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Behavioral alterations of zebrafish larvae after early embryonic exposure to ketamine

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Abstract

Rationale

Ketamine has been associated with pediatric risks that include neurocognitive impairment and long-term behavioral disorders. However, the neurobehavioral effects of ketamine exposure in early development remain uncertain.

Objectives

This study aimed to test stage- and dose-dependent effects of ketamine exposure on certain brain functions by evaluating alterations in locomotion, anxiety-like and avoidance behaviors, as well as socialization.

Methods

Embryos were exposed to different concentrations of ketamine (0, 0.2, 0.4, and 0.8 mg mL−1) for 20 min during the 256-cell (2.5 h post fertilization—hpf), 50% epiboly (5.5 hpf), and 1–4 somites (10.5 hpf) stages. General exploratory activities, natural escape-like responses, and social interactions were analyzed under continuous light or under a moving light stimulus.

Results

A dose-dependent decrease in the overall mean speed was perceived in the embryos exposed during the 256-cell stage. These results were related to previously observed head and eye malformations, following ketamine exposure at this stage and may indicate possible neurobehavioral disorders when ketamine exposure is performed at this stage. Results also showed that ketamine exposure during the 50% epiboly and 1–4 somites stages induced a significant increment of the anxiety-like behavior and a decrease in avoidance behavior in all exposed groups.

Conclusions

Overall, the results validate the neurodevelopmental risks of early-life exposure to ketamine.

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References

  • Aizawa H (2013) Habenula and the asymmetric development of the vertebrate brain. Anat Sci Int 88:1–9

    Article  PubMed  Google Scholar 

  • Ben-Ari Y, Spitzer NC (2010) Phenotypic checkpoints regulate neuronal development. Trends Neurosci 33:485–492

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Brambrink AM, Evers AS, Avidan MS, Farber NB, Smith DJ, Martin LD, Dissen GA, Creeley CE, Olney JW (2012) Ketamine-induced neuroapoptosis in the fetal and neonatal rhesus macaque brain. Anesthesiology 116:372–384

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Burgess HA, Granato M (2007) Sensorimotor gating in larval zebrafish. J Neurosci 27:4984–4994

    Article  CAS  PubMed  Google Scholar 

  • Buske C, Gerlai R (2011a) Early embryonic ethanol exposure impairs shoaling and the dopaminergic and serotoninergic systems in adult zebrafish. Neurotoxicol Teratol 33:698–707

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Buske C, Gerlai R (2011b) Shoaling develops with age in zebrafish (Danio rerio). Prog Neuro-Psychopharmacol Biol Psychiatry 35:1409–1415

    Article  Google Scholar 

  • Buske C, Gerlai R (2014) Diving deeper into zebrafish development of social behavior: analyzing high resolution data. J Neurosci Methods 234:66–72

    Article  PubMed  Google Scholar 

  • Chatterjee M, Guo Q, Weber S, Scholpp S, Li JY (2014) Pax6 regulates the formation of the habenular nuclei by controlling the temporospatial expression of Shh in the diencephalon in vertebrates. BMC Biol 12:13

    Article  PubMed  PubMed Central  Google Scholar 

  • Clift DE, Thorn RJ, Passarelli EA, Kapoor M, LoPiccolo MK, Richendrfer HA, Colwill RM, Creton R (2015) Effects of embryonic cyclosporine exposures on brain development and behavior. Behav Brain Res 282:117–124

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Colwill RM, Creton R (2011) Locomotor behaviors in zebrafish (Danio rerio) larvae. Behav Process 86:222–229

    Article  Google Scholar 

  • Cox JA, Kucenas S, Voigt MM (2005) Molecular characterization and embryonic expression of the family of N-methyl-D-aspartate receptor subunit genes in the zebrafish. Dev Dyn 234:756–766

    Article  CAS  PubMed  Google Scholar 

  • Creton R (2009) Automated analysis of behavior in zebrafish larvae. Behav Brain Res 203:127–136

    Article  PubMed  Google Scholar 

  • Creton R, Speksnijder JE, Jaffe LF (1998) Patterns of free calcium in zebrafish embryos. J Cell Sci 111(Pt 12):1613–1622

    CAS  PubMed  Google Scholar 

  • Cuevas E, Trickler WJ, Guo X, Ali SF, Paule MG, Kanungo J (2013) Acetyl L-carnitine protects motor neurons and Rohon-Beard sensory neurons against ketamine-induced neurotoxicity in zebrafish embryos. Neurotoxicol Teratol 39:69–76

