
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    ABSTRACT: The continued addition of new neurons to mature olfactory circuits represents a remarkable mode of cellular and structural brain plasticity. However, the anatomical configuration of newly established circuits, the types and numbers of neurons that form new synaptic connections, and the effect of sensory experience on synaptic connectivity in the olfactory bulb remain poorly understood. Using in vivo electroporation and monosynaptic tracing, we show that postnatal-born granule cells form synaptic connections with centrifugal inputs and mitral/tufted cells in the mouse olfactory bulb. In addition, newly born granule cells receive extensive input from local inhibitory short axon cells, a poorly understood cell population. The connectivity of short axon cells shows clustered organization, and their synaptic input onto newborn granule cells dramatically and selectively expands with odor stimulation. Our findings suggest that sensory experience promotes the synaptic integration of new neurons into cell type-specific olfactory circuits.
    PLoS ONE 12/2011; 6(12):e29423. DOI:10.1371/journal.pone.0029423 · 3.23 Impact Factor
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