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 Specific Regulation of Adult Hippocampal Neural Precursor Cells

JoVE Journal
Neuroscience

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JoVE Journal Neuroscience

Assaying Circuit Specific Regulation of Adult Hippocampal Neural Precursor Cells

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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.

L'obiettivo di questo protocollo è descrivere un approccio per l'analisi del comportamento delle cellule staminali/progenitrici neurali adulte in risposta alla manipolazione chemiogenetica di uno specifico circuito neurale locale.

Questo protocollo può rispondere a come vari circuiti nel cervello regolano la neurogenesi adulta e in particolare come i circuiti neurali stimolanti o inibitori influenzano la proliferazione delle cellule staminali neurali adulte. Il vantaggio di questa tecnica è che è in grado di indirizzare specificamente i circuiti neurali desiderati, oltre a ridurre la quantità di stress introdotta agli animali. Questo metodo può fornire informazioni su come i diversi circuiti cerebrali regolano la neurogenesi adulta, in particolare su come specifici tipi di cellule che rilasciano determinati neurotrasmettitori regolano la proliferazione delle cellule staminali neurali adulte.

Per iniziare questa procedura, posizionare le sezioni tissutali in PBS e montare da cinque a otto sezioni su uno scivolo caricato positivamente in ordine seriale da anteriore a posteriore. Lasciare asciugare le sezioni tissutali a temperatura ambiente per due o cinque minuti e aderire completamente alle diapositive. Quindi, preparare il buffer di citrato in un contenitore.

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