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Targeted invalidation of CCK2 receptor gene induces anxiolytic-like action in light–dark exploration, but not in fear conditioning test

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Abstract

Rationale

Evidence suggests that γ-aminobutyric acid (GABA) and cholecystokinin (CCK) have opposite roles in the regulation of anxiety.

Objectives

The aim of our work was to study the behaviour of CCK2 receptor deficient mice in light–dark exploration and fear conditioning tests. Moreover, the action of diazepam and methyl-6,7-dimethoxy-4-ethyl-β-carboline-3-carboxylate (DMCM), having the opposite effect on GABAA receptors, was evaluated on the exploratory behaviour in these mice. Expression levels of GABAA receptor subunit genes were also measured.

Methods

Light–dark exploration and fear conditioning tests were used to determine changes in anxiety of mice. The action of diazepam (0.5–2 mg/kg i.p.) and DMCM (0.25–1 mg/kg i.p.) was studied in the light–dark box. The effect of DMCM was also evaluated in the motor activity test to demonstrate that its anti-exploratory action was not related to motor suppression. Expression levels of GABAA receptor subunit genes were determined by means of real-time polymerase chain reaction (qRT-PCR).

Results

Female mice lacking CCK2 receptors displayed increased exploratory activity in the light–dark box compared to their wild-type (+/+) littermates. Locomotor activity in the motility boxes and the intensity of freezing did not differ in wild-type (+/+) and homozygous (−/−) mice. Treatment with diazepam (0.5 mg/kg) increased the number of transitions in wild-type (+/+) animals, whereas in homozygous (−/−) mice diazepam (0.5–2 mg/kg) reduced exploratory activity. Administration of DMCM (0.25–1 mg/kg) induced an anxiogenic-like effect in homozygous (−/−) mice, but did not change their locomotor activity. Gene expression analysis established a 1.6-fold increase in the expression of the α2 subunit of GABAA receptors in the frontal cortex of homozygous (−/−) mice.

Conclusion

Genetic invalidation of CCK2 receptors induced an anxiolytic-like action in exploratory, but not in conditioned models of anxiety. The observed reduction in anxiety in homozygous (−/−) mice is probably related to an increased function of GABAergic system in the brain.

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References

  • Abramov U, Raud S, Kõks S, Innos J, Kurrikoff K, Matsui T, Vasar E (2004) Targeted mutation of CCK2 receptor gene antagonises behavioural changes induced by social isolation in female, but not in male mice. Behav Brain Res 155:1–11

    Article  PubMed  CAS  Google Scholar 

  • Becker C, Thiebot MH, Touitou Y, Hamon M, Cesselin F, Benoliel JJ (2001) Enhanced cortical extracellular levels of cholecystokinin-like material in a model of anticipation of social defeat in the rat. J Neurosci 21:262–269

    PubMed  CAS  Google Scholar 

  • Bradwejn J, de Montigny C (1984) Benzodiazepines antagonize cholecystokinin-induced activation of rat hippocampal neurones. Nature 312:363–364

    Article  PubMed  CAS  Google Scholar 

  • Chaouloff F, Durand M, Mormède P (1999) Anxiety- and activity-related effects of diazepam and chlordiazepoxide in the rat light/dark and dark/light tests. Behav Brain Res 85:27–35

    Article  Google Scholar 

  • Chesler EJ, Wilson SG, Lariviere WR, Rodriguez-Zas SL, Mogil JS (2002) Identification and ranking of genetic and laboratory environment factors influencing a behavioral trait, thermal nociception, via computational analysis of a large data archive. Neurosci Biobehav Rev 26:907–923

    Article  PubMed  Google Scholar 

  • Contet C, Rawlins JN, Deacon RM (2001) A comparison of 129S2/SvHsd and C57BL/6JOlaHsd mice on a test battery assessing sensorimotor, affective and cognitive behaviours: implications for the study of genetically modified mice. Behav Brain Res 124:33–46

    Article  PubMed  CAS  Google Scholar 

  • Cope DW, Maccaferri G, Márton LF, Roberts JDB, Cobden PM, Somogyi P (2002) Cholecystokinin-immunopositive basket and Schaffer collateral-associated interneurones target different domains of pyramidal cells in the CA1 area of the rat hippocampus. Neurosci 109:63–80

    Article  CAS  Google Scholar 

  • Cope DW, Wulff P, Oberto A, Aller MI, Capogna M, Ferraguti F, Halbsguth C, Hoeger H, Jolin HE, Jones A, McKenzie AN, Ogris W, Poeltl A, Sinkkonen ST, Vekovischeva OY, Korpi ER, Sieghart W, Sigel E, Somogyi P, Wisden W (2004) Abolition of zolpidem sensitivity in mice with a point mutation in the GABAA receptor gamma2 subunit. Neuropharmacol 47:17–34

