{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,12,11]],"date-time":"2025-12-11T11:51:30Z","timestamp":1765453890430,"version":"3.46.0"},"reference-count":54,"publisher":"MDPI AG","issue":"24","license":[{"start":{"date-parts":[[2025,12,11]],"date-time":"2025-12-11T00:00:00Z","timestamp":1765411200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"Natural Sciences and Engineering Research Council (NSERC) of Canada","award":["1021837"],"award-info":[{"award-number":["1021837"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["IJMS"],"abstract":"<jats:p>Arginine vasotocin (AVT) has recently emerged as a local regulator of testicular function in fish. Using ex vivo culture system, it was demonstrated that AVT directly stimulates androgen-dependent basal spermatogenesis in zebrafish. In the presence of gonadotropins, AVT enhanced FSH-induced development of early phases of spermatogonial proliferation while blocking FSH-mediated spermiogenesis. However, AVT promoted proliferation of LH-induced pre-meiotic and meiotic germ cell populations without affecting the final stages of spermiogenesis. These findings led to the hypothesis that AVT plays a role by promoting early germ cell proliferation and differentiation while simultaneously inhibiting premature progression through spermiogenesis. To test this hypothesis, we investigated the chronic effects of AVT on adult zebrafish testes, in vivo. Prolonged AVT treatment for 21 days led to dose-dependent accumulation of undifferentiated type A spermatogonia and reduced post-meiotic germ cells and spermatozoa. We also observed decreased plasma 11-ketotestosterone (11-KT) levels and downregulation of fshr. This was accompanied by a basal suppression of avt and its receptors, avpr1aa, avpr1ab, avpr2aa, avpr2ab, avpr2l, in both brain and testis during the pre-spawning phase. The present findings, along with those of previously published studies, collectively demonstrate that AVT presence during the early stages of testicular development promotes spermatogonia proliferation while diminishing FSH-induced premature progress toward spermatogenesis. This occurs until later stages, when AVT expression is diminished, allowing for optimal LH-induced spermiogenesis in zebrafish.<\/jats:p>","DOI":"10.3390\/ijms262411938","type":"journal-article","created":{"date-parts":[[2025,12,11]],"date-time":"2025-12-11T11:17:27Z","timestamp":1765451847000},"page":"11938","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Integrated Role of Arginine Vasotocin in the Control of Spermatogenesis in Zebrafish"],"prefix":"10.3390","volume":"26","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-1635-6269","authenticated-orcid":false,"given":"Maya","family":"Zanardini","sequence":"first","affiliation":[{"name":"Department of Biological Sciences, University of Calgary, 2500 University Drive NW, Calgary, AB T2N 1N4, Canada"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-4368-6853","authenticated-orcid":false,"given":"Hamid R.","family":"Habibi","sequence":"additional","affiliation":[{"name":"Department of Biological Sciences, University of Calgary, 2500 University Drive NW, Calgary, AB T2N 1N4, Canada"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2025,12,11]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"1588","DOI":"10.1210\/en.2004-0818","article-title":"Gronadotropin-releasing hormone induction of apoptosis in the testes of goldfish (Caraasius auratus)","volume":"146","author":"Buret","year":"2005","journal-title":"Endocrinology"},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"204","DOI":"10.1038\/s42003-021-01708-y","article-title":"Insulin-like 3 affects zebrafish spermatogenic cells directly and via Sertoli cells","volume":"4","author":"Crespo","year":"2021","journal-title":"Commun. Biol."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"336","DOI":"10.1016\/j.bbrc.2007.12.136","article-title":"Discovery of a gonad-specific IGF subtype in teleost","volume":"367","author":"Wang","year":"2008","journal-title":"Biochem. Biophys. Res. Commun."