Assaying Circuit Specific Regulation of Adult Hippocampal Neural Precursor Cells

Luis J. Quintanilla; Chia-Yu Yeh; Hechen Bao; Christina Catavero; Juan Song

doi:10.3791/59237

Assaying Circuit Spezifische Regulation von adulthippocampal neuralen Vorläuferzellen

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Neuroscience

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JoVE Journal Neuroscience

Assaying Circuit Specific Regulation of Adult Hippocampal Neural Precursor Cells

Assaying Circuit Spezifische Regulation von adulthippocampal neuralen Vorläuferzellen

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08:52 min

July 24, 2019

DOI: 10.3791/59237-v

Luis J. Quintanilla^1,2,3, Chia-Yu Yeh^1,2, Hechen Bao^1,2, Christina Catavero^1,2,3, Juan Song^1,2,3

¹Department of Pharmacology,University of North Carolina Chapel Hill, ²Neuroscience Center,University of North Carolina Chapel Hill, ³Neuroscience Curriculum,University of North Carolina Chapel Hill

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Overview

This protocol describes a method for analyzing the behavior of adult neural stem/progenitor cells when subjected to chemogenetic manipulation of specific local neural circuits. The approach aims to elucidate how neural circuit stimulation or inhibition impacts adult neurogenesis.

Key Study Components

Area of Science

Neuroscience
Neurogenesis
Cell Biology

Background

Investigation of adult neural stem cell proliferation.
Understanding the regulation by specific neural circuits.
Role of neurotransmitters in adult neurogenesis.
Micro-manipulation techniques to reduce stress in animal models.

Purpose of Study

To evaluate the effect of targeted neural circuit manipulation on adult neurogenesis.
To assess the proliferation of neural stem cells.
To determine the functionality of specific neurotransmitter-releasing cell types.

Methods Used

Use of tissue sections and immunohistochemistry for analysis.
Adult neural stem/progenitor cells manipulated via chemogenetic techniques.
No multiomics workflow is mentioned in the study.
Key steps include tissue preparation, staining, and microscopy imaging.
Quantification of cell density and neurogenesis using imaging software.

Main Results

Confirmed the effectiveness of targeted neural circuit manipulation on neural stem cell proliferation.
Demonstrated fluorescent labeling of specific cell types involved in neurogenesis.
Proliferation rates of nestin-positive cells and their morphological characteristics analyzed.
Effective quantification methods established for evaluating neurogenesis across different experimental conditions.

Conclusions

The study illustrates the impact of local circuit activity on the regulation of neural stem cells.
The methodology enhances the understanding of neurogenic processes in the adult brain.
Possible implications for therapeutic strategies targeting neurogenesis in neurological diseases.

Frequently Asked Questions

What are the advantages of this protocol?

This protocol allows for precise manipulation of neural circuits while minimizing stress in animal subjects, enhancing the study’s reliability.

How is the neural stem cell proliferation measured?

Proliferation is assessed through fluorescence microscopy, identifying cells labeled with thymidine analogs and counting their density.

What type of tissue is used in this study?

Adult brain tissue sections are used to analyze the behavior of neural stem/progenitor cells under specific conditions.

How can this method be adapted for other studies?

The methodology can be tailored to investigate other neurotransmitter systems or different neural pathways associated with neurogenesis.

What are potential limitations of this approach?

The results may be influenced by the specific conditions set during circuit manipulation or the accessibility of target cells in the tissue sections.

What data outcomes can be obtained from this protocol?

Outcomes include cell density measurements, identification of proliferating progenitor cells, and insights into the regulatory effects of neurotransmitters on neurogenesis.

Das Ziel dieses Protokolls ist es, einen Ansatz zur Analyse des Verhaltens von adulten neuronalen Stamm-/Vorläuferzellen als Reaktion auf die chemogenetische Manipulation eines bestimmten lokalen neuronalen Schaltkreises zu beschreiben.

Dieses Protokoll kann beantworten, wie verschiedene Schaltkreise im Gehirn die adulte Neurogenese regulieren und insbesondere, wie stimulierende oder hemmende neuronale Schaltkreise die Proliferation adulter neuronaler Stammzellen beeinflussen. Der Vorteil dieser Technik ist, dass sie in der Lage ist, gezielt gewünschte neuronale Schaltkreise anzusprechen und die Belastung der Tiere zu reduzieren. Diese Methode kann Einen einblick in die Art und Weise geben, wie verschiedene Gehirnkreise die Adult Neurogenese regulieren, insbesondere, wie bestimmte Zelltypen, die bestimmte Neurotransmitter freisetzen, die Proliferation adulter neuronaler Stammzellen regulieren.

Um dieses Verfahren zu beginnen, legen Sie die Gewebeabschnitte in PBS und montieren Sie fünf bis acht Abschnitte auf einem positiv geladenen Dia in serieller Reihenfolge von vorder nach hinten. Lassen Sie die Gewebeabschnitte bei Raumtemperatur für zwei bis fünf Minuten trocknen und halten Sie sich vollständig an den Dias. Bereiten Sie als Nächstes den Citratpuffer in einem Container vor.

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