Cholecystokinin-Induced Anxiety in Rats: Relevance of Pre-Experimental Stress and Seasonal Variations Sulev Koks, MD, Phd; Pekka T

Cholecystokinin-induced anxiety in rats: relevance of pre-experimental stress and seasonal variations Sulev Koks, MD, PhD; Pekka T. Mannisto, MD, PhD; Michel Bourin, PhD; Jakov Shlik, MD, PhD; Veiko Vasar, MD, PhD; Eero Vasar, MD, PhD K6ks - Department of Physiology and Institute of Molecular and Cell Biology, University of Tartu, Tartu, Estonia; Mannisto - Department of Pharmacology and Toxicology, University of Kuopio, Kuopio, Finland; Bourin - Department of Pharmacology, University of Nantes, Nantes, France; Shlik, V. Vasar - Department of Psychiatry, University of Tartu, Tartu; E. Vasar Department of Physiology University of Tartu, Tartu Objective: To examine the influence of pre-experimental stress on the anxiogenic-like action of caerulein, an agonist of cholecystokinin (CCK) receptors. Differences in the anxiety levels of rats in summer and winter, and the role of CCK in these behavioural alterations, were also examined. Design: Prospective animal study. Interventions: Male Wistar rats were injected with the CCK agonist caerulein, or the CCK antagonists L-365,260 or devazepide, after being exposed to pre-experimental stress (handling and isolation).Outcome measures: Performance in the plus-maze model of anxiety; serum levels of prolactin, thyrotropin and growth hormone; brain density and affinity of dopamine D2, serotonin 5-HT2 and CCK receptors. Results: Caerulein (5 pg/kg, subcutaneous injection) caused the strongest action in animals brought to the experimental room immediately before the experiment and kept in isolation after the administration of caerulein. Caerulein did not cause any reduction of exploratory activity in rats made familiar with the experimental room and kept in the home-cage after the injection of the CCK agonist. The anti- exploratory action of caerulein in stressed rats was reversed by the CCK antagonist L-365,260 (I100 pg/kg, intraperitoneal injection), demonstrating the involvement of the CCKB receptor subtype. In addition, seasonal fluctuations occur in the exploratory activity of rats; such activity was much lower in July than in November. The rats displaying the reduced exploratory activity had an increased number of CCK receptors in the frontal cortex and hippocampus. Simultaneously, the density of serotonin 5-HT2 receptors in the frontal cortex, but not that of dopamine D2 receptors in the striatum, was elevated. The blood level of growth hormone was also higher in July. Conclusions: The anti-exploratory action of caerulein appears to be dependent on the pre-experimental stress of rats. Moreover, the seasonal variations of exploratory behaviour of rats are evident in the plus-maze model of anxiety. The reduced exploratory activity in summer appears to be related to the elevated density of CCK and 5-HT2 receptors in the brain. Correspondence to: S. K6ks, Department of Physiology, University of Tartu, 19 Ravila St., 5041 1 Tartu Estonia; fax 3727 374332; [email protected] Medical subject headings: anxiety; brain; caerulein; cholecystokinin; devazepide; exploratory behavior; hormones; prolactin; radioligand assay; rats, Wistar; receptors, cholecystokinin; receptors, dopamine; receptors, serotonin; seasons; stress; thyrotropin J Psychiatry Neurosci 2000;25(1):33-42. Submitted April 15, 1998 Revised Mar. 2, 1999; June 30, 1999 Accepted July 20, 1999 © 2000 Canadian Medical Association Li= O' v.,%w.. -.:i-- ... i" I"M.W-MaWWR Objectif: Etudier l'influence du stress pre-experimental sur 1'effet quasi anxiogene de la ceruleine, agoniste des recepteurs de la cholecystokinine (CCK). On a aussi etudie les differences entre les niveaux d'anxiete chez les rats l'6te et l'hiver et le r6le de la CCK dans ces modifications du comportement. Conception: etude prospective sur animaux. Interventions: On a injecte a des rats Wistar males de la ceruleine, agoniste de la CCK, les antagonistes L-365,260 de la CCK ou du devazepide, apres les avoir expose a un stress pre-experimental (manipu- lation et isolement). Mesures de resultats: Performance dans le labyrinthe (un modele d'anxiete); taux seriques de prolactine, de thyrotropine et d'hormone de croissance; densite cerebrale et affinite des recepteurs de la dopamine D2, de la serotonine 5-HT2 et de la CCK. Resultats: La ceruleine (injection sous-cutanee de 5 pg/kg) a eu 1'effet le plus marque chez les animaux transportes a la salle experimentale immediatement avant l'experience et gardes en isolement apres I'administration de la ceruleine. La ceruleine n'a pas provoque de reduction de l'activite exploratoire chez les rats qui ont appris a connaitre la salle experimentale et ont ete gardes dans leur cage habituelle apres l'injection de l'agoniste de la CCK. L'action anti-exploratoire de la ceruleine chez les rats stresses a ete contree par I'antagoniste L-365,260 de la CCK (injection intraperitoneale de 100 pg/kg), ce qui demontre le role du sous-type recepteur