},{"key":"ref_4","doi-asserted-by":"crossref","unstructured":"Xie, X., N\u00f3brega, R., and P\u0161eni\u010dka, M. (2020). Spermatogonial stem cells in fish: Characterization, isolation, enrichment, and recent advances of in vitro culture systems. Biomolecules, 10.","DOI":"10.3390\/biom10040644"},{"key":"ref_5","doi-asserted-by":"crossref","unstructured":"Doretto, L.B., Butzge, A.J., Nakajima, R.T., Martinez, E.R., de Souza, B.M., Rodrigues, M.D.S., Rosa, I.F., Ricci, J.M., Tovo-Neto, A., and Costa, D.F. (2022). Gdnf Acts as a Germ Cell-Derived Growth Factor and Regulates the Zebrafish Germ Stem Cell Niche in Autocrine- and Paracrine-Dependent Manners. Cells, 11.","DOI":"10.3390\/cells11081295"},{"key":"ref_6","doi-asserted-by":"crossref","unstructured":"Habibi, H.R., and Andreu-Vieyra, C.V. (2007). Hormonal regulation of follicular atresia in teleost fish. The Fish Oocyte: From Basic Studies to Biotechnological Applications, Springer.","DOI":"10.1007\/978-1-4020-6235-3_9"},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"68","DOI":"10.4161\/org.4.2.5851","article-title":"Wnt Secretion and Extra-Cellular Regulators","volume":"4","author":"Komiya","year":"2008","journal-title":"Organogenesis"},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"3804","DOI":"10.1210\/en.2015-1157","article-title":"Fsh stimulates spermatogonial proliferation and differentiation in zebrafish via Igf3","volume":"156","author":"Morais","year":"2015","journal-title":"Endocrinology"},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"75","DOI":"10.1016\/S0303-7207(99)00250-6","article-title":"Direct action of GnRH variants on goldfish oocyte meiosis and follicular steroidogenesis","volume":"160","author":"Pati","year":"2000","journal-title":"Mol. Cell. Endocrinol."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1095\/biolreprod.115.137547","article-title":"Conserved anti-m\u00fcllerian hormone: Anti-m\u00fcllerian hormone type-2 receptor specific interaction and intracellular signaling in teleosts","volume":"94","author":"Rocha","year":"2016","journal-title":"Biol. Reprod."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"dev178665","DOI":"10.1242\/dev.178665","article-title":"Endocrine and local signaling interact to regulate spermatogenesis in zebrafish: Follicle-stimulating hormone, retinoic acid and androgens","volume":"146","author":"Crespo","year":"2019","journal-title":"Development"},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"3527","DOI":"10.1210\/en.2010-1469","article-title":"Proteolytically activated, recombinant anti-M\u00fcllerian hormone inhibits androgen secretion, proliferation, and differentiation of spermatogonia in adult zebrafish testis organ cultures","volume":"152","author":"KSkaar","year":"2011","journal-title":"Endocrinology"},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"110440","DOI":"10.1016\/j.mce.2019.04.020","article-title":"Paracrine\/autocrine control of spermatogenesis by gonadotropin-inhibitory hormone","volume":"492","author":"Fallah","year":"2019","journal-title":"Mol. Cell. Endocrinol."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"111087","DOI":"10.1016\/j.mce.2020.111087","article-title":"Effects of gonadotropin-inhibitory hormone on early and late stages of spermatogenesis in ex-vivo culture of zebrafish testis","volume":"520","author":"Fallah","year":"2021","journal-title":"Mol. Cell. Endocrinol."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"bqaa004","DOI":"10.1210\/endocr\/bqaa004","article-title":"Role of GnRH Isoforms in Paracrine\/Autocrine Control of Zebrafish (Danio rerio) Spermatogenesis","volume":"161","author":"Fallah","year":"2020","journal-title":"Endocrinology"},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"54","DOI":"10.1016\/j.ygcen.2015.02.013","article-title":"Vasotocin\u2014A new player in the control of oocyte maturation and ovulation in fish","volume":"221","author":"Joy","year":"2015","journal-title":"Gen. Comp. Endocrinol."