    Article  CAS  Google Scholar 

  • Costall B, Domeney AM, Hughes J, Kelly ME, Naylor RJ, Woodruff GN (1991) Anxiolytic effects of CCK-B antagonists. Neuropeptides 19:65–73

    Article  PubMed  CAS  Google Scholar 

  • Crawley J, Goodwin FK (1980) Preliminary report of a simple animal behavior model for the anxiolytic effects of benzodiazepines. Pharmacol Biochem Behav 13:167–170

    Article  PubMed  CAS  Google Scholar 

  • Crestani F, Lorez M, Baer K, Essrich C, Benke D, Laurent JP, Belzung C, Fritschy JM, Luscher B, Mohler H (1999) Decreased GABAA-receptor clustering results in enhanced anxiety and a bias for threat cues. Nat Neurosci 2:780–782

    Article  PubMed  Google Scholar 

  • Daugé V, Beslot F, Matsui T, Roques BP (2001) Mutant mice lacking the n receptor show a dopamine-dependent hyperactivity and a behavioral sensitization to morphine. Neurosci Lett 306:41–44

    Article  PubMed  Google Scholar 

  • Davidson RJ, Irwin W (1999) The functional neuroanatomy of emotion and affective style. Trends Cogn Sci 3:11–21

    Article  PubMed  Google Scholar 

  • Dawson GR, Rupniak NM, Iversen SD, Curnow R, Tye S, Stanhope KJ, Tricklebank MD (1995) Lack of effect of CCKB receptor antagonists in ethological and conditioned animal screens for anxiolytic drugs. Psychopharmacology 121:109–117

    Article  PubMed  CAS  Google Scholar 

  • Dockstader CL, van der Kooy D (2001) Mouse strain differences in opiate reward learning are explained by differences in anxiety, not reward or learning. Neurosci 21:9077–9081

    CAS  Google Scholar 

  • Farook JM, Zhu YZ, Wang H, Moochhala S, Lee L, Wong PT (2001) Strain differences in freezing behavior of PVG hooded and Sprague–Dawley rats: differential cortical expression of cholecystokinin2 receptors. Neuroreport 12:2717–2720

    Article  PubMed  CAS  Google Scholar 

  • Fekete M, Lengyel A, Hegedüs B, Penke B, Zarandy M, Toth G, Telegdy G (1984) Further analysis of the effects of cholecystokinin octapeptides on avoidance behaviour in rats. Eur J Pharmacol 98:79–91

    Article  PubMed  CAS  Google Scholar 

  • Ferraro L, Beani L, Trist D, Reggiani A, Bianchi C (1999) Effects of cholecystokinin peptides and GV 150013, a selective cholecystokininB receptor antagonist, on electrically evoked endogenous GABA release from rat cortical slices. J Neurochem 73:1973–1981

    PubMed  CAS  Google Scholar 

  • Harro J, Vasar E, Bradwejn J (1993) CCK in animal and human research on anxiety. Trends Pharmacol Sci 14:244–249

    Article  PubMed  CAS  Google Scholar 

  • Hendry SH, Jones EG, DeFelipe J, Schmechel D, Brandon C, Emson PC (1984) Neuropeptide-containing neurons of the cerebral cortex are also GABAergic. Proc Natl Acad Sci U S A 81:6526–6530

    Article  PubMed  CAS  Google Scholar 

  • Hernandez-Gomez AM, Aguilar-Roblero R, Perez de la Mora M (2002) Role of cholecystokinin-A and cholecystokinin-B receptors in anxiety. Amino Acids 23:283–290

    Article  PubMed  CAS  Google Scholar 

  • Holmes A, Lit Q, Murphy DL, Gold E, Crawley JN (2003) Abnormal anxiety-related behavior in serotonin transporter null mutant mice: the influence of genetic background. Genes Brain Behav 2:365–380

    Article  PubMed  CAS  Google Scholar 

  • Horinouchi Y, Akiyoshi J, Nagata A, Matsushita H, Tsutsumi T, Isogawa K, Noda T, Nagayama H (2004) Reduced anxious behavior in mice lacking the CCK2 receptor gene. Eur Neuropsychopharmacol 14:157–161

