Objective To verify the neurotypicality of AAV-PHP.eB after tail vein injection in adult mice and its efficiency in crossing the blood-brain barrier(BBB).Methods The rAAV-SYN-GFP plasmid was constructed, and adult C57...Objective To verify the neurotypicality of AAV-PHP.eB after tail vein injection in adult mice and its efficiency in crossing the blood-brain barrier(BBB).Methods The rAAV-SYN-GFP plasmid was constructed, and adult C57 BL mice were injected with AAV PHP.eB: SYN-GFP in the tail vein(300 nL, virus titer 3 × 10~9 vg) and in the prefrontal lobe(50 L, virus tite5 × 10^(11) vg). The green fluorescent protein(GFP) signal in the brain was observed at two weeks, while the GFP signal in the peripheral organs was observed at four weeks. Results Two weeks after tail vein injection, GFP expression was observed throughout the brain especially in the cortex, hippocampus, and geniculate nucleus. No GFP signal was observed or detected by western blotting in the peripheral organs after four weeks. GFP signal was observed mainly at the loca site after prefrontal lobe injection.Conclusion AAV-PHP.eB: SYN-GFP can effectively cross the BBB in adult mice. Using a neuron-specific promoter allows exogenous gene expression in neurons; therefore, AAV-PHP.eB can be used as an effective carrier for studying diseases in the central nervous system(CNS).展开更多
Astrocytes are the most abundant glial cells in the central nervous system.They perform a diverse array of functions,with a critical role in structural integrity,synapse formation,and neurotransmission.These cells exh...Astrocytes are the most abundant glial cells in the central nervous system.They perform a diverse array of functions,with a critical role in structural integrity,synapse formation,and neurotransmission.These cells exhibit substantial regional heterogeneity and display variable responses to different neurological diseases.Such diversity in astrocyte morphology and function is essential for understanding both normal brain function and the underlying mechanisms of neurological disorders.To investigate this heterogeneity,we developed a novel method for the selective and sparse labeling of astrocytes in various brain regions.This technique utilizes a dual adeno-associated virus system that allows for the expression of Cre recombinase and enhanced green fluorescent protein under the control of the glial fibrillary acidic protein(GfaABC1D)promoter.The system was tested in C57BL/6J mice and successfully labeled astrocytes across multiple brain regions.The method enabled the detailed visualization of individual astrocytes-including their intricate peripheral processes-through three-dimensional reconstructions from confocal microscopy images.Furthermore,the labeling efficiency of this dual adeno-associated virus technology was validated by examining astrocyte function in a spared nerve injury model and through chemogenetic modulation.This innovative approach holds great promise for future research because it enables a more comprehensive understanding of astrocyte variation not only in spared nerve injury but also in a broad spectrum of neurological diseases.The ability to selectively label and study astrocytes in different brain regions provides a powerful tool for exploring the complexities of these essential cells and their roles in physiological and pathological conditions.展开更多
基金Supported by grants from the Innovation of Science and Technology Talents in Harbin(No.2017RAXQJ045)the Fundamental Research Funds for the Central Universities
文摘Objective To verify the neurotypicality of AAV-PHP.eB after tail vein injection in adult mice and its efficiency in crossing the blood-brain barrier(BBB).Methods The rAAV-SYN-GFP plasmid was constructed, and adult C57 BL mice were injected with AAV PHP.eB: SYN-GFP in the tail vein(300 nL, virus titer 3 × 10~9 vg) and in the prefrontal lobe(50 L, virus tite5 × 10^(11) vg). The green fluorescent protein(GFP) signal in the brain was observed at two weeks, while the GFP signal in the peripheral organs was observed at four weeks. Results Two weeks after tail vein injection, GFP expression was observed throughout the brain especially in the cortex, hippocampus, and geniculate nucleus. No GFP signal was observed or detected by western blotting in the peripheral organs after four weeks. GFP signal was observed mainly at the loca site after prefrontal lobe injection.Conclusion AAV-PHP.eB: SYN-GFP can effectively cross the BBB in adult mice. Using a neuron-specific promoter allows exogenous gene expression in neurons; therefore, AAV-PHP.eB can be used as an effective carrier for studying diseases in the central nervous system(CNS).
基金National Natural Science Foundation of China,No.32271148(to JW)the National Key Research and the Development Program of China,No.2023M740625(to ML)+1 种基金the Natural Science Foundation of Guangdong Province,Nos.2021B1515120050(to HW)and 2023A1515110782(to ML)and Key R&D Program of Ningxia Hui Autonomous Region,No.2024BEG02027(to JW).
文摘Astrocytes are the most abundant glial cells in the central nervous system.They perform a diverse array of functions,with a critical role in structural integrity,synapse formation,and neurotransmission.These cells exhibit substantial regional heterogeneity and display variable responses to different neurological diseases.Such diversity in astrocyte morphology and function is essential for understanding both normal brain function and the underlying mechanisms of neurological disorders.To investigate this heterogeneity,we developed a novel method for the selective and sparse labeling of astrocytes in various brain regions.This technique utilizes a dual adeno-associated virus system that allows for the expression of Cre recombinase and enhanced green fluorescent protein under the control of the glial fibrillary acidic protein(GfaABC1D)promoter.The system was tested in C57BL/6J mice and successfully labeled astrocytes across multiple brain regions.The method enabled the detailed visualization of individual astrocytes-including their intricate peripheral processes-through three-dimensional reconstructions from confocal microscopy images.Furthermore,the labeling efficiency of this dual adeno-associated virus technology was validated by examining astrocyte function in a spared nerve injury model and through chemogenetic modulation.This innovative approach holds great promise for future research because it enables a more comprehensive understanding of astrocyte variation not only in spared nerve injury but also in a broad spectrum of neurological diseases.The ability to selectively label and study astrocytes in different brain regions provides a powerful tool for exploring the complexities of these essential cells and their roles in physiological and pathological conditions.
