Effects Of Different Titers Of RAAV5 On Transcriptome Of Hippocampal Pyramidal Neurons And Astrocytes

Recombinant adeno-associated virus(rAAV)vectors have been widely used in neuroscience for gene transduction,and multiple commonly used rAAV serotypes have been characterized with their tropism and efficiency.However,little is known regarding the impact of diffenrent titers rAAV vectors on brain cells.Here,we evaluated the effects of three different titers rAAV 5 vectors(1011,1012 and 1013v.g./mL)on transcriptome of hippocampal pyramidal neurons and astrocytes,playing important roles in learning,memory,navigation and mood regulation.rAAV5-DIO-mCherry was injected into the Camk Ⅱα-cre mice and Aldh1L1-cre-ERT2 mice for labeling pyramidal neurons and astrocytes,respectively.mCherry positive and DAPI negative cells were isolated for RNA sequencing.We found that high-titer rAAV5 up-regulates the genes involved in neuron death and mitochondrial respiratory chain in pyramidal neurons and astrocytes respectively,meanwhile it down-regulates the genes related to synapse part and golgi vescle transport in pyramidal neurons and astrocytes,respectively.These results reveal that as titer increases,the immunogenicity and cytotoxicity of rAAV5 gradually emerge.High-titer rAAV5 may enhance the sensitivity of pyramidal neurons to external stimuli and impair the synaptic signal transmission and neural plasticity.Vesicle transport and signal transmission in astrocytes may be disturbed by high-titer rAAV 5.Given that rAAV vectors are widely used for gene delivery,we suggest that it is better to test the infection efficiency of rAAV with different titers before experiment,and to avoid the use of high-titer rAAV as far as possible under the premise of meeting the experimental requirements.