Transneuronal circuit tracing with neurotropic viruses

ArticleinCurrent opinion in neurobiology 18(6):617-23 · May 2009with17 Reads
DOI: 10.1016/j.conb.2009.03.007 · Source: PubMed
Because neurotropic viruses naturally traverse neural pathways, they are extremely valuable for elucidating neural circuits. Naturally occurring herpes and rabies viruses have been used for transneuronal circuit tracing for decades. Depending on the type of virus and strain, virus can travel preferentially in the anterograde or the retrograde direction. More recently, genetic modifications have allowed for many improvements. These include: reduced pathogenicity; addition of marker genes; control of synaptic spread; pseudotyping for infection of selected cells; addition of ancillary genetic elements for combining circuit tracing with manipulation of activity or functional assays. These modifications, along with the likelihood of future developments, suggest that neurotropic viruses will be increasingly important and effective tools for future studies of neural circuits.
    • "We envision future experiments using effector genes for genetic profiling (e.g. RIBO-TRAP [Sanz et al., 2009] ), rabiesmediated monosynaptic anatomical tracing (Callaway, 2008 ), and functional perturbation of behavior (Liu et al., 2012) (see Figure 3—figuresupplement 4e,j). In cases where other, larger promoters Representative images of prefrontal cortex showing EGFP (green), mKate2 (red) and GAD67 (blue) fluorescence in rats subjected to IS or HC conditions. "
    [Show abstract] [Hide abstract] ABSTRACT: Understanding how the brain captures transient sensory experience and converts it into long lasting changes in neural circuits requires the identification and investigation of the specific ensembles of neurons that are responsible for the encoding of each experience. We have developed a Robust Activity Marking (RAM) system that allows for the identification and interrogation of ensembles of neurons. The RAM system provides unprecedented high sensitivity and selectivity through the use of an optimized synthetic activity-regulated promoter that is strongly induced by neuronal activity and a modified Tet-Off system that achieves improved temporal control. Due to its compact design, RAM can be packaged into a single adeno-associated virus (AAV), providing great versatility and ease of use, including application to mice, rats, flies, and potentially many other species. Cre-dependent RAM, CRAM, allows for the study of active ensembles of a specific cell type and anatomical connectivity, further expanding the RAM system’s versatility.
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    • "As shortly mentioned in the clearing section, this caveat can be overcome by using transsynaptic viruses that can cross synapses and thus circumvent the requirement to confirm connectivity at a resolution achieved by electron microscopy only, because the connectivity can be inferred from the known direction and mechanism of spread of the transsynaptic tracer [33][34][35]. Especially transsynaptic tracers based on rabies virus which cross synaptic connections in a strictly retrograde direction are powerful tools for studying neuronal networks [13, 27,[36][37][38]. Currently, there are two approaches to the automation of light microscopic imaging of 3D whole brain datasets. "
    [Show abstract] [Hide abstract] ABSTRACT: Magnetic resonance imaging, positron emission tomography, and optical imaging have emerged as key tools to understand brain function and neurological disorders in preclinical mouse models. They offer the unique advantage of monitoring individual structural and functional changes over time. What remained unsolved until recently was to generate whole-brain microscopy data which can be correlated to the 3D in vivo neuroimaging data. Conventional histological sections are inappropriate especially for neuronal tracing or the unbiased screening for molecular targets through the whole brain. As part of the European Society for Molecular Imaging (ESMI) meeting 2016 in Utrecht, the Netherlands, we addressed this issue in the Molecular Neuroimaging study group meeting. Presentations covered new brain clearing methods, light sheet microscopes for large samples, and automatic registration of microscopy to in vivo imaging data. In this article, we summarize the discussion; give an overview of the novel techniques; and discuss the practical needs, benefits, and limitations.
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    • "On the other hand, cytotoxicity is a major concern when using neurotropic viruses. As a consequence, these viruses are less suited for long-term experiments unlike WGA–Cre-dependent labeling (Callaway 2008; Kelly and Strick 2003; Viral et al. 2012 ). Besides, Gradinaru et al. showed that WGA–Credependent labeling provides ample topologic expression for circuit manipulation by channelrhodopsins (Gradinaru et al. 2010). "
    [Show abstract] [Hide abstract] ABSTRACT: Novel neuromodulation techniques in the field of brain research, such as optogenetics, prompt to target specific cell populations. However, not every subpopulation can be distinguished based on brain area or activity of specific promoters, but rather on topology and connectivity. A fascinating tool to detect neuronal circuitry is based on the transsynaptic tracer, wheat germ agglutinin (WGA). When expressed in neurons, it is transported throughout the neuron, secreted, and taken up by synaptically connected neurons. Expression of a WGA and Cre recombinase fusion protein using a viral vector technology in Cre-dependent transgenic animals allows to trace neuronal network connections and to induce topological transgene expression. In this study, we applied and evaluated this technology in specific areas throughout the whole rodent brain, including the hippocampus, striatum, substantia nigra, and the motor cortex. Adeno-associated viral vectors (rAAV) encoding the WGA–Cre fusion protein under control of a CMV promoter were stereotactically injected in Rosa26-STOP-EYFP transgenic mice. After 6 weeks, both the number of transneuronally labeled YFP+/mCherry− cells and the transduced YFP+/mCherry+ cells were quantified in the connected regions. We were able to trace several connections using WGA–Cre transneuronal labeling; however, the labeling efficacy was region-dependent. The observed transneuronal labeling mostly occurred in the anterograde direction without the occurrence of multi-synaptic labeling. Furthermore, we were able to visualize a specific subset of newborn neurons derived from the subventricular zone based on their connectivity.
    Article · Jun 2016
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