Viral Neuronal Tracing

Viral neuronal tracing is the use of a virus to trace neural pathways, providing a self-replicating tracer. Viruses have the advantage of self replication over molecular tracers, but can also spread too quickly and cause degradation of neural tissue. Viruses which can infect the nervous system, called neurotropic viruses, spread through spatially close assemblies of neurons through synapses, allowing for their use in studying functionally connected neural networks. The use of viruses to label functionally connected neurons stems from work done by Albert Sabin who developed a bioassay which could assess the infection of viruses across neurons. Subsequent research allowed for incorporation of immunohistochemical techniques to systematically label neuronal connections. To date, viruses have been used to study multiple circuits in the nervous system.

Most neuroanatomists would agree that understanding how the brain is connected to itself and the body is of paramount importance. As such, it is of equal importance to have a way to visualize and study the connections among neurons. Neuronal tracing methods offer an unprecedented view into the morphology and connectivity of neural networks. Depending on the tracer used, this can be limited to a single neuron or can progress trans-synaptically to adjacent neurons. After the tracer has spread sufficiently, the extent may be measured either by fluorescence (for dyes) or by immunohistochemistry (for biological tracers). An important innovation in this field is the use of neurotropic viruses as tracers. These not only spread throughout the initial site of infection, but can jump across synapses. The use of a virus provides a self-replicating tracer. This can allow for the elucidation of neural microcircuitry to an extent that was previously unobtainable. This has significant implications f...

The life cycle of viruses, such as those used in neuronal tracing, is different from cellular organisms. Viruses are parasitic in nature and cannot proliferate on their own. Therefore, they must infect another organism and effectively hijack cellular machinery to complete their life cycle. The first stage of the viral life cycle is called viral entry. This defines the manner in which a virus infects a new host cell. In nature, neurotropic viruses are usually transmitted through bites or scratches, as in the case of Rabies virus or certain strains of Herpes viruses. In tracing studies, this step occurs artificially, typically through the use of a syringe. The next stage of the viral life cycle is called viral replication. During this stage, the virus takes over the host cell's machinery to cause the cell to create more viral proteins and assemble more viruses. Once the cell has produced a sufficient number of viruses, the virus enters the viral shedding stage. During this stage, vi...

Infection edit 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 ...

Viruses can be transmitted in one of two directions. First, one must understand the underlying mechanism of axoplasmic transport. Within the axon are long slender protein complexes called microtubules. They act as a cytoskeleton to help the cell maintain its shape. These can also act as highways within the axon and facilitate transport of neurotransmitter-filled vesicles and enzymes back and forth between the cell body, or soma and the axon terminal, or synapse. Transport can proceed in either direction: anterograde (from soma to synapse), or retrograde (from synapse to soma). Neurons naturally transport proteins, neurotransmitters, and other macromolecules via these cellular pathways. Neuronal tracers, including viruses, take advantage of these transport mechanisms to distribute a tracer throughout a cell. Researchers can use this to study synaptic circuitry. Anterograde transport edit Anterograde tracing is the use of a tracer that moves from soma to synapse. Anterograde transport us...

The use of viruses as tracers has its benefits and its drawbacks. As such, there are some applications in which viruses are an excellent tracer, and other applications in which there are better methods to use. Benefits edit One of the benefits of using viral tracers is the ability of the virus to jump across synapses. This allows for tracing of microcircuitry as well as projection studies. Few molecular tracers are able to do this, and those that can usually have a decreased signal in secondary neurons. Therefore, another benefit of viral tracing is the ability of viruses to self-replicate. As soon as the secondary neuron is infected, the virus begins multiplying and replicating. There is no loss of signal as the tracer propagates through the brain. Drawbacks edit Although some characteristics of viruses present a number of advantages in tracing, others present potential problems. As they propagate through the nervous system, the viral tracers infect neurons and ultimately destroy them...

Viral tracing is primarily used to trace neuronal circuits. Researchers use one of the previously mentioned viruses to study how neurons in the brain are connected to each other with a very fine level of detail. Connectivity largely determines how the brain functions. Viruses have been used to study retinal ganglion circuits, cortical circuits, and spinal circuits, among others.