Brain-wide N2cG compensation permits glycoprotein-deleted rabies virus to trace neural circuits across multiple synapses

Rabies-viruses-based retrograde tracers can spread across multiple synapses in a retrograde direction in the nervous system of rodents and primates, making them powerful tools for determining the structure and function of the complicated neural circuits of the brain. However, they have some limitations, such as posing high risks to human health and the inability to retrograde trans-synaptic label inputs from genetically-defined starter neurons. Here, we established a new retrograde trans-multi-synaptic tracing method through brain-wide rabies virus glycoprotein (RVG) compensation, followed by glycoprotein-deleted rabies virus (RV-[Formula: see text]G) infection in specific brain regions. Furthermore, in combination with the avian tumor virus receptor A (TVA) controlled by a cell-type-specific promoter, we found that EnvA-pseudotyped RV-[Formula: see text]G can mediate efficient retrograde trans-multi-synaptic transduction from cell-type-specific starter neurons. This study provides new alternative methods for neuroscience researchers to analyze the input neural networks of rodents and nonhuman primates

AAV13 Enables Precise Targeting of Local Neural Populations

As powerful tools for local gene delivery, adeno-associated viruses (AAVs) are widely used for neural circuit studies and therapeutical purposes. However, most of them have the characteristics of large diffusion range and retrograde labeling, which may result in off-target transduction during in vivo application. Here, in order to achieve precise gene delivery, we screened AAV serotypes that have not been commonly used as gene vectors and found that AAV13 can precisely transduce local neurons in the brain, with a smaller diffusion range than AAV2 and rigorous anterograde labeling. Then, AAV13-based single-viral and dual-viral strategies for sparse labeling of local neurons in the brains of C57BL/6 or Cre transgenic mice were developed. Additionally, through the neurobehavioral test in the ventral tegmental area, we demonstrated that AAV13 was validated for functional monitoring by means of carrying Cre recombinase to drive the expression of Cre-dependent calcium-sensitive indicator. In summary, our study provides AAV13-based toolkits for precise local gene delivery, which can be used for in situ small nuclei targeting, sparse labeling and functional monitoring

AAV11 enables efficient retrograde targeting of projection neurons and enhances astrocyte-directed transduction

Viral tracers that enable efficient retrograde labeling of projection neurons are powerful vehicles for structural and functional dissections of the neural circuit and for the treatment of brain diseases. Currently, some recombinant adeno-associated viruses (rAAVs) based on capsid engineering are widely used for retrograde tracing, but display undesirable brain area selectivity due to inefficient retrograde transduction in certain neural connections. Here we developed an easily editable toolkit to produce high titer AAV11 and demonstrated that it exhibits potent and stringent retrograde labeling of projection neurons in adult male wild-type or Cre transgenic mice. AAV11 can function as a powerful retrograde viral tracer complementary to AAV2-retro in multiple neural connections. In combination with fiber photometry, AAV11 can be used to monitor neuronal activities in the functional network by retrograde delivering calcium-sensitive indicator under the control of a neuron-specific promoter or the Cre-lox system. Furthermore, we showed that GfaABC1D promoter embedding AAV11 is superior to AAV8 and AAV5 in astrocytic tropism in vivo, combined with bidirectional multi-vector axoastrocytic labeling, AAV11 can be used to study neuron-astrocyte connection. Finally, we showed that AAV11 allows for analyzing circuit connectivity difference in the brains of the Alzheimer’s disease and control mice. These properties make AAV11 a promising tool for mapping and manipulating neural circuits and for gene therapy of some neurological and neurodegenerative disorders