دورية أكاديمية
Cell-Specific Single Viral Vector CRISPR/Cas9 Editing and Genetically Encoded Tool Delivery in the Central and Peripheral Nervous Systems.
العنوان: | Cell-Specific Single Viral Vector CRISPR/Cas9 Editing and Genetically Encoded Tool Delivery in the Central and Peripheral Nervous Systems. |
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المؤلفون: | Moffa JC; Washington University Pain Center, Department of Anesthesiology, Washington University School of Medicine, St. Louis, Missouri 63110.; Washington University Medical Scientist Training Program, Washington University School of Medicine, St. Louis, Missouri 63110., Bland IN; Washington University Pain Center, Department of Anesthesiology, Washington University School of Medicine, St. Louis, Missouri 63110., Tooley JR; Washington University Pain Center, Department of Anesthesiology, Washington University School of Medicine, St. Louis, Missouri 63110.; Washington University Division of Biological and Behavioral Sciences, Washington University School of Medicine, St. Louis, Missouri 63110., Kalyanaraman V; Washington University Pain Center, Department of Anesthesiology, Washington University School of Medicine, St. Louis, Missouri 63110., Heitmeier M; Washington University Pain Center, Department of Anesthesiology, Washington University School of Medicine, St. Louis, Missouri 63110., Creed MC; Washington University Pain Center, Department of Anesthesiology, Washington University School of Medicine, St. Louis, Missouri 63110.; Departments of Neuroscience, Psychiatry, and Biomedical Engineering, Washington University School of Medicine, St. Louis, Missouri 63110., Copits BA; Washington University Pain Center, Department of Anesthesiology, Washington University School of Medicine, St. Louis, Missouri 63110 bcopits@wustl.edu. |
المصدر: | ENeuro [eNeuro] 2024 Jul 05; Vol. 11 (7). Date of Electronic Publication: 2024 Jul 05 (Print Publication: 2024). |
نوع المنشور: | Journal Article |
اللغة: | English |
بيانات الدورية: | Publisher: Society for Neuroscience Country of Publication: United States NLM ID: 101647362 Publication Model: Electronic-Print Cited Medium: Internet ISSN: 2373-2822 (Electronic) Linking ISSN: 23732822 NLM ISO Abbreviation: eNeuro Subsets: MEDLINE |
أسماء مطبوعة: | Original Publication: [Washington, DC] : Society for Neuroscience, [2014]- |
مواضيع طبية MeSH: | CRISPR-Cas Systems* , Dependovirus*/genetics , Gene Editing*/methods , Genetic Vectors*, Animals ; Mice ; Optogenetics/methods ; Central Nervous System/metabolism ; Peripheral Nervous System/metabolism ; Male ; Mice, Inbred C57BL ; Neurons/metabolism ; Female ; Mice, Transgenic |
مستخلص: | CRISPR/Cas9 gene editing represents an exciting avenue to study genes of unknown function and can be combined with genetically encoded tools such as fluorescent proteins, channelrhodopsins, DREADDs, and various biosensors to more deeply probe the function of these genes in different cell types. However, current strategies to also manipulate or visualize edited cells are challenging due to the large size of Cas9 proteins and the limited packaging capacity of adeno-associated viruses (AAVs). To overcome these constraints, we developed an alternative gene editing strategy using a single AAV vector and mouse lines that express Cre-dependent Cas9 to achieve efficient cell-type specific editing across the nervous system. Expressing Cre-dependent Cas9 from a genomic locus affords space to package guide RNAs for gene editing together with Cre-dependent, genetically encoded tools to manipulate, map, or monitor neurons using a single virus. We validated this strategy with three common tools in neuroscience: ChRonos, a channelrhodopsin, for studying synaptic transmission using optogenetics, GCaMP8f for recording Ca 2+ transients using photometry, and mCherry for tracing axonal projections. We tested these tools in multiple brain regions and cell types, including GABAergic neurons in the nucleus accumbens, glutamatergic neurons projecting from the ventral pallidum to the lateral habenula, dopaminergic neurons in the ventral tegmental area, and proprioceptive neurons in the periphery. This flexible approach could help identify and test the function of novel genes affecting synaptic transmission, circuit activity, or morphology with a single viral injection. Competing Interests: The authors declare no competing financial interests. (Copyright © 2024 Moffa et al.) |
التعليقات: | Update of: bioRxiv. 2023 Oct 10:2023.10.10.561249. doi: 10.1101/2023.10.10.561249. (PMID: 37873336) |
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معلومات مُعتمدة: | R01 DA058755 United States DA NIDA NIH HHS; R01 NS130046 United States NS NINDS NIH HHS |
فهرسة مساهمة: | Keywords: CRISPR/Cas9; gene editing; imaging; optogenetics; photometry; tool |
تواريخ الأحداث: | Date Created: 20240613 Date Completed: 20240705 Latest Revision: 20240710 |
رمز التحديث: | 20240710 |
مُعرف محوري في PubMed: | PMC11228695 |
DOI: | 10.1523/ENEURO.0438-23.2024 |
PMID: | 38871457 |
قاعدة البيانات: | MEDLINE |
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