Housing and husbandry factors affecting zebrafish novel tank test responses: a global multi-laboratory study

Courtney Hillman; Barbara D. Fontana; Tamara G. Amstislavskaya; Maria A. Gorbunova; Stefani Altenhofen; Karissa Barthelson; Leonardo M. Bastos; João V. Borba; Carla D. Bonan; Caroline H. Brennan; Amaury Farías-Cea; Austin Cooper; Jamie Corcoran; Eduardo R. Dondossola; Luis M. Martinez-Duran; Matheus Gallas-Lopes; David S. Galstyan; Ella O. Garcia; Ewan Gerken; Robert Hindges; Justin W. Kenney; Maxim A. Kleshchev; Tatiana O. Kolesnikova; Adele Leggieri; Sergey L. Khatsko; Michael Lardelli; Guilherme Lodetti; Giulia Lombardelli; Ana C. Luchiari; Stefani M. Portela; Violeta Medan; Lirane M. Moutinho; Evgeny V. Nekhoroshev; Barbara D. Petersen; Maureen L. Petrunich-Rutherford; Angelo Piato; Maurizio Porfiri; Emily Read; Cássio M. Resmim; Eduardo P. Rico; Denis B. Rosemberg; Murilo S. de Abreu; Catia A. Salazar; Thailana Stahlhofer-Buss; Júlia R. Teixeira; Ana M. Valentim; Alexander V. Zhdanov; Patricio Iturriaga-Vásquez; Xian Wang; Ryan Y. Wong; Allan V. Kalueff; Matthew O. Parker

doi:10.1038/s41684-025-01548-x

Housing and husbandry factors affecting zebrafish novel tank test responses: a global multi-laboratory study

Courtney Hillman, Barbara D. Fontana, Tamara G. Amstislavskaya, Maria A. Gorbunova, Stefani Altenhofen, Karissa Barthelson, Leonardo M. Bastos, João V. Borba, Carla D. Bonan, Caroline H. Brennan, Amaury Farías-Cea, Austin Cooper, Jamie Corcoran, Eduardo R. Dondossola, Luis M. Martinez-Duran, Matheus Gallas-Lopes, David S. Galstyan, Ella O. Garcia, Ewan Gerken, Robert HindgesJustin W. Kenney, Maxim A. Kleshchev, Tatiana O. Kolesnikova, Adele Leggieri, Sergey L. Khatsko, Michael Lardelli, Guilherme Lodetti, Giulia Lombardelli, Ana C. Luchiari, Stefani M. Portela, Violeta Medan, Lirane M. Moutinho, Evgeny V. Nekhoroshev, Barbara D. Petersen, Maureen L. Petrunich-Rutherford, Angelo Piato, Maurizio Porfiri, Emily Read, Cássio M. Resmim, Eduardo P. Rico, Denis B. Rosemberg, Murilo S. de Abreu, Catia A. Salazar, Thailana Stahlhofer-Buss, Júlia R. Teixeira, Ana M. Valentim, Alexander V. Zhdanov, Patricio Iturriaga-Vásquez, Xian Wang, Ryan Y. Wong, Allan V. Kalueff^*, Matthew O. Parker^*

^*Corresponding author for this work

Department of Biosciences and Bioinformatics

Research output: Contribution to journal › Article › peer-review

Abstract

The reproducibility crisis in bioscience, characterized by inconsistent study results, impedes our understanding of biological processes. Global collaborative studies offer a unique solution to this problem. Here, we present a global collaboration using the zebrafish (Danio rerio) novel tank test, a popular behavioral assay for anxiety-like responses. We analyzed data from 20 laboratories worldwide, focusing on housing conditions and experimental setups. Our study included 488 adult zebrafish, tested for 5 min, focusing on a variety of variables. Key findings show that female zebrafish exhibit more anxiety-like behavior than males, highlighting sex as a critical variable. Housing conditions, including higher stocking densities and specific feed types, also influenced anxiety levels. Optimal conditions (5 fish/L) and nutritionally rich feeds (for example, rotifers) mitigated anxiety-like behaviors. Environmental stressors, such as noise and transportation, significantly impacted behavior. We recommend standardizing testing protocols to account for sex differences, optimal stocking densities, nutritionally rich feeds and minimizing stressors to improve the reliability of zebrafish behavioral studies.