},{"key":"ref_17","doi-asserted-by":"crossref","unstructured":"Mennigen, J.A., Ramachandran, D., Shaw, K., Chaube, R., Joy, K.P., and Trudeau, V.L. (2022). Reproductive roles of the vasopressin\/oxytocin neuropeptide family in teleost fishes. Front. Endocrinol., 13.","DOI":"10.3389\/fendo.2022.1005863"},{"key":"ref_18","doi-asserted-by":"crossref","unstructured":"Zanardini, M., Zhang, W., and Habibi, H.R. (2024). Arginine Vasotocin Directly Regulates Spermatogenesis in Adult Zebrafish (Danio rerio) Testes. Int. J. Mol. Sci., 25.","DOI":"10.3390\/ijms25126564"},{"key":"ref_19","doi-asserted-by":"crossref","unstructured":"Ramachandran, D., Sharma, K., Saxena, V., Nipu, N., Rajapaksha, D.C., and Mennigen, J.A. (2023). Knock-out of vasotocin reduces reproductive success in female zebrafish, Danio rerio. Front. Endocrinol., 14.","DOI":"10.3389\/fendo.2023.1151299"},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"51","DOI":"10.1016\/j.theriogenology.2021.03.001","article-title":"Vasotocin stimulates maturation-inducing hormone, oocyte maturation and ovulation in the catfish Heteropneustes fossilis: Evidence for a preferential calcium involvement","volume":"167","author":"Singh","year":"2021","journal-title":"Theriogenology"},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"172","DOI":"10.1016\/j.ygcen.2009.03.016","article-title":"Effects of hCG and ovarian steroid hormones on vasotocin levels in the female catfish Heteropneustes fossilis","volume":"162","author":"Singh","year":"2009","journal-title":"Gen. Comp. Endocrinol."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"504","DOI":"10.1016\/j.ygcen.2010.02.020","article-title":"An involvement of vasotocin in oocyte hydration in the catfish Heteropneustes fossilis: A comparison with effects of isotocin and hCG","volume":"166","author":"Singh","year":"2010","journal-title":"Gen. Comp. Endocrinol."},{"key":"ref_23","doi-asserted-by":"crossref","unstructured":"Ferr\u00e9, A., Chauvign\u00e9, F., Gozdowska, M., Kulczykowska, E., Finn, R.N., and Cerd\u00e0, J. (2023). Neurohypophysial and paracrine vasopressinergic signaling regulates aquaporin trafficking to hydrate marine teleost oocytes. Front. Endocrinol., 14.","DOI":"10.3389\/fendo.2023.1222724"},{"key":"ref_24","doi-asserted-by":"crossref","unstructured":"Zanardini, M., and Habibi, H.R. (2025). The Role of Neurohypophysial Hormones in the Endocrine and Paracrine Control of Gametogenesis in Fish. Cells, 14.","DOI":"10.3390\/cells14141061"},{"key":"ref_25","doi-asserted-by":"crossref","unstructured":"Kelly, A.M., and Goodson, J.L. (2013). Behavioral relevance of species-specific vasotocin anatomy in gregarious finches. Front. Neurosci., 7.","DOI":"10.3389\/fnins.2013.00242"},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"3","DOI":"10.1016\/S0079-6123(08)00401-9","article-title":"Nonapeptides and the evolutionary patterning of sociality","volume":"170","author":"Goodson","year":"2008","journal-title":"Prog. Brain Res."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"282","DOI":"10.1159\/000316867","article-title":"Individual behavioral and neuronal phenotypes for arginine vasotocin mediated courtship and aggression in a territorial teleost","volume":"75","author":"Santangelo","year":"2010","journal-title":"Brain Behav. Evol."},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"230","DOI":"10.1016\/j.yhbeh.2011.12.016","article-title":"Nonapeptides and Social Behavior in Fishes","volume":"61","author":"Godwin","year":"2012","journal-title":"Horm. Behav."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"314","DOI":"10.1016\/j.physbeh.2014.11.052","article-title":"Arginine vasotocin reduces levels of cooperative behaviour in a cleaner fish","volume":"139","author":"Cardoso","year":"2015","journal-title":"Physiol. Behav."