    Article  CAS  PubMed  Google Scholar 

  • Danos N, Lauder GV (2007) The ontogeny of fin function during routine turns in zebrafish Danio rerio. J Exp Biol 210:3374–3386

    Article  PubMed  Google Scholar 

  • Marron Fdez de Velasco E, Law PY, Rodriguez RE (2009) Mu opioid receptor from the zebrafish exhibits functional characteristics as those of mammalian mu opioid receptor. Zebrafish 6:259–268

    Article  PubMed  Google Scholar 

  • Dong C, Anand KJ (2013) Developmental neurotoxicity of ketamine in pediatric clinical use. Toxicol Lett 220:53–60

    Article  CAS  PubMed  Google Scholar 

  • Felix LM, Antunes LM, Coimbra AM (2014) Ketamine NMDA receptor-independent toxicity during zebrafish (Danio rerio) embryonic development. Neurotoxicol Teratol 41:27–34

    Article  CAS  PubMed  Google Scholar 

  • Felix LM, Serafim C, Valentim AM, Antunes LM, Campos S, Matos M, Coimbra AM (2016) Embryonic stage-dependent teratogenicity of ketamine in zebrafish (Danio rerio). Chem Res Toxicol 29:1298–1309

    Article  CAS  PubMed  Google Scholar 

  • Granato M, van Eeden FJ, Schach U, Trowe T, Brand M, Furutani-Seiki M, Haffter P, Hammerschmidt M, Heisenberg CP, Jiang YJ, Kane DA, Kelsh RN, Mullins MC, Odenthal J, Nusslein-Volhard C (1996) Genes controlling and mediating locomotion behavior of the zebrafish embryo and larva. Development 123:399–413

    CAS  PubMed  Google Scholar 

  • Halluin C, Madelaine R, Naye F, Peers B, Roussigne M, Blader P (2016) Habenular neurogenesis in zebrafish is regulated by a hedgehog, Pax6 Proneural Gene Cascade. PLoS One 11:e0158210

    Article  PubMed  PubMed Central  Google Scholar 

  • Huang L, Liu Y, Zhang P, Kang R, Liu Y, Li X, Bo L, Dong Z (2013) In vitro dose-dependent inhibition of the intracellular spontaneous calcium oscillations in developing hippocampal neurons by ketamine. PLoS One 8:e59804

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Hwang J, Kim HS, Seok JW, Kim JD, Koun S, Park SY, Lee J, Kim HS, Kim HS, Kim KS, Chang KT, Ryoo ZY, Wang SM, Huh TL, Lee S (2009) Transcriptome analysis of the zebrafish mind bomb mutant. Mol Gen Genomics 281:77–85

    Article  CAS  Google Scholar 

  • Ikonomidou C, Bosch F, Miksa M, Bittigau P, Vockler J, Dikranian K, Tenkova TI, Stefovska V, Turski L, Olney JW (1999) Blockade of NMDA receptors and apoptotic neurodegeneration in the developing brain. Science 283:70–74

    Article  CAS  PubMed  Google Scholar 

  • Kalueff AV, Gebhardt M, Stewart AM, Cachat JM, Brimmer M, Chawla JS, Craddock C, Kyzar EJ, Roth A, Landsman S, Gaikwad S, Robinson K, Baatrup E, Tierney K, Shamchuk A, Norton W, Miller N, Nicolson T, Braubach O, Gilman CP, Pittman J, Rosemberg DB, Gerlai R, Echevarria D, Lamb E, Neuhauss SC, Weng W, Bally-Cuif L, Schneider H, Zebrafish Neuroscience Research C (2013) Towards a comprehensive catalog of zebrafish behavior 1.0 and beyond. Zebrafish 10:70–86

    Article  PubMed  PubMed Central  Google Scholar 

  • Kalueff AV, Stewart AM, Gerlai R (2014) Zebrafish as an emerging model for studying complex brain disorders. Trends Pharmacol Sci 35:63–75

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Kanungo J, Cuevas E, Ali SF, Paule MG (2013) Ketamine induces motor neuron toxicity and alters neurogenic and proneural gene expression in zebrafish. J Appl Toxicol 33:410–417

    Article  CAS  PubMed  Google Scholar 

  • Kim E, Clark AL, Kiss A, Hahn JW, Wesselschmidt R, Coscia CJ, Belcheva MM (2006) Mu- and kappa-opioids induce the differentiation of embryonic stem cells to neural progenitors. J Biol Chem 281:33749–33760

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Kimmel CB, Ballard WW, Kimmel SR, Ullmann B, Schilling TF (1995) Stages of embryonic development of the zebrafish. Dev Dyn 203:253–310

