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Valutazione dell'omeostasi dopaminergici in topi tramite l'uso di analisi di cromatografia liquid...
Valutazione dell'omeostasi dopaminergici in topi tramite l'uso di analisi di cromatografia liquid...
JoVE Journal
Neuroscience
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JoVE Journal Neuroscience
Assessment of Dopaminergic Homeostasis in Mice by Use of High-performance Liquid Chromatography Analysis and Synaptosomal Dopamine Uptake

Valutazione dell'omeostasi dopaminergici in topi tramite l'uso di analisi di cromatografia liquida ad alte prestazioni e assorbimento della dopamina sinaptosomiali

Full Text
13,056 Views
11:26 min
September 21, 2017

DOI: 10.3791/56093-v

Kathrine L. Jensen1, Annika H. Runegaard1, Pia Weikop2, Ulrik Gether1, Mattias Rickhag1

1Molecular Neuropharmacology and Genetics Laboratory, Lundbeck Foundation Center for Biomembranes in Nanomedicine, Department of Neuroscience and Pharmacology, Faculty of Health and Medical Sciences,University of Copenhagen, 2Laboratory of Neuropsychiatry, Psychiatric Center Copenhagen and Department of Neuroscience and Pharmacology,University of Copenhagen

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Overview

This study presents two techniques to investigate dopamine homeostasis in mice: the assessment of dopamine transporter function through synaptosomal dopamine uptake and the analysis of dopamine levels in striatal tissue using high-performance liquid chromatography (HPLC). These methods allow for the evaluation of dopamine tissue content and transporter functionality, especially after in-vivo manipulations.

Key Study Components

Area of Science

  • Neuroscience
  • Dopamine Transporter Function
  • High-Performance Liquid Chromatography (HPLC)

Background

  • Dopamine plays a crucial role in various neural functions.
  • The dopamine transporter is critical for dopamine reuptake and homeostasis.
  • Understanding dopamine dynamics can provide insights into neurological disorders.
  • HPLC is a reliable method for quantifying neurotransmitter levels.

Purpose of Study

  • To provide protocols for assessing dopamine transporter function.
  • To measure dopamine content in the striatum of mice.
  • To facilitate the understanding of dopamine homeostasis post-manipulation.

Methods Used

  • The study utilizes synaptosomal preparations and HPLC for analysis.
  • Mice serve as the biological model, particularly focusing on their striatal tissue.
  • The protocols can be executed following chemogenetic manipulations or drug treatments.
  • Detailed steps are provided for brain dissection, tissue preparation, and analysis.

Main Results

  • Both methods enable robust measurement of dopamine levels and transporter activity.
  • The study provides insights into the parameters of dopamine homeostasis in a controlled setting.

Conclusions

  • The protocols established in this study enhance the understanding of dopamine transport dynamics.
  • They enable researchers to assess the impact of various manipulations on dopamine neurotransmission.

Frequently Asked Questions

What advantages does synaptosomal preparation offer?
Synaptosomal preparation allows for the isolation of synaptic terminals, providing a focused assessment of neurotransmitter dynamics without interference from other cell components.
How are dopamine levels measured in the study?
Dopamine levels are quantified using high-performance liquid chromatography (HPLC), which allows for precise analysis of neurotransmitter concentrations in tissue samples.
What interventions can affect dopamine homeostasis?
The methods implemented can be applied following chemogenetic manipulations, drug treatments, or behavioral training, influencing dopamine transporter function and levels.
What is the role of the dopamine transporter?
The dopamine transporter regulates dopamine signaling by reabsorbing dopamine from the synaptic cleft, thus playing a crucial role in maintaining neurotransmitter homeostasis.
How can this study's methods be adapted?
The techniques can be adapted for various experimental manipulations and different neuronal pathways, facilitating broader applications in neuroscience research.

Assorbimento della dopamina sinaptosomiali e strumenti sperimentali rappresentano di analisi di cromatografia liquida ad alte prestazioni per indagare l'omeostasi della dopamina nei topi valutando la funzione del trasportatore della dopamina e livelli della dopamina in tessuto striatal, rispettivamente. Qui presentiamo i protocolli per misurare il contenuto di tessuto di dopamina e di valutare la funzionalità del trasportatore della dopamina.

Stiamo introducendo due tecniche in questo protocollo. L'obiettivo del primo è quello di valutare la funzione del trasportatore della dopamina misurando l'assorbimento della dopamina dai preparati sinaptosomiali. Il secondo è per valutare i livelli di dopamina nei tessuti mediante analisi HPLC.

Questi metodi forniscono importanti parametri dell'omeostasi della dopamina, dimostrando come misurare il contenuto tissutale della dopamina e valutare la funzionalità del trasportatore della dopamina nei topi. Il vantaggio principale di queste tecniche è che possono essere eseguite dopo manipolazioni in vivo come manipolazioni chemiogenetiche, trattamenti farmacologici in vivo o addestramento comportamentale degli animali. A dimostrare la procedura HPLC sarà Pia Weikop, Professore Associato presso il Laboratorio di Neuropsichiatria.

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