A versatile viral toolkit for functional discovery in the nervous system

Advances in approaches to study cell-type specific cortical circuits throughout development

“…Several currently available mouse lines target Cre or Flp expression to different populations of excitatory projection neurons ( Gerfen et al, 2013 ; Daigle et al, 2018 ; Matho et al, 2021 ; Vaasjo et al, 2022 ) and inhibitory interneurons ( Taniguchi et al, 2011 ; He et al, 2016 ). Furthermore, several labs are expanding available AAVs for cell-type specific targeting ( Nair et al, 2020 ; Vormstein-Schneider et al, 2020 ; Graybuck et al, 2021 ; Mich et al, 2021 ; Pouchelon et al, 2022 ). Experimental strategies that combine Cre and Flp refine targeting of cell types in an intersectional manner by considering multiple genetic features ( Fenno et al, 2014 , 2017 ).…”

“… He et al (2016) successfully targeted these distinct VIP/CR and VIP/CCK subtypes by crossing VIP-Flp mice to CR-ires-Cre or CCK-ires-Cre mice and an Ai65 dual conditional Cre/Flp reporter line ( Figure 1A ). Advances in designing AAV vectors promise to make such intersectional experiments more tractable ( Nair et al, 2020 ; Vormstein-Schneider et al, 2020 ; Graybuck et al, 2021 ; Mich et al, 2021 ; Pouchelon et al, 2022 ). For example, Graybuck et al (2021) generated AAVs that use cell-type specific enhancers to drive expression of Cre, Flp, or Nigri recombinases.…”

“…Using these tools, they successfully tagged PT-type cells, IT-type cells, and interneurons with unique fluorophores throughout the cortex without the need for complicated transgenic crosses ( Figure 1B ). Finally, Pouchelon et al (2022) developed a set of AAV backbones that allow for boolean intersectional experiments that include: Cre-ON, Flp-ON, Cre-ON;Flp-ON, Cre-ON;Flp-OFF, and Cre-OFF;Flp-ON. These backbones can be integrated with several existing technologies (e.g., optogenetics, calcium-indicators, and DREADDs) and then used with available conditional transgenic mice.…”

A Novel Single Vector Intersectional AAV Strategy for Interrogating Cellular Diversity and Brain Function

“…Adeno-associated virus (AAV)-based reporter tools are an alternative approach that offer regional and temporal specificity and are relatively quick and inexpensive to generate. Because of these benefits, several elegant intersectional AAV tools with varying levels of in vivo functionality have recently emerged [8][9][10][11][12][13] . While useful, these tools also present a variety of challenges for the end user that may discourage their widespread adoption, including complex design parameters, use of large stop-cassettes that impede viral packaging capacity limits, dependence on multiple vectors for payload expression, or reliance on tetracycline-based expression systems which have reported issues with toxicity or leak expression [14][15][16][17] .…”

“…Yet, as our understanding of cellular diversity in the brain expands, so does the need for more sophisticated tools that can selectively target cells based on multiple features. To address this need, several transgenic [5][6][7] and viral-based [8][9][10][11][12][13] intersectional strategies have been developed, each with unique strengths and weaknesses. Compared to viral strategies, transgenic approaches can better accommodate reporter transgenes that utilize large stop cassettes and may also be preferable for driving intersectional expression during development or in cell populations that are difficult to access [5][6][7] .…”