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JoVE Journal
Neuroscience
Pomiar wychwytu glukozy w modelach proteinopatii TDP-43 u drosophila
Pomiar wychwytu glukozy w modelach proteinopatii TDP-43 u drosophila
JoVE Journal
Neuroscience
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JoVE Journal Neuroscience
Measuring Glucose Uptake in Drosophila Models of TDP-43 Proteinopathy

Pomiar wychwytu glukozy w modelach proteinopatii TDP-43 u drosophila

Full Text
3,203 Views
07:07 min
August 3, 2021

DOI: 10.3791/62936-v

Suvithanandhini Loganathan*1, Hannah E. Ball*1, Ernesto Manzo1,2, Daniela C. Zarnescu1,3

1Department of Molecular and Cellular Biology,University of Arizona, Tucson, 2Vollum Institute,Oregon Health and Science University, 3Department of Neuroscience,University of Arizona, Tucson

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

Overview

This study investigates glucose uptake in Drosophila motor neurons affected by TAR DNA binding protein (TDP-43) proteinopathy using a FRET-based, genetically encoded glucose sensor. The technique allows for real-time measurement of intracellular glucose levels, highlighting metabolic changes in neurons related to neurodegenerative conditions.

Key Study Components

Area of Science

  • Neuroscience
  • Cell Biology
  • Neurodegeneration

Background

  • TDP-43 is associated with neurodegenerative diseases such as ALS.
  • Understanding glucose metabolism is critical for insights into neuronal health.
  • This study utilizes a FRET-based sensor for dynamic measurements in living tissues.
  • Previous methods may not have allowed for real-time analysis within specific neuron types.

Purpose of Study

  • To measure changes in glucose uptake in motor neurons under TDP-43 proteinopathy.
  • To compare glucose metabolism between ALS models and control neurons.
  • To utilize a live imaging approach for direct observations of metabolic changes.

Methods Used

  • Live imaging of Drosophila motor neurons using a FRET-based glucose sensor.
  • Dissections were performed on wandering third instar larvae to prepare neuronal samples.
  • Image acquisition utilized a confocal microscope to capture changes every 10 seconds.
  • FRET efficiency was calculated to determine glucose uptake in response to stimulation.

Main Results

  • Control motor neurons exhibited a small increase in glucose uptake, while TDP-43 mutant neurons showed a significantly higher increase.
  • FRET by CFP ratios indicated alterations in intracellular glucose concentrations upon stimulation.
  • No baseline differences were observed between genotypes, emphasizing the impact of glucose stimulation.
  • Results suggest critical differences in metabolic responses between healthy and impaired neurons.

Conclusions

  • This study demonstrates the utility of FRET-based sensors in assessing metabolic changes in live neurons.
  • The findings provide insights into glucose metabolism's role in neuronal health and disease.
  • Implications for understanding metabolic dysregulation in neurodegenerative diseases are significant.

Frequently Asked Questions

What are the advantages of using Drosophila for this study?
Drosophila serve as a powerful model for studying neurodegeneration due to their genetic manipulability and physiological similarities to humans.
How is the FRET-based glucose sensor used in live tissues?
The FRET-based sensor reports changes in glucose levels in real-time by measuring the fluorescence resonance energy transfer signals in specific neurons.
What outcomes can be measured using this technique?
This technique allows for the measurement of intracellular glucose concentrations and changes in metabolism within living motor neurons.
How does the imaging process work in this study?
The larvae are dissected and immobilized, followed by imaging with a confocal microscope to capture glucose uptake changes sequentially.
What are the limitations of this study?
A limitation includes potential variations in neuronal health over time which could affect imaging results and interpretations.

Wychwyt glukozy jest zwiększony w neuronach ruchowych Drosophila dotkniętych proteinopatią białka wiążącego DNA TAR (TDP-43), na co wskazuje genetycznie kodowany czujnik glukozy oparty na FRET.

Technika ta jest istotna, ponieważ może mierzyć wychwyt glukozy w określonych komórkach w żywych tkankach Drosophila. Czujnik oparty na fret zapewnia bezpośredni pomiar zmian wewnątrzkomórkowego poziomu glukozy. Technika ta pozwala naukowcom oceniać zmiany poziomu glukozy w czasie rzeczywistym, w żywych neuronach ruchowych na podstawie modeli ALS w porównaniu z grupą kontrolną.

Wyzwaniem jest obrazowanie tego samego zestawu neuronów po zmianie bufora. Pozostawienie próbki na mikroskopie pomaga zapewnić, że VMC porusza się tak mało, jak to możliwe. Przed rozpoczęciem sekcji włącz mikroskop obrazowy i lasery.

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Wychwyt glukozy modele Drosophila proteinopatia TDP-43 czujnik oparty na FRET wewnątrzkomórkowe poziomy glukozy obrazowanie na żywo neurony ruchowe modele ALS rozwarstwienie larw mikroskop konfokalny technika obrazowania układ nerwowo-mięśniowy selekcja optyczna stymulacja glukozy

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