    Article  PubMed  CAS  Google Scholar 

  • Hughes J, Boden P, Costall B, Domeney A, Kelly E,Horwell DC, Hunter JC, Pinnock RD, Woodruff GN (1990) Development of a class of selective cholecystokinin type B receptor antagonists having potent anxiolytic activity. Proc Natl Acad Sci U S A 87:6728–6732

    Article  PubMed  CAS  Google Scholar 

  • Kõks S, Vasar E, Soosaar A, Lang A, Volke V, Võikar V, Bourin M, Männistö PT (1997) Relation of exploratory behavior of rats in elevated plus-maze to brain receptor binding properties and serum growth hormone levels. Eur Neuro-Psychopharmacol 7:289–294

    Google Scholar 

  • Kõks S, Männisto PT, Bourin M, Shlik J, Vasar V, Vasar E (2000) Cholecystokinin–induced anxiety in rats: relevance of pre–experimental stress and seasonal variations. J Psychiatry Neurosci 25:33–42

    PubMed  Google Scholar 

  • Kõks S, Volke V, Veraksitš A, Rünkorg K, Sillat T, Abramov U, Bourin M, Huotari M, Männistö PT, Matsui T, Vasar E (2001) Cholecystokinin2 receptor-deficient mice display altered function of brain dopaminergic system. Psychopharmacology 158:198–204

    Article  PubMed  Google Scholar 

  • Kosaka T, Kosaka K, Tateishi K, Hamaoka Y, Yanaihara N, Wu JY, Hama K (1985) GABAergic neurons containing CCK-8-like and/or VIP-like immunoreactivities in the rat hippocampus and dentate gyrus. J Comp Neurol 239:420–430

    Article  PubMed  CAS  Google Scholar 

  • Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the \(2^{{ - \Delta \Delta C}}_{T} \) method. Methods 25:402–408

    Article  PubMed  CAS  Google Scholar 

  • Löw K, Crestani F, Keist R, Benke D, Brünig I, Benson JA, Fritschy J-M, Rülicke T, Bluethmann H, Möhler H, Rudolph U (2000) Molecular and neuronal substrate for the selective attenuation of anxiety. Science 290:131–134

    Article  PubMed  Google Scholar 

  • Miller KK, Hoffer A, Svoboda KR, Lupica CR (1997) Cholecystokinin increases GABA release by inhibiting a resting K+ conductance in hippocampal interneurons. J Neurosci 17:4994–5003

    PubMed  CAS  Google Scholar 

  • Miyasaka K, Kobayashi S, Ohta M, Kanai S, Yoshida Y, Nagata A, Matsui T, Noda T, Takiguchi S, Takata Y, Kawanami T, Funakoshi A (2002) Anxiety-related behaviors in cholecystokinin-A, B, and AB receptor gene knockout mice in the plus-maze. Neurosci Lett 335:115–118

    Article  PubMed  CAS  Google Scholar 

  • Möhler H, Fritschy JM, Rudolph U (2002) A new benzodiazepine pharmacology. Pharmacol Exp Ther 300:2–8

    Article  Google Scholar 

  • Nagata A, Ito M, Iwata N, Kuno J, Takano H, Minowa O, Chihara K, Matsui T, Noda T (1996) G protein-coupled cholecystokinin-B/gastrin receptors are responsible for physiological cell growth of the stomach mucosa in vivo. Proc Natl Acad Sci U S A 93:11825–11830

    Article  PubMed  CAS  Google Scholar 

  • Nobel F, Wank SA, Crawley JN, Bradwejn J, Seroogy KB, Hamon M, Roques BP (1999) International Union of Pharmacology.XXI. Structure, distribution, and functions of cholecystokinin receptors. Pharmacol Rev 51:745–781

    PubMed  Google Scholar 

  • Paylor R, Nguyen M, Crawley JN, Patrick J, Beaudet A, Orr-Urtreger A (1998) Alpha7 nicotinic receptor subunits are not necessary for hippocampal-dependent learning or sensorimotor gating: a behavioral characterization of Acra7-deficient mice. Learn Mem 5:302–316

    PubMed  CAS  Google Scholar 

  • Perez de la Mora M, Hernandez-Gomez AM, Mendez-Franco J, Fuxe K (1993) Cholecystokinin-8 increases K(+)-evoked [3H] gamma-aminobutyric acid release in slices from various brain areas. Eur J Pharmacol 250:423–430

    Article  PubMed  CAS  Google Scholar 

  • Pommier B, Beslot F, Simon A, Pophillat M, Matsui T, Daugé V, Roques BP, Noble F (2002) Deletion of CCK2 receptor in mice results in an upregulation of the endogenous opioid system. J Neurosci 22:2005–2011

