Protocol for in vivo analysis of pre- and post-synaptic protein function in mice

TL;DR

This paper introduces a protocol to study pre- and post-synaptic proteins in mice brains using AAV gene transfer and immunofluorescence techniques.

Contribution

A novel protocol combining AAV-mediated gene transfer and ACSF perfusion for in vivo synaptic protein analysis in mice.

Findings

AAV gene transfer allows manipulation of specific neuron subtypes in mice.

ACSF perfusion and immunofluorescence improve visualization of synaptic proteins in brain sections.

The protocol enables in vivo analysis of synaptic protein function.

Abstract

Studying synapses in vivo presents challenges due to the complexity of accurately targeting and visualizing specific synaptic proteins within the brain. Here, we present a protocol for in vivo analysis of pre- and post-synaptic protein function in mice. We describe steps for combining adeno-associated virus (AAV)-mediated gene transfer to manipulate specific neuron subtypes. We also describe immunofluorescence on artificial cerebrospinal fluid (ACSF)-perfused brain sections to enhance the visualization of synaptic proteins.

For complete details on the use and execution of this protocol, please refer to Cramer et al.1

•ACSF perfusion technique for investigating synaptic proteins in vivo•Steps for AAV gene transfer for broad expression or in specific neurons•Instructions for immunofluorescent labeling of free-floating brain sections•Guidance on improved detection of synaptic proteins using microscopy

ACSF perfusion technique for investigating synaptic proteins in vivo

Steps for AAV gene transfer for broad expression or in specific neurons

Instructions for immunofluorescent labeling of free-floating brain sections

Guidance on improved detection of synaptic proteins using microscopy

Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Studying synapses in vivo presents challenges due to the complexity of accurately targeting and visualizing specific synaptic proteins within the brain. Here, we present a protocol for in vivo analysis of pre- and post-synaptic protein function in mice. We describe steps for combining adeno-associated virus (AAV)-mediated gene transfer to manipulate specific neuron subtypes. We also describe immunofluorescence on artificial cerebrospinal fluid (ACSF)-perfused brain sections to enhance the visualization of synaptic proteins.

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