    Article  CAS  PubMed  Google Scholar 

  • Klee EW, Schneider H, Clark KJ, Cousin MA, Ebbert JO, Hooten WM, Karpyak VM, Warner DO, Ekker SC (2012) Zebrafish: a model for the study of addiction genetics. Hum Genet 131:977–1008

    Article  CAS  PubMed  Google Scholar 

  • Kohrs R, Durieux ME (1998) Ketamine: teaching an old drug new tricks. Anesth Analg 87:1186–1193

    CAS  PubMed  Google Scholar 

  • Kurdi MS, Theerth KA, Deva RS (2014) Ketamine: current applications in anesthesia, pain, and critical care. Anesth Essays Res 8:283–290

    Article  PubMed  PubMed Central  Google Scholar 

  • Leclerc C, Neant I, Moreau M (2012) The calcium: an early signal that initiates the formation of the nervous system during embryogenesis. Front Mol Neurosci 5:3

    Article  PubMed  PubMed Central  Google Scholar 

  • Lee A, Mathuru AS, Teh C, Kibat C, Korzh V, Penney TB, Jesuthasan S (2010) The habenula prevents helpless behavior in larval zebrafish. Curr Biol 20:2211–2216

    Article  CAS  PubMed  Google Scholar 

  • Mahabir S, Chatterjee D, Buske C, Gerlai R (2013) Maturation of shoaling in two zebrafish strains: a behavioral and neurochemical analysis. Behav Brain Res 247:1–8

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Mathuru AS, Jesuthasan S (2013) The medial habenula as a regulator of anxiety in adult zebrafish. Front Neural Circuits 7:99

    Article  PubMed  PubMed Central  Google Scholar 

  • Mellon RD, Simone AF, Rappaport BA (2007) Use of anesthetic agents in neonates and young children. Anesth Analg 104:509–520

    Article  CAS  PubMed  Google Scholar 

  • Miller N, Gerlai R (2012) Automated tracking of zebrafish shoals and the analysis of shoaling behavior. In: Kalueff VA, Stewart MA (eds) Zebrafish protocols for neurobehavioral research. Humana Press, Totowa, NJ, pp. 217–230

    Chapter  Google Scholar 

  • Mion G, Villevieille T (2013) Ketamine pharmacology: an update (pharmacodynamics and molecular aspects, recent findings). CNS Neurosci Ther 19:370–380

    Article  CAS  PubMed  Google Scholar 

  • Morgan CJ, Curran HV, Independent Scientific Committee on D (2012) Ketamine use: a review. Addiction 107:27–38

    Article  PubMed  Google Scholar 

  • Narita M, Kuzumaki N, Miyatake M, Sato F, Wachi H, Seyama Y, Suzuki T (2006) Role of delta-opioid receptor function in neurogenesis and neuroprotection. J Neurochem 97:1494–1505

    Article  CAS  PubMed  Google Scholar 

  • Ozil JP, Banrezes B, Toth S, Pan H, Schultz RM (2006) Ca2+ oscillatory pattern in fertilized mouse eggs affects gene expression and development to term. Dev Biol 300:534–544

    Article  CAS  PubMed  Google Scholar 

  • Padilla S, Hunter DL, Padnos B, Frady S, MacPhail RC (2011) Assessing locomotor activity in larval zebrafish: influence of extrinsic and intrinsic variables. Neurotoxicol Teratol 33:624–630

    Article  CAS  PubMed  Google Scholar 

  • Palanisamy A (2012) Maternal anesthesia and fetal neurodevelopment. Int J Obstet Anesth 21:152–162

    Article  CAS  PubMed  Google Scholar 

  • Patthey C, Gunhaga L (2014) Signaling pathways regulating ectodermal cell fate choices. Exp Cell Res 321:11–16

    Article  CAS  PubMed  Google Scholar 

  • Paule MG, Li M, Allen RR, Liu F, Zou X, Hotchkiss C, Hanig JP, Patterson TA, Slikker W Jr, Wang C (2011) Ketamine anesthesia during the first week of life can cause long-lasting cognitive deficits in rhesus monkeys. Neurotoxicol Teratol 33:220–230

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Pelkowski SD, Kapoor M, Richendrfer HA, Wang X, Colwill RM, Creton R (2011) A novel high-throughput imaging system for automated analyses of avoidance behavior in zebrafish larvae. Behav Brain Res 223:135–144

    Article  PubMed  PubMed Central  Google Scholar 

  • Perrine SA, Hoshaw BA, Unterwald EM (2006) Delta opioid receptor ligands modulate anxiety-like behaviors in the rat. Br J Pharmacol 147:864–872