Original language	English
Article number	15081
Pages (from-to)	156-164
Number of pages	9
Journal	Lab Animal
Volume	54
Issue number	6
DOIs	https://doi.org/10.1038/s41684-025-01548-x
Publication status	Published - Jun 2025

Access to Document

10.1038/s41684-025-01548-x

Cite this

Hillman, C., Fontana, B. D., Amstislavskaya, T. G., Gorbunova, M. A., Altenhofen, S., Barthelson, K., Bastos, L. M., Borba, J. V., Bonan, C. D., Brennan, C. H., Farías-Cea, A., Cooper, A., Corcoran, J., Dondossola, E. R., Martinez-Duran, L. M., Gallas-Lopes, M., Galstyan, D. S., Garcia, E. O., Gerken, E., ... Parker, M. O. (2025). Housing and husbandry factors affecting zebrafish novel tank test responses: a global multi-laboratory study. Lab Animal, 54(6), 156-164. Article 15081. https://doi.org/10.1038/s41684-025-01548-x

@article{76d0272b4cba4de9912c9fe95222036e,

title = "Housing and husbandry factors affecting zebrafish novel tank test responses: a global multi-laboratory study",

abstract = "The reproducibility crisis in bioscience, characterized by inconsistent study results, impedes our understanding of biological processes. Global collaborative studies offer a unique solution to this problem. Here, we present a global collaboration using the zebrafish (Danio rerio) novel tank test, a popular behavioral assay for anxiety-like responses. We analyzed data from 20 laboratories worldwide, focusing on housing conditions and experimental setups. Our study included 488 adult zebrafish, tested for 5 min, focusing on a variety of variables. Key findings show that female zebrafish exhibit more anxiety-like behavior than males, highlighting sex as a critical variable. Housing conditions, including higher stocking densities and specific feed types, also influenced anxiety levels. Optimal conditions (5 fish/L) and nutritionally rich feeds (for example, rotifers) mitigated anxiety-like behaviors. Environmental stressors, such as noise and transportation, significantly impacted behavior. We recommend standardizing testing protocols to account for sex differences, optimal stocking densities, nutritionally rich feeds and minimizing stressors to improve the reliability of zebrafish behavioral studies.",

author = "Courtney Hillman and Fontana, {Barbara D.} and Amstislavskaya, {Tamara G.} and Gorbunova, {Maria A.} and Stefani Altenhofen and Karissa Barthelson and Bastos, {Leonardo M.} and Borba, {Jo{\~a}o V.} and Bonan, {Carla D.} and Brennan, {Caroline H.} and Amaury Far{\'i}as-Cea and Austin Cooper and Jamie Corcoran and Dondossola, {Eduardo R.} and Martinez-Duran, {Luis M.} and Matheus Gallas-Lopes and Galstyan, {David S.} and Garcia, {Ella O.} and Ewan Gerken and Robert Hindges and Kenney, {Justin W.} and Kleshchev, {Maxim A.} and Kolesnikova, {Tatiana O.} and Adele Leggieri and Khatsko, {Sergey L.} and Michael Lardelli and Guilherme Lodetti and Giulia Lombardelli and Luchiari, {Ana C.} and Portela, {Stefani M.} and Violeta Medan and Moutinho, {Lirane M.} and Nekhoroshev, {Evgeny V.} and Petersen, {Barbara D.} and Petrunich-Rutherford, {Maureen L.} and Angelo Piato and Maurizio Porfiri and Emily Read and Resmim, {C{\'a}ssio M.} and Rico, {Eduardo P.} and Rosemberg, {Denis B.} and {de Abreu}, {Murilo S.} and Salazar, {Catia A.} and Thailana Stahlhofer-Buss and Teixeira, {J{\'u}lia R.} and Valentim, {Ana M.} and Zhdanov, {Alexander V.} and Patricio Iturriaga-V{\'a}squez and Xian Wang and Wong, {Ryan Y.} and Kalueff, {Allan V.} and Parker, {Matthew O.}",

note = "Publisher Copyright: {\textcopyright} The Author(s) 2025.",

year = "2025",

month = jun,

doi = "10.1038/s41684-025-01548-x",

language = "English",

volume = "54",

pages = "156--164",

journal = "Lab Animal",

issn = "0093-7355",

number = "6",

}

Hillman, C, Fontana, BD, Amstislavskaya, TG, Gorbunova, MA, Altenhofen, S, Barthelson, K, Bastos, LM, Borba, JV, Bonan, CD, Brennan, CH, Farías-Cea, A, Cooper, A, Corcoran, J, Dondossola, ER, Martinez-Duran, LM, Gallas-Lopes, M, Galstyan, DS, Garcia, EO, Gerken, E, Hindges, R, Kenney, JW, Kleshchev, MA, Kolesnikova, TO, Leggieri, A, Khatsko, SL, Lardelli, M, Lodetti, G, Lombardelli, G, Luchiari, AC, Portela, SM, Medan, V, Moutinho, LM, Nekhoroshev, EV, Petersen, BD, Petrunich-Rutherford, ML, Piato, A, Porfiri, M, Read, E, Resmim, CM, Rico, EP, Rosemberg, DB, de Abreu, MS, Salazar, CA, Stahlhofer-Buss, T, Teixeira, JR, Valentim, AM, Zhdanov, AV, Iturriaga-Vásquez, P, Wang, X, Wong, RY, Kalueff, AV & Parker, MO 2025, 'Housing and husbandry factors affecting zebrafish novel tank test responses: a global multi-laboratory study', Lab Animal, vol. 54, no. 6, 15081, pp. 156-164. https://doi.org/10.1038/s41684-025-01548-x

TY - JOUR

T1 - Housing and husbandry factors affecting zebrafish novel tank test responses

T2 - a global multi-laboratory study

AU - Hillman, Courtney

AU - Fontana, Barbara D.