    PubMed  CAS  Google Scholar 

  • Rasmussen K, Helton DR, Berger JE, Scearce E (1993) The CCK-B antagonist LY288513 blocks effects of diazepam withdrawal on auditory startle. Neuroreport 5:154–156

    Article  PubMed  CAS  Google Scholar 

  • Raud S, Rünkorg K, Veraksitš A, Reimets A, Nelovkov A, Abramov U, Volke V, Kõks S, Vasar E, Matsui T, Bourin M (2003) Targeted mutation of CCK2 receptor gene modifies the behavioral effects of diazepam in female mice. Psychopharmacology 168:417–425

    Article  PubMed  CAS  Google Scholar 

  • Rodgers RJ, Boullier E, Chatzimichalaki P, Cooper GD, Shorten A (2002) Contrasting phenotypes of C57BL/6JOlaHsd, 129S2/SvHsd and 129/SvEv mice in two exploration-based tests of anxiety-related behaviour. Physiol Behav 77:301–310

    Article  PubMed  CAS  Google Scholar 

  • Sebret A, Lena I, Crete D, Matsui T, Roques BP, Daugé V (1999) Rat hippocampal neurons are critically involved in physiological improvement of memory processes induced by cholecystokinin-B receptor stimulation. J Neurosci 19:7230–7237

    PubMed  CAS  Google Scholar 

  • Shlik J, Vasar E, Bradwejn J (1997) Cholecystokinin and psychiatric disorders: role in aetiology and potential of receptor antagonists in therapy. CNS Drugs 8:134–152

    Article  CAS  Google Scholar 

  • Singh L, Field MJ, Vass CA, Hughes J, Woodruff GN (1992) The antagonism of benzodiazepine withdrawal effects by the selective cholecystokininB receptor antagonist CI-988. Br J Pharmacol 105:8–10

    PubMed  CAS  Google Scholar 

  • Tsutsumi T, Akiyoshi J, Isogawa K, Kohno Y, Hikichi T, Nagayama H (1999) Suppression of conditioned fear by administration of CCKB receptor antagonist PD135158. Neuropeptides 33:483–486

    Article  PubMed  CAS  Google Scholar 

  • Vasar E, Peuranen E, Harro J, Lang A, Oreland L, Männistö PT (1993) Social isolation of rats increases the density of cholecystokinin receptors in the frontal cortex and abolishes the anti-exploratory effect of caerulein. Naunyn-Schmiedeberg’s Arch Pharmacol 348:96–101

    Article  PubMed  CAS  Google Scholar 

  • Võikar V, Kõks S, Vasar E, Rauvala H (2001) Strain and gender differences in the behavior of mouse lines commonly used in transgenic studies. Physiol Behav 72:271–278

    Article  PubMed  Google Scholar 

  • Võikar V, Vasar E, Rauvala H (2004) Behavioral alterations induced by repeated testing in C57BL/6J and 129S2/Sv mice: implications for phenotyping screens. Genes Brain Behav 3:27–38

    Article  PubMed  Google Scholar 

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Acknowledgements

This study was financially supported by the grants from the Estonian Science foundation (GARFS 5528, GARFS 5529, GARFS 5688), from the Estonian Ministry of Education and Science (0182584Bs03) and VARMC-TIPP.

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Authors and Affiliations

  1. Department of Physiology, Biomedicum, University of Tartu, 19 Ravila Street, 50411, Tartu, Estonia

    Sirli Raud, Jürgen Innos, Urho Abramov, Ain Reimets, Sulev Kõks, Andres Soosaar & Eero Vasar

  2. Centre of Molecular and Clinical Medicine, University of Tartu, 19 Ravila Street, 50411, Tartu, Estonia

    Sirli Raud, Jürgen Innos, Urho Abramov, Ain Reimets, Sulev Kõks, Andres Soosaar & Eero Vasar

  3. Division of Hematology/Oncology, Department of Medicine, Kobe University School of Medicine, 650-0017, Kobe, Japan

    Toshimitsu Matsui

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  1. Sirli Raud
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  2. Jürgen Innos
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  3. Urho Abramov
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  4. Ain Reimets
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  7. Toshimitsu Matsui
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Correspondence to Sulev Kõks.

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Raud, S., Innos, J., Abramov, U. et al. Targeted invalidation of CCK2 receptor gene induces anxiolytic-like action in light–dark exploration, but not in fear conditioning test. Psychopharmacology 181, 347–357 (2005). https://doi.org/10.1007/s00213-005-2255-x

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  • DOI: https://doi.org/10.1007/s00213-005-2255-x

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