de la CCKB. On enregistre en outre des fluctuations saisonnieres de l'activit6 exploratoire des rats, qui est beaucoup plus basse en juillet qu'en novembre. Les rats qui ont montre une baisse de l'activite exploratoire avaient davantage de recepteurs de la CCK dans le cortex frontal et l'hippo- campe. En meme temps, la densite des recepteurs de la serotonine 5-HT2 dans le cortex frontal etait elevee, mais celle des recepteurs de la dopamine D2 dans le striatum ne l'etait pas. Le taux sanguin d'hormone de croissance etait aussi plus eleve en juillet. Conclusions: L'action anti-exploratoire de la ceruleine semble liMe au stress pre- exp6rimental chez les rats. De plus, le labyrinthe montre les variations saisonnieres du comportement exploratoire des rats. La baisse de l'activite exploratoire au cours de l'ete semble liee a l'elevation de la densite des recepteurs de la CCK et 5-HT2 dans le cerveau. Introduction the panicogenic action of CCKB agonists, demonstrating a role for CCK, receptors.6 Cholecystokinin octapeptide (CCK-8) is a widely dis- There is some evidence that the response of rats to the tributed neuropeptide in the mammalian brain.' Recent anxiogenic-like action of CCK may be dependent on the evidence suggests that CCK-8 is involved in the neuro- level of pre-experimental stress.2 Biro et all' have biology of anxiety.2 The anxiogenic-like action of CCK demonstrated that the anxiogenic-like action of CCK is agonists is established in various animal species, related to the release of corticotropin-releasing hor- including the monkey, guinea-pig, rat, mouse and cat.3 mone (CRH). CRH obviously plays a key role in the CCKB-receptor (brain subtype) antagonists are effective behavioural and hormonal mechanisms of stress." in antagonizing the anxiogenic-like action of CCK ago- Recent studies have shown that mice lacking the CRH1 nists, but they also cause an anxiolytic-like effect in var- receptor display impaired stress response and reduced ious animal models of anxiety.2'" There is a growing anxiety in the light/dark compartment test.'2"13 The body of evidence that the administration of the CCK8- results obtained from the clinical studies provide some receptor agonists CCK-4 and pentagastrin induces evidence for an altered activity of CRH in anxiety dis- panic-like attacks in healthy volunteers and patients orders, but apparently less than that seen in depres- suffering from panic disorder.4'79 The sensitivity of sion.""l' Patients suffering from panic disorder display patients with panic disorder to the panicogenic action increased sensitivity to CRH-induced corticotropin and of CCKB agonists is significantly higher than that of cortisol release.18 The administration of the CCK8-recep- healthy subjects.8 The striking similarity between CCK- tor agonists CCK-4 and pentagastrin has been shown to induced panic-like attacks and natural attacks has been activate the hypothalamic-pituitary-adrenal axis.7'19'20 In established in patients with panic disorder. The aug- their recent study, Koszycki et a121 found that the mented response to CCK agonists is also described in administration of CCK-4 significantly enhanced corti- patients suffering from other anxiety disorders.3 cotropin secretion in healthy volunteers responding Accordingly, increased sensitivity to CCKB-receptor with panic-like attacks compared with nonresponders. agonist-induced panic-like attacks seems to be a com- In the present study, an attempt was made to clarify the mon feature of anxiety disorders. The pretreatment of significance of pre-experimental stress on the anxio- healthy subjects with CCK8-receptor antagonists blocks genic-like action of CCK. I... AWO W7--- NIWw*_*t5z^#0' CC: in: a rat model o::X(anx Several investigators have described the seasonal tion of caerulein (5 pg/kg, subcutaneous injection; fluctuations in anxiety. Recent reports have described a Sigma, US), an agonist of CCK receptors. Caerulein or higher incidence in the occurrence of first panic attacks saline solution was injected 15 minutes before the in the summer, irrespective of the hemisphere in which beginning of the plus-maze study. One half of the ani- the study was carried out.22-24 Lelliott et aP have found mals were isolated after the injection, whereas the other that, of 57 patients with panic disorder with agorapho- half were placed back into the home-cage. The action of bia, more had their first panic attack in late spring and the CCKB antagonist L-365,260 (1 to 100 pg/kg, summer than in fall and winter, and in warm weather intraperitoneal injection; Merck Sharp & Dohme, UK) than in cold weather. Therefore, the second major goal and the CCKA antagonist devazepide (1 to 100 pg/kg, of the present study was to investigate the differences in intraperitoneal injection; Merck Sharp & Dohme) on the the anxiety levels of rats in summer and winter, and the anxiogenic-like action of caerulein was studied in the role of CCK in these behavioural alterations.

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