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"110333","DOI":"10.1016\/j.cbpb.2019.110333","article-title":"A reproductive role for the nonapeptides vasotocin and isotocin in male zebrafish (Danio rerio)","volume":"238","author":"Altmieme","year":"2019","journal-title":"Comp. Biochem. Physiol. Part B Biochem. Mol. Biol."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"221","DOI":"10.1016\/j.ygcen.2012.08.015","article-title":"Arginine-vasotocin expression and participation in reproduction and social behavior in males of the cichlid fish Cichlasoma dimerus","volume":"179","author":"Ramallo","year":"2012","journal-title":"Gen. Comp. Endocrinol."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"142","DOI":"10.1111\/jne.12239","article-title":"Arginine Vasotocin, Isotocin and Nonapeptide Receptor Gene Expression Link to Social Status and Aggression in Sex-Dependent Patterns","volume":"27","author":"Lema","year":"2015","journal-title":"J. Neuroendocrinol."},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"330","DOI":"10.1016\/j.cbpb.2005.12.004","article-title":"Arginine vasotocin (AVT) and isotocin (IT) in fish brain: Diurnal and seasonal variations","volume":"143","author":"Gozdowska","year":"2006","journal-title":"Comp. Biochem. Physiol. Part B Biochem. Mol. Biol."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"3394","DOI":"10.1210\/en.2011-2022","article-title":"Time-of-Day-Dependent Changes in GnRH1 Neuronal Activities and Gonadotropin mRNA Expression in a Daily Spawning Fish, Medaka","volume":"153","author":"Karigo","year":"2012","journal-title":"Endocrinology"},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"9","DOI":"10.1016\/j.ygcen.2005.12.022","article-title":"Arginine vasotocin a key hormone in fish physiology and behaviour: A review with insights from mammalian models","volume":"147","author":"Balment","year":"2006","journal-title":"Gen. Comp. Endocrinol."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"541","DOI":"10.1016\/j.ygcen.2010.11.013","article-title":"Diurnal rhythms in hypothalamic\/pituitary AVT synthesis and secretion in rainbow trout: Evidence for a circadian regulation","volume":"170","author":"Soengas","year":"2011","journal-title":"Gen. Comp. Endocrinol."},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"105677","DOI":"10.1016\/j.yhbeh.2025.105677","article-title":"Dominance and aggressiveness are associated with vasotocin neuron numbers in a cooperatively breeding cichlid fish","volume":"168","author":"Ruberto","year":"2024","journal-title":"Horm. Behav."},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"e22500","DOI":"10.1002\/jbt.22500","article-title":"Molecular characterization and expression of arginine vasotocin V1a2 receptor in Atlantic croaker brain: Potential mechanisms of its downregulation by PCB77","volume":"34","author":"Rahman","year":"2020","journal-title":"J. Biochem. Mol. Toxicol."},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"2393","DOI":"10.1098\/rspb.2008.0622","article-title":"Expression of arginine vasotocin in distinct preoptic regions is associated with dominant and subordinate behaviour in an African cichlid fish","volume":"275","author":"Greenwood","year":"2008","journal-title":"Proc. R. Soc. B Biol. Sci."},{"key":"ref_40","doi-asserted-by":"crossref","unstructured":"Zanardini, M., Parker, N., Ma, Y., and Habibi, H.R. (2025). Arginine vasotocin is a player in the multifactorial control of spermatogenesis in zebrafish. Front. Endocrinol., 16.","DOI":"10.3389\/fendo.2025.1672823"},{"key":"ref_41","doi-asserted-by":"crossref","unstructured":"Soko\u0142owska, E., Gozdowska, M., and Kulczykowska, E. (2020). Nonapeptides Arginine Vasotocin and Isotocin in Fishes: Advantage of Bioactive Molecules Measurement. Front. Mar. Sci., 7.","DOI":"10.3389\/fmars.2020.00610"},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"885","DOI":"10.1007\/s10695-018-0590-1","article-title":"In situ localization of vasotocin receptor gene transcripts in the brain-pituitary-gonadal axis of the catfish Heteropneustes fossilis: A morpho-functional study","volume":"45","author":"Rawat","year":"2019","journal-title":"Fish Physiol. Biochem."