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Pietri T, Roman AC, Guyon N, Romano SA, Washbourne P, Moens CB, de Polavieja GG, Sumbre G (2013) The first n2-null zebrafish model shows altered motor behaviors. Front Neural Circuits 7:118

    Article  PubMed  PubMed Central  Google Scholar 

  • Pobbe RL, Zangrossi H Jr (2008) Involvement of the lateral habenula in the regulation of generalized anxiety- and panic-related defensive responses in rats. Life Sci 82:1256–1261

    Article  CAS  PubMed  Google Scholar 

  • Reif DM, Truong L, Mandrell D, Marvel S, Zhang G, Tanguay RL (2016) High-throughput characterization of chemical-associated embryonic behavioral changes predicts teratogenic outcomes. Arch Toxicol 90:1459–1470

    Article  CAS  PubMed  Google Scholar 

  • Richendrfer H, Creton R (2013) Automated high-throughput behavioral analyses in zebrafish larvae. J Vis Exp: e50622.

  • Richendrfer H, Pelkowski SD, Colwill RM, Creton R (2012) On the edge: pharmacological evidence for anxiety-related behavior in zebrafish larvae. Behav Brain Res 228:99–106

    Article  CAS  PubMed  Google Scholar 

  • Riehl R, Kyzar E, Allain A, Green J, Hook M, Monnig L, Rhymes K, Roth A, Pham M, Razavi R, Dileo J, Gaikwad S, Hart P, Kalueff AV (2011) Behavioral and physiological effects of acute ketamine exposure in adult zebrafish. Neurotoxicol Teratol 33:658–667

    Article  CAS  PubMed  Google Scholar 

  • Rofael HZ, Abdel-Rahman MS (2002) The role of ketamine on plasma cocaine pharmacokinetics in rat. Toxicol Lett 129:167–176

    Article  CAS  PubMed  Google Scholar 

  • Root CM, Velazquez-Ulloa NA, Monsalve GC, Minakova E, Spitzer NC (2008) Embryonically expressed GABA and glutamate drive electrical activity regulating neurotransmitter specification. J Neurosci 28:4777–4784

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Sanchez-Simon FM, Rodriguez RE (2008) Developmental expression and distribution of opioid receptors in zebrafish. Neuroscience 151:129–137

    Article  CAS  PubMed  Google Scholar 

  • Scallet AC, Schmued LC, Slikker W Jr, Grunberg N, Faustino PJ, Davis H, Lester D, Pine PS, Sistare F, Hanig JP (2004) Developmental neurotoxicity of ketamine: morphometric confirmation, exposure parameters, and multiple fluorescent labeling of apoptotic neurons. Toxicol Sci 81:364–370

    Article  CAS  PubMed  Google Scholar 

  • Schmidt R, Strahle U, Scholpp S (2013) Neurogenesis in zebrafish—from embryo to adult. Neural Dev 8:3

    Article  PubMed  PubMed Central  Google Scholar 

  • Schnorr SJ, Steenbergen PJ, Richardson MK, Champagne DL (2012) Measuring thigmotaxis in larval zebrafish. Behav Brain Res 228:367–374

    Article  CAS  PubMed  Google Scholar 

  • Solnica-Krezel L, Stemple DL, Mountcastle-Shah E, Rangini Z, Neuhauss SC, Malicki J, Schier AF, Stainier DY, Zwartkruis F, Abdelilah S, Driever W (1996) Mutations affecting cell fates and cellular rearrangements during gastrulation in zebrafish. Development 123:67–80

    CAS  PubMed  Google Scholar 

  • Sprung J, Flick RP, Wilder RT, Katusic SK, Pike TL, Dingli M, Gleich SJ, Schroeder DR, Barbaresi WJ, Hanson AC, Warner DO (2009) Anesthesia for cesarean delivery and learning disabilities in a population-based birth cohort. Anesthesiology 111:302–310

    Article  PubMed  PubMed Central  Google Scholar 

  • Steele SL, Ekker M, Perry SF (2011) Interactive effects of development and hypoxia on catecholamine synthesis and cardiac function in zebrafish (Danio rerio). J Comp Physiol B 181:527–538

    CAS  PubMed  Google Scholar 

  • Stewart AM, Braubach O, Spitsbergen J, Gerlai R, Kalueff AV (2014) Zebrafish models for translational neuroscience research: from tank to bedside. Trends Neurosci 37:264–278

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Su PH, Chang YZ, Chen JY (2010) Infant with in utero ketamine exposure: quantitative measurement of residual dosage in hair. Pediatr Neonatol 51:279–284

