Retrograde tracing is a research method used in neuroscience to track the neural connections from the termination point (synapse) to the source (cell body). Retrograde retrieval techniques allow detailed assessment of neural connections between the target population of neurons and their inputs across the nervous system. These techniques allow "mapping" connections between neurons in certain structures (eg the eye) and target neurons in the brain. The opposite technique is anterograde tracing, which is used to trace the neural connections from their source to their cessation point (ie from body cells to synapses). The anterograde and retrograde techniques of tracing are based on axonal transport visualization.
Video Retrograde tracing
Technique
Retrograde search can be achieved in a variety of ways, including the use of virus strains as a cell connectivity marker to the injection site. The pseudorabies virus (PRV, Bartha strain), for example, can be used as a suitable tracer because of infectious tendencies to spread upstream through synaptic linked neuronal pathways, thus revealing the nature of their circuits.
Rabies has been shown to be effective for circuit tracking systems because of the low level of damage to infected cells, specificity only infects neurons, and strict limitations of viral spread between neurons to synaptic regions. These factors allow for very specific traces that can reveal individual nerve connections in the circuit without inflicting physical damage to the cell.
Another technique involves injecting special "beads" into the anesthetic brain core of the animal. The animals were left alive for several days and then euthanized. The cells in the origin of the projection are visualized through an inverse fluorescence microscope.
The specialist technique was developed by Wickersham and colleagues, who used a modified rabies virus. The virus is capable of infecting single cells and jumping across a synapse; this allows researchers to investigate the local connectivity of neurons.
Rabies virus
Once taken at the synaptic terminal or axon of the target neuron, the rabies virus is enveloped in vesicles that are transported to the cells of the body through axonal dyneins. In the wildtype rabies virus, the virus will continue to replicate and spread throughout the central nervous system until it systematically infects the entire brain. The removal of genes encoding glycoprotein (protein G) in rabies limits strict viral spread to infected cells initially. Transsynaptic viral spread may be limited to monosynaptic transmission to the neurons of origin with pseudotyping of G proteins and placing the gene under Cre-control. The spread of this virus can be visualized through methods including the addition of fluorescence genes such as green fluorescent protein to viral tapes or through immunohistochemistry.
Pseudorabies Virus
A member of the herpesviridae family, the pseudorabies virus spreads through the CNS in both retrograde and anterograde modes, moving up the neural axons into the soma and dendrites in retrograde applications. The elimination of three key membrane protein genes in the PRV-Bartha pseudorabies strain precludes the spread of anterograde virus and allows additional manipulation into viral DNA such as fluorescence to be added, enabling retrograde circuit tracking.
Fluoro-Gold
Fluoro-Gold, also known as hydroxystilbamidine, is a non-viral fluorescent retrograde tracer that moves up axons and traverses dendritic trees can be visualized through fluorescent or immunohistochemical microscopy.
Maps Retrograde tracing
See also
- Anterograde tracing
- Neural pathways
- Virus neuronal tracking
References
Further reading
Source of the article : Wikipedia
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