},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"747","DOI":"10.1038\/s41586-020-03040-7","article-title":"Universal nomenclature for oxytocin\u2013vasotocin ligand and receptor families","volume":"592","author":"Theofanopoulou","year":"2021","journal-title":"Nature"},{"key":"ref_44","doi-asserted-by":"crossref","unstructured":"Kwon, W.S., Park, Y.J., Kim, Y.H., You, Y.A., Kim, I.C., and Pang, M.G. (2013). Vasopressin Effectively Suppresses Male Fertility. PLoS ONE, 8.","DOI":"10.1371\/journal.pone.0054192"},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"e202201488","DOI":"10.26508\/lsa.202201488","article-title":"Vasopressin V1a receptor and oxytocin receptor regulate murine sperm motility differently","volume":"6","author":"Tsuchiya","year":"2023","journal-title":"Life Sci. Alliance"},{"key":"ref_46","first-page":"3865","article-title":"Blood Collection for Biochemical Analysis in Adult Zebrafish","volume":"26","author":"Pedroso","year":"2012","journal-title":"J. Vis. Exp."},{"key":"ref_47","unstructured":"Westerfield, M. (2000). The zebrafish book. A Guide for the Laboratory Use of Zebrafish (Danio rerio), University of Oregon Press. [4th ed.]."},{"key":"ref_48","first-page":"e4196","article-title":"Regular Care and Maintenance of a Zebrafish Laboratory: An Introduction","volume":"69","author":"Avdesh","year":"2012","journal-title":"J. Vis. Exp."},{"key":"ref_49","doi-asserted-by":"crossref","first-page":"40","DOI":"10.1089\/154585404774101662","article-title":"Characterization and Development of Courtship in Zebrafish, Danio rerio","volume":"1","author":"Darrow","year":"2004","journal-title":"Zebrafish"},{"key":"ref_50","doi-asserted-by":"crossref","first-page":"676","DOI":"10.1038\/nmeth.2019","article-title":"Fiji: An open-source platform for biological-image analysis","volume":"9","author":"Schindelin","year":"2019","journal-title":"Nat. Methods"},{"key":"ref_51","doi-asserted-by":"crossref","first-page":"59","DOI":"10.1038\/nprot.2007.514","article-title":"High-resolution in situ hybridization to whole-mount zebrafish embryos","volume":"3","author":"Thisse","year":"2008","journal-title":"Nat. Protoc."},{"key":"ref_52","doi-asserted-by":"crossref","first-page":"112494","DOI":"10.1016\/j.mce.2025.112494","article-title":"Brain monoamine changes modulate the corticotropin-releasing hormone receptor 1-mediated behavioural response to acute thermal stress in zebrafish larvae","volume":"600","author":"Rajeswari","year":"2025","journal-title":"Mol. Cell. Endocrinol."},{"key":"ref_53","doi-asserted-by":"crossref","unstructured":"Shaw, K., Lu, C., Liu, X., and Trudeau, V.L. (2023). Arginine vasopressin injection rescues delayed oviposition in cyp19a1b-\/- mutant female zebrafish. Front. Endocrinol., 14.","DOI":"10.3389\/fendo.2023.1308675"},{"key":"ref_54","doi-asserted-by":"crossref","first-page":"599","DOI":"10.1016\/j.mex.2018.05.011","article-title":"Optimization of phenol-chloroform RNA extraction","volume":"5","author":"Toni","year":"2018","journal-title":"MethodsX"}],"container-title":["International Journal of Molecular Sciences"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1422-0067\/26\/24\/11938\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,12,11]],"date-time":"2025-12-11T11:20:01Z","timestamp":1765452001000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1422-0067\/26\/24\/11938"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2025,12,11]]},"references-count":54,"journal-issue":{"issue":"24","published-online":{"date-parts":[[2025,12]]}},"alternative-id":["ijms262411938"],"URL":"https:\/\/doi.org\/10.3390\/ijms262411938","relation":{},"ISSN":["1422-0067"],"issn-type":[{"value":"1422-0067","type":"electronic"}],"subject":[],"published":{"date-parts":[[2025,12,11]]}}}