1. Neuroscience

Generation of a CRF1-Cre transgenic rat and the role of central amygdala CRF1 cells in nociception and anxiety-like behavior

  1. Marcus M Weera  Is a corresponding author
  2. Abigail E Agoglia
  3. Eliza Douglass
  4. Zhiying Jiang
  5. Shivakumar Rajamanickam
  6. Rosetta S Shackett
  7. Melissa Herman
  8. Nicholas J Justice
  9. Nicholas W Gilpin
  1. Louisiana State University Health Sciences Center New Orleans, United States
  2. University of North Carolina at Chapel Hill, United States
  3. The University of Texas Health Science Center, United States
https://doi.org/10.7554/eLife.67822
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Cite this article
  1. Marcus M Weera
  2. Abigail E Agoglia
  3. Eliza Douglass
  4. Zhiying Jiang
  5. Shivakumar Rajamanickam
  6. Rosetta S Shackett
  7. Melissa Herman
  8. Nicholas J Justice
  9. Nicholas W Gilpin
(2022)
Generation of a CRF1-Cre transgenic rat and the role of central amygdala CRF1 cells in nociception and anxiety-like behavior
eLife 11:e67822.
https://doi.org/10.7554/eLife.67822
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Abstract

Corticotropin-releasing factor type-1 (CRF1) receptors are critical to stress responses because they allow neurons to respond to CRF released in response to stress. Our understanding of the precise role of CRF1-expressing neuronal populations in CRF-mediated behaviors has been largely limited to mouse experiments due to the lack of genetic tools available to selectively visualize and manipulate CRF1+ cells in rats. Here, we describe the generation and validation of a transgenic CRF1-Cre-tdTomato rat, which expresses a bicistronic iCre-2A-tdTomato transgene directed by 200kb of promoter and enhancer sequence surrounding the Crhr1 cDNA present within a BAC clone, that has been transgenically inserted into the rat genome. We report that Crhr1 and Cre mRNA expression are highly colocalized in CRF1-Cre-tdTomato rats within both the central amygdala (CeA), composed of mostly GABAergic neurons, and in the basolateral amygdala (BLA), composed of mostly glutamatergic neurons. In the CeA, membrane properties, inhibitory synaptic transmission, and responses to CRF bath application in tdTomato+ neurons are similar to those previously reported in GFP+ cells in CRFR1-GFP mice. We show that stimulatory DREADD receptors can be selectively targeted to CeA CRF1+ cells via virally delivered Cre-dependent transgenes, that transfected Cre/tdTomato+ cells are activated by clozapine-n-oxide in vitro and in vivo, and that activation of these cells in vivo increases anxiety-like behavior and nocifensive responses. Outside the amygdala, we show that Cre-tdTomato is expressed in several brain areas across the rostrocaudal axis of the CRF1-Cre-tdTomato rat brain, and that the expression pattern of Cre-tdTomato cells is similar to the known expression pattern of CRF1 cells. Given the accuracy of expression in the CRF1-Cre rat, modern genetic techniques used to investigate the anatomy, physiology, and behavioral function of CRF1+ neurons and circuits can now be performed in assays that require the use of rats as the model organism.

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All data generated during this study are included in the manuscript.

Article and author information

Author details

  1. Marcus M Weera

    Department of Physiology, Louisiana State University Health Sciences Center New Orleans, New Orleans, United States
    For correspondence
    mweera@lsuhsc.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-2451-0350

  2. Abigail E Agoglia

    Department of Pharmacology, University of North Carolina at Chapel Hill, Chapel Hill, United States
    Competing interests
    The authors declare that no competing interests exist.

  3. Eliza Douglass

    Department of Pharmacology, University of North Carolina at Chapel Hill, Chapel Hill, United States
    Competing interests
    The authors declare that no competing interests exist.

  4. Zhiying Jiang

    Institute Of Molecular Medicine, The University of Texas Health Science Center, Houston, United States
    Competing interests
    The authors declare that no competing interests exist.

  5. Shivakumar Rajamanickam

    Institute of Molecular Medicine, The University of Texas Health Science Center, Houston, United States
    Competing interests
    The authors declare that no competing interests exist.

  6. Rosetta S Shackett

    Department of Physiology, Louisiana State University Health Sciences Center New Orleans, New Orleans, United States
    Competing interests
    The authors declare that no competing interests exist.

  7. Melissa Herman

    Department of Pharmacology, University of North Carolina at Chapel Hill, Chapel Hill, United States
    Competing interests
    The authors declare that no competing interests exist.

  8. Nicholas J Justice

    Institute of Molecular Medicine, The University of Texas Health Science Center, Houston, United States
    Competing interests
    The authors declare that no competing interests exist.

  9. Nicholas W Gilpin

    Department of Physiology, Louisiana State University Health Sciences Center New Orleans, New Orleans, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-8901-8917

Funding

National Institute on Alcohol Abuse and Alcoholism (R01,AA023305)

  • Nicholas W Gilpin

National Institute on Alcohol Abuse and Alcoholism (R21,AA026022)

  • Melissa Herman
  • Nicholas W Gilpin

National Institute on Alcohol Abuse and Alcoholism (R00,AA023002)

  • Melissa Herman

National Institute on Alcohol Abuse and Alcoholism (National Research Service Award,AA027145)

  • Marcus M Weera

National Institute on Alcohol Abuse and Alcoholism (Institutional Training Grant,AA007577)

  • Marcus M Weera

United States Department of Veterans Affairs (Merit Award,#I01 BX003451)

  • Nicholas W Gilpin

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Ethics

Animal experimentation: All animal procedures were conducted in accordance with recommendations in the Guide for the Care and Use of Laboratory Animals of the National Institutes of Health, and were approved by the Institutional Animal Care and Use Committee of the respective institutions at which procedures occurred (Louisiana State University Health Sciences Center, University of North Carolina - Chapel Hill, University of Texas Health Sciences Center). (LSUHSC IACUC Protocol #3749; UNC IACUC Protocol #19-190; UTHSC IACUC Protocol #21-075)

Reviewing Editor

  1. Matthew N Hill, University of Calgary, Canada

Version history

  1. Preprint posted: February 23, 2021 (view preprint)
  2. Received: February 24, 2021
  3. Accepted: April 6, 2022
  4. Accepted Manuscript published: April 7, 2022 (version 1)
  5. Version of Record published: April 22, 2022 (version 2)

Copyright

This is an open-access article, free of all copyright, and may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose. The work is made available under the Creative Commons CC0 public domain dedication.

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  1. Marcus M Weera
  2. Abigail E Agoglia
  3. Eliza Douglass
  4. Zhiying Jiang
  5. Shivakumar Rajamanickam
  6. Rosetta S Shackett
  7. Melissa Herman
  8. Nicholas J Justice
  9. Nicholas W Gilpin
(2022)
Generation of a CRF1-Cre transgenic rat and the role of central amygdala CRF1 cells in nociception and anxiety-like behavior
eLife 11:e67822.
https://doi.org/10.7554/eLife.67822

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