Visualizing Shifts on Neuron-Glia Circuit with the Calcium Imaging Technique

Matheus H. Tempone; Hercules R. Freitas; Clarissa S. Schitine; Ricardo A. de Melo Reis

doi:10.3791/63338

Visualización de cambios en el circuito Neuron-Glia con la técnica de imágenes de calcio

JoVE Journal
Neuroscience
Author Produced

A subscription to JoVE is required to view this content. Sign in or start your free trial.

JoVE Journal Neuroscience

Visualizing Shifts on Neuron-Glia Circuit with the Calcium Imaging Technique

Visualización de cambios en el circuito Neuron-Glia con la técnica de imágenes de calcio

Full Text

5,238 Views

11:41 min

April 8, 2022

DOI: 10.3791/63338-v

Matheus H. Tempone¹, Hercules R. Freitas², Clarissa S. Schitine³, Ricardo A. de Melo Reis¹

¹Laboratory of Neurochemistry, Institute of Biophysics Carlos Chagas Filho,Universidade Federal do Rio de Janeiro, ²Laboratory of Neuroenergetics and Inborn Errors of Metabolism, Institute of Medical Biochemistry Leopoldo de Meis,Universidade Federal do Rio de Janeiro, ³Laboratory of Neurochemistry and Cell Biology, Institute of Life Sciences,Universidade Federal da Bahia

Please note that some of the translations on this page are AI generated. Click here for the English version.

Overview

This study focuses on cell calcium imaging as a method to monitor cytosolic calcium concentration changes in live cells. Using a chicken retina culture, the work differentiates responses of neurons and glial cells to potassium chloride and ATP stimuli, highlighting the involvement of calcium dynamics in signaling processes.

Key Study Components

Area of Science

Neuroscience
Cell Biology
Imaging Techniques

Background

Calcium is a crucial second messenger in various cellular processes.
High selective fluorescent calcium dyes enable advanced imaging techniques.
The response of different cell types to specific stimuli can provide insight into their functional roles.
This procedure allows for the investigation of neuronal and glial signaling in cultured cells.

Purpose of Study

To monitor changes in cytosolic calcium concentrations in live cells.
To differentiate the calcium responses of neurons and glia based on specific stimuli.
To elucidate the functional roles of these cell types in calcium signaling.

Methods Used

The platform used is a cell culture of chicken retina cells.
This study primarily focuses on enriched neuron cultures and glial cells.
The method involves the use of Fura-2 AM for calcium imaging.
Cells are treated with potassium chloride and ATP to measure intracellular calcium shifts.
Data is analyzed using Metafluor software and presented in Excel tables.

Main Results

The findings reveal distinct calcium responses in neurons when stimulated with potassium chloride.
Glial cells primarily respond to ATP stimuli, underscoring their role in calcium signaling.
The study provides insights into the spatial and temporal dynamics of calcium signaling in mixed cultures.
Results indicate that the method effectively differentiates cellular responses based on treatment.

Conclusions

This study demonstrates the efficacy of calcium imaging in understanding cell-specific responses.
The methodology enables detailed tracking of calcium dynamics, enhancing understanding of neuronal mechanisms.
These insights can inform future research on plasticity and cellular communication in the nervous system.

Frequently Asked Questions

What are the advantages of using chicken retina cells?

Chicken retina cells are accessible for dissection and provide a well-defined model for studying neuronal and glial interactions in a controlled environment.

How are neuronal and glial responses differentiated?

Responses are differentiated based on their distinct reaction to potassium chloride and ATP, enabling specific insights into calcium dynamics in each cell type.

What types of data are obtained from calcium imaging?

Calcium imaging provides quantifiable data on intracellular calcium concentration changes, allowing analysis of cell excitability and signaling mechanisms.

Can this method be adapted for other cell types?

Yes, the procedure can be adapted to other neuronal or glial cultures based on the specific calcium imaging needs and targeted stimuli.

What are some key considerations when performing this procedure?

It is essential to maintain sterile conditions, accurately prepare reagents, and carefully monitor incubation times to ensure reliable results.

La imagen de calcio celular es una metodología versátil para estudiar la señalización dinámica de células individuales, en poblaciones mixtas en cultivo o incluso en animales despiertos, basada en la expresión de canales / receptores permeables al calcio que dan firmas funcionales únicas.

El objetivo general de este procedimiento es monitorear los cambios en las concentraciones de calcio citosólico en las células vivas para diferenciar las respuestas neuronales o glías basadas en cloruro de potasio o estímulos de ATP. El calcio es un segundo mensajero importante involucrado en varios procesos celulares, incluyendo la neurotransmisión, la plasticidad y la apoptosis. El advenimiento de un colorante de calcio fluorescente de alta selección, como Fura-2 AM, asociado con el desarrollo de mejores microscopios fluorescentes y métodos de computación, produjo datos ópticos de alta resolución con un alto grado de resolución espacial y temporal para obtener imágenes de la señalización de calcio en células y organismos vivos.

Este protocolo es adaptable a neuronas enriquecidas, glía purificada o cultivos de población mixta. Rastrea qué tipos de células respondieron a estímulos determinados en función de la respuesta diferencial al cloruro de potasio y ATP. El cloruro de potasio cambia el potencial de membrana de la célula.

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