AU - Amstislavskaya, Tamara G.

AU - Gorbunova, Maria A.

AU - Altenhofen, Stefani

AU - Barthelson, Karissa

AU - Bastos, Leonardo M.

AU - Borba, João V.

AU - Bonan, Carla D.

AU - Brennan, Caroline H.

AU - Farías-Cea, Amaury

AU - Cooper, Austin

AU - Corcoran, Jamie

AU - Dondossola, Eduardo R.

AU - Martinez-Duran, Luis M.

AU - Gallas-Lopes, Matheus

AU - Galstyan, David S.

AU - Garcia, Ella O.

AU - Gerken, Ewan

AU - Hindges, Robert

AU - Kenney, Justin W.

AU - Kleshchev, Maxim A.

AU - Kolesnikova, Tatiana O.

AU - Leggieri, Adele

AU - Khatsko, Sergey L.

AU - Lardelli, Michael

AU - Lodetti, Guilherme

AU - Lombardelli, Giulia

AU - Luchiari, Ana C.

AU - Portela, Stefani M.

AU - Medan, Violeta

AU - Moutinho, Lirane M.

AU - Nekhoroshev, Evgeny V.

AU - Petersen, Barbara D.

AU - Petrunich-Rutherford, Maureen L.

AU - Piato, Angelo

AU - Porfiri, Maurizio

AU - Read, Emily

AU - Resmim, Cássio M.

AU - Rico, Eduardo P.

AU - Rosemberg, Denis B.

AU - de Abreu, Murilo S.

AU - Salazar, Catia A.

AU - Stahlhofer-Buss, Thailana

AU - Teixeira, Júlia R.

AU - Valentim, Ana M.

AU - Zhdanov, Alexander V.

AU - Iturriaga-Vásquez, Patricio

AU - Wang, Xian

AU - Wong, Ryan Y.

AU - Kalueff, Allan V.

AU - Parker, Matthew O.

PY - 2025/6

Y1 - 2025/6

N2 - The reproducibility crisis in bioscience, characterized by inconsistent study results, impedes our understanding of biological processes. Global collaborative studies offer a unique solution to this problem. Here, we present a global collaboration using the zebrafish (Danio rerio) novel tank test, a popular behavioral assay for anxiety-like responses. We analyzed data from 20 laboratories worldwide, focusing on housing conditions and experimental setups. Our study included 488 adult zebrafish, tested for 5 min, focusing on a variety of variables. Key findings show that female zebrafish exhibit more anxiety-like behavior than males, highlighting sex as a critical variable. Housing conditions, including higher stocking densities and specific feed types, also influenced anxiety levels. Optimal conditions (5 fish/L) and nutritionally rich feeds (for example, rotifers) mitigated anxiety-like behaviors. Environmental stressors, such as noise and transportation, significantly impacted behavior. We recommend standardizing testing protocols to account for sex differences, optimal stocking densities, nutritionally rich feeds and minimizing stressors to improve the reliability of zebrafish behavioral studies.

AB - The reproducibility crisis in bioscience, characterized by inconsistent study results, impedes our understanding of biological processes. Global collaborative studies offer a unique solution to this problem. Here, we present a global collaboration using the zebrafish (Danio rerio) novel tank test, a popular behavioral assay for anxiety-like responses. We analyzed data from 20 laboratories worldwide, focusing on housing conditions and experimental setups. Our study included 488 adult zebrafish, tested for 5 min, focusing on a variety of variables. Key findings show that female zebrafish exhibit more anxiety-like behavior than males, highlighting sex as a critical variable. Housing conditions, including higher stocking densities and specific feed types, also influenced anxiety levels. Optimal conditions (5 fish/L) and nutritionally rich feeds (for example, rotifers) mitigated anxiety-like behaviors. Environmental stressors, such as noise and transportation, significantly impacted behavior. We recommend standardizing testing protocols to account for sex differences, optimal stocking densities, nutritionally rich feeds and minimizing stressors to improve the reliability of zebrafish behavioral studies.

UR - http://www.scopus.com/inward/record.url?scp=105006488764&partnerID=8YFLogxK

U2 - 10.1038/s41684-025-01548-x

DO - 10.1038/s41684-025-01548-x

M3 - Article

C2 - 40419676

AN - SCOPUS:105006488764

SN - 0093-7355

VL - 54

SP - 156

EP - 164

JO - Lab Animal

JF - Lab Animal

IS - 6

M1 - 15081

ER -

Housing and husbandry factors affecting zebrafish novel tank test responses: a global multi-laboratory study

Abstract

Access to Document

Other files and links

Fingerprint

Cite this