Methods

Infectionedit

The viral tracer may be introduced in peripheral organs, such as a muscle or gland. Certain viruses, such as adeno-associated virus can be injected into the blood stream and cross the blood–brain barrier to infect the brain. It may also be introduced into a ganglion or injected directly into the brain using a stereotactic device. These methods offer unique insight into how the brain and its periphery are connected. Viruses are introduced into neuronal tissue in many different ways. There are two major methods to introduce tracer into the target tissues. Pressure injection requires the tracer, in liquid form, to be injected directly into the cell. This is the most common method. Iontophoresis involves the application of current to the tracer solution within an electrode. The tracer molecules pick up a charge and are driven into the cell via the electric field. This is a useful method if you wish to label a cell after performing the patch clamp technique. Once the tracer is introduced into the cell, the aforementioned transport mechanisms take over. Once introduced into the nervous system, the virus will begin to infect cells in the local area. The viruses function by incorporating their own genetic material into the genome of the infected cells. The host cell will then produce the proteins encoded by the gene. Researchers are able to incorporate numerous genes into the infected neurons, including fluorescent proteins used for visualization. Further advances in neuronal tracing allow for targeted expression of fluorescent proteins to specific cell types.

Histology and imagingedit

Once the virus has spread to the desired extent, the brain is sliced and mounted on slides. Then, fluorescent antibodies specific for the virus or fluorescent complementary DNA probes for viral DNA are washed over the slices and imaged under a fluorescence microscope.