<em>Ex Vivo</em> Optogenetic Interrogation of Long-Range Synaptic Transmission and Plasticity from Medial Prefrontal Cortex to Lateral Entorhinal Cortex

Lisa Kinnavane; Paul J. Banks

doi:10.3791/63077

Ex Vivo Optogenetic Interrogation of Long-Range Synaptic Transmission and Plasticity fro...

JoVE Journal
Neuroscience

This content is Free Access.

JoVE Journal Neuroscience

Ex Vivo Optogenetic Interrogation of Long-Range Synaptic Transmission and Plasticity from Medial Prefrontal Cortex to Lateral Entorhinal Cortex

Ex Vivo Optogenetic Interrogation of Long-Range Synaptic Transmission and Plasticity from Medial Prefrontal Cortex to Lateral Entorhinal Cortex

Full Text

2,928 Views

11:31 min

February 25, 2022

DOI: 10.3791/63077-v

Lisa Kinnavane¹, Paul J. Banks¹

¹School of Physiology, Pharmacology and Neuroscience,University of Bristol

Overview

This study presents a detailed protocol for viral transduction of specific brain regions using optogenetic constructs, allowing for synapse-specific electrophysiological characterization in acute rodent brain slices. The method facilitates the investigation of long-range pathways and selectively stimulates axons that are not anatomically separated, enhancing our understanding of synaptic physiology and plasticity.

Key Study Components

Area of Science

Neuroscience
Electrophysiology
Optogenetics

Background

Viral delivery of optogenetic constructs enables precise control over neuronal activity.
This method allows for the selective stimulation of specific synaptic connections.
Exploiting optogenetics enhances the study of synaptic physiology in acute brain slices.
Detailed procedures for brain slicing and neuron patch clamping are discussed.

Purpose of Study

To establish a protocol for studying the electrophysiological properties of specific synapses.
To demonstrate the advantages of using optogenetic tools in neuronal research.
To provide a clear methodology for acute brain slice preparations and recordings.

Methods Used

The study utilizes acute brain slices from rodents as the main platform.
Optogenetic constructs are virally delivered to specific brain regions for targeted synapse activation.
Key procedural steps include viral injection, brain slicing, and whole cell patch clamping of neurons.
Timelines include a two-week period for viral expression before slice preparation.
Electrophysiological recordings are performed to assess synaptic properties following illumination.

Main Results

The method allows for selective stimulation of synapses, facilitating detailed analysis of electrophysiological properties.
Changes in excitability and synaptic responses can be directly measured.
The protocol establishes effective approaches for investigating synaptic mechanisms and neuronal plasticity.
Key insights into the functioning of specific synaptic pathways are demonstrated.

Conclusions

This study provides a robust framework for investigating synaptic behavior using optogenetic methods.
The approach enhances understanding of neuronal mechanisms and their implications for neural circuit function.
Future applications could advance the knowledge of synaptic plasticity in various models of neurological conditions.

Frequently Asked Questions

What are the advantages of using optogenetics in this study?

Optogenetics allows for precise temporal control of synapse activation, enabling researchers to study specific pathways and synaptic mechanisms that traditional methods cannot achieve.

How is the viral delivery method implemented?

The viral delivery is performed using a Hamilton syringe connected to a microinjection syringe pump to infuse the viral construct into predefined brain areas after making a burr hole in the skull.

What types of data outcomes are obtained from this methodology?

The methodology provides electrophysiological data on synaptic responses, including capacitance and resistance changes, as well as insights into neuronal excitability.

Can this protocol be adapted for other models?

Yes, despite focusing on rodents, the protocol could potentially be adapted for other species or models depending on the specific research questions being addressed.

What are some limitations of this method?

Some limitations include the need for precise anatomical targeting and potential variability in viral expression across different subjects or brain regions.

How essential is the slicing technique in this protocol?

The slicing technique is critical as it allows for the preservation of synaptic architecture and ensures that electrophysiological recordings accurately reflect how neurons function in vivo.

Here we present a protocol describing viral transduction of discrete brain regions with optogenetic constructs to permit synapse-specific electrophysiological characterization in acute rodent brain slices.

Virally delivered optogenetic constructs permit detailed electrophysiological characterization of the physiology and plasticity of specific synapses in acute brain slices. The primary advantages of activating synapses optogenetically is the ability to study long-range pathways, and the selective stimulation of axons which are not anatomically separated. Demonstrating the procedure will be Dr.Lisa Kinnavane, a research associate from our laboratory.

To begin, load a Hamilton syringe into a microinjection syringe pump attached to a movable arm mounted to a stereotaxic frame. Then place a five-microliter aliquot of the virus in a microcentrifuge. Spin the tube for a few seconds and pipette two microliters of the viral preparation into the tube's lid.

View the full transcript and gain access to thousands of scientific videos