Can transsynaptic viral strategies be used to reveal functional aspects of neural circuitry?

Viruses have proved instrumental to elucidating neuronal connectivity relationships in a variety of organisms. Recent advances in genetic technologies have facilitated analysis of neurons directly connected to a defined starter population. These advances have also made viral transneuronal mapping available to the broader neuroscience community, where one-step rabies virus mapping has become routine. This method is commonly used to identify inputs onto defined cell populations, to demonstrate the quantitative proportion of inputs coming from specific brain regions, or to compare input patterns between two or more cell populations. Furthermore, the number of inputs labeled is often assumed to reflect the number of synaptic connections, and these viruses are commonly believed to label strong synapses more efficiently than weak synapses. While these maps are often interpreted to provide a quantitative estimate of the synaptic landscape onto starter cell populations, in fact very little is known about how transneuronal transmission takes place. We do not know how these viruses transmit between neurons, if they display biases in the cell types labeled, or even if transmission is synapse-specific. In this review, we discuss the experimental evidence against or in support of key concepts in viral tracing, focusing mostly on the use of one-step rabies input mapping and related methods. Does spread of these viruses occur specifically through synaptic connections, preferentially through synapses, or non-specifically? How efficient is viral transneuronal transmission, and is this efficiency equal in all cell types? And lastly, to what extent does viral labeling reflect functional connectivity?