Front Cell Neurosci. 2020 Jun 5
Di Chen, Keke Ren, Haiying Liu, Honghui Mao, Zongyan Li, Huiming Mo, Shengjun Xie, Yiwu Shi, Qian Chen, and Wenting Wang
Products used in the paper Details Operation
AAV vector packaging The AAV was packaged by PackGene Biotech in China. Mice were given retro-ocular injection of AAV as previously described. Briefly, mice were anesthetized with 3% isoflurane. Then, 100 ml of AAVPhP:eB-DIO-EGFP-P2A-EGFPf with a titer of 1.3E + 12 gc/ml (PackGene Biotech, LLC, China) was injected into the retro-orbital sinus with a 27-G needle and a 1-ml syringe. Request Quote

Research Field: CNS

Keywords: sparse neuron labeling, neuronal morphology reconstruction, dendritic spine, autism, CaMKIIα, Shank3

AAV Serotype: AAVPhP:eB

Dose: 100 ml of AAVPhP:eB-DIO-EGFP-P2A-EGFPf with a titer of 1.3E + 12 gc/ml (PackGene Biotech, LLC, China) was injected into the retro-orbital sinus with a 27-G needle and a 1-ml syringe.

Routes of Administration: retro-ocular injection

Targeted organ: brain

Animal or cell line strain: The mice were then placed in a warm and moist environment to wait for resuscitation. After resuscitation, the mice were put back in the cage and housed for 3 weeks before the next step.

Abstract

Single neurons, as the basic unit of the brain, consist of a cell body and processes, including dendrites and axons. Even neurons of the same type show various subtle process characteristics to fit into the diverse neural circuits. Different cell types of neurons form complicated circuits in the brain. Therefore, detailed neuronal morphology is required to understand normal neuronal function and pathological mechanisms, such as those that occur in autism. Here, we developed a strategy to sparsely label the same type of neurons throughout the whole brain and tested its application in an autistic animal model—Shank3 knockout (KO) mice. To achieve this, we designed an adeno-associated virus (AAV) that expresses Cre recombinase-dependent regular and membrane-targeted enhanced green fluorescent protein (EGFP) under a human synapsin 1 promoter and verified it in several Cre transgenic mice. We could sparsely label the projection neurons in multiple brain areas by retro-ocular injection of the virus into CaMKIIα-Cre mice. Then, we analyzed the morphology of the projection neurons in Shank3 KO mice with this method. We found differential dendritic complexity and dendritic spine changes in projection neurons in Shank3 KO mice crossed with CaMKIIα-Cre mice compared with littermate control mice in the striatum, cortex, and hippocampus. By combining this method with various Cre mouse lines crossed with mouse models of disease, we can screen the morphological traits of distinct types of neurons throughout the whole brain that will help us to understand the exact role of the specific cell types of neurons not only in autism spectrum disorder (ASD) mouse models but also in other psychiatric disorder mouse models.

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