    Article  PubMed  Google Scholar 

  • Suen MF, Chan WS, Hung KW, Chen YF, Mo ZX, Yung KK (2013) Assessments of the effects of nicotine and ketamine using tyrosine hydroxylase-green fluorescent protein transgenic zebrafish as biosensors. Biosens Bioelectron 42:177–185

    Article  CAS  PubMed  Google Scholar 

  • Toth AB, Shum AK, Prakriya M (2016) Regulation of neurogenesis by calcium signaling. Cell Calcium 59:124–134

    Article  CAS  PubMed  Google Scholar 

  • Tufi S, Leonards P, Lamoree M, de Boer J, Legler J, Legradi J (2016) Changes in neurotransmitter profiles during early zebrafish (Danio rerio) development and after pesticide exposure. Environ Sci Technol 50:3222–3230

    Article  CAS  PubMed  Google Scholar 

  • Varga ZM (2011) Aquaculture and husbandry at the zebrafish international resource center. Methods Cell Biol 104:453–478

    Article  PubMed  Google Scholar 

  • Viberg H, Ponten E, Eriksson P, Gordh T, Fredriksson A (2008) Neonatal ketamine exposure results in changes in biochemical substrates of neuronal growth and synaptogenesis, and alters adult behavior irreversibly. Toxicology 249:153–159

    Article  CAS  PubMed  Google Scholar 

  • Webb SE, Miller AL (2000) Calcium signalling during zebrafish embryonic development. BioEssays 22:113–123

    Article  CAS  PubMed  Google Scholar 

  • Westerfield M (2007) The zebrafish book: a guide for the laboratory use of zebrafish (Danio rerio), 5th edn. University of Oregon press.

  • Wilder RT, Flick RP, Sprung J, Katusic SK, Barbaresi WJ, Mickelson C, Gleich SJ, Schroeder DR, Weaver AL, Warner DO (2009) Early exposure to anesthesia and learning disabilities in a population-based birth cohort. Anesthesiology 110:796–804

    Article  PubMed  PubMed Central  Google Scholar 

  • Wolman MA, Jain RA, Liss L, Granato M (2011) Chemical modulation of memory formation in larval zebrafish. Proc Natl Acad Sci U S A 108:15468–15473

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Yan J, Li YR, Zhang Y, Lu Y, Jiang H (2014) Repeated exposure to anesthetic ketamine can negatively impact neurodevelopment in infants: a prospective preliminary clinical study. J Child Neurol 29:1333–1338

    Article  PubMed  Google Scholar 

  • Zakhary SM, Ayubcha D, Ansari F, Kamran K, Karim M, Leheste JR, Horowitz JM, Torres G (2011) A behavioral and molecular analysis of ketamine in zebrafish. Synapse 65:160–167

    Article  CAS  PubMed  PubMed Central  Google Scholar 

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Acknowledgements

This work was supported by FEDER funds through the Operational Competitiveness Programme—COMPETE and by the Operational Competitiveness and Internationalization Programme—POCI and by National Funds through FCT—Fundação para a Ciência e a Tecnologia under the project FCOMP-01-0124-FEDER-028683 (PTDC/CVT-WEL/4672/2012), and POCI-01-0145-FEDER-006958 (UID/AGR/04033/2013). Financial support for Ana M. Valentim was provided by postdoctoral fellowship SFRH/BPD/103006/2014 issued by FCT.

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Authors and Affiliations

  1. Centre for the Research and Technology of Agro-Environmental and Biological Sciences (CITAB), University of Trás-os-Montes and Alto Douro (UTAD), Vila Real, Portugal

    Luís M. Félix, Luís M. Antunes, Ana M. Coimbra & Ana M. Valentim

  2. Institute for Research and Innovation in Health (i3S), University of Porto (UP), Porto, Portugal

    Luís M. Félix, Luís M. Antunes & Ana M. Valentim

  3. Laboratory Animal Science (LAS), Institute for Molecular and Cell Biology (IBMC), University of Porto (UP), Porto, Portugal

    Luís M. Félix, Luís M. Antunes & Ana M. Valentim

  4. School of Agrarian and Veterinary Sciences (ECAV), University of Trás-os-Montes and Alto Douro (UTAD), Vila Real, Portugal

    Luís M. Antunes

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  1. Luís M. Félix
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Correspondence to Luís M. Félix.

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Félix, L.M., Antunes, L.M., Coimbra, A.M. et al. Behavioral alterations of zebrafish larvae after early embryonic exposure to ketamine. Psychopharmacology 234, 549–558 (2017). https://doi.org/10.1007/s00213-016-4491-7

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  • DOI: https://doi.org/10.1007/s00213-016-4491-7

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