Optimization of virus-mediated functional imaging based on quantitative evaluation of transduction efficiency in pigeon (Columba livia domestica)

Currently, virus-mediated functional imaging integrated with optogenetic techniques has been used successfully in both neuroanatomical track tracing and accurate regulation of animal's social and cognitive behaviors. In avian, only a few studies focused on the mesencephalon and brainstem, compared to those on the telencephalon and diencephalon. To widen adeno-associated viruses (AAV) in avian studies, we tested the transduction efficiency of different types of sera and promoters and designed a Python script to analyze quantitatively the fluorescence intensity of brain slices. Our experiments on pigeons consisted of the serotypes for anterograde and retrograde labeling, promoters for virus gene transcription, injection doses and periods for transduction, and different optogenetic components. We found serotypes 1 and 11 with ubiquitous promoters transducing well at all brain regions, AAV transduction being more efficient in the telencephalon than mesencephalon and the hChR2 component significantly affecting transduction efficiency. The optimum injection dose and transduction period guideline for each location and serotype were determined. Our findings can be refereed during the related study and will aid future research in avian.