Quantitative analysis of rabies virus-based synaptic connectivity tracing

Monosynaptically restricted rabies viruses have been used for more than a decade for synaptic connectivity tracing. However, the verisimilitude of quantitative conclusions drawn from these experiments is largely unknown. The primary reason is the simple metrics commonly used, which generally disregard the effect of starter cell numbers. Here we present an experimental dataset with a broad range of starter cell numbers and explore their relationship with the number of input cells across the brain using descriptive statistics and modelling. We show that starter cell numbers strongly affect input fraction and convergence index measures, making quantitative comparisons unreliable. Furthermore, we suggest a principled way to analyse rabies derived connectivity data by taking advantage of the starter vs input cell relationship that we describe and validate across independent datasets.