Differences in behavioural effects of amphetamine and dopamine-related gene expression in wild-type and homozygous CCK2 receptor deficient mice

Neuropeptide cholecystokinin (CCK) interacts with dopamine in the regulation of motor activity and motivations. Therefore, in CCK2 receptor deficient mice the behavioural effects of repeated amphetamine administration and changes in dopamine-related gene expression were studied. Four-day amphetamine (1 mg/kg) treatment induced a significantly stronger motor sensitization in homozygous mice compared to their wild-type littermates. However, in the conditioned place preference test the action of amphetamine was more pronounced in wild-type animals. As opposed to wild-type mice, amphetamine (1–3 mg/kg) did not cause a significant conditioned place preference in homozygous mice. The expression of Tyhy gene was elevated in the mesolimbic structures and Drd2 gene was down-regulated in the mesencephalon of saline-treated homozygous mice in comparison with respective wild-type group. Four-day treatment with amphetamine induced a significant increase in the expression of Tyhy in the mesencephalon, striatum and mesolimbic structures of wild-type mice, whereas in homozygous mice a similar change was evident only in the mesencephalon. Also, the expression of Drd1 gene in the striatum and Drd2 gene in the mesolimbic structures of wild-type mice were up-regulated under the influence of amphetamine. In conclusion, the present study established differences in the behavioural effects of amphetamine in wild-type and homozygous mice. The increased tone of dopaminergic projections from the mesencephalon to mesolimbic structures is probably related to increased amphetamine-induced motor sensitization in homozygous mice. The lack of development of up-regulation of Drd1 and Drd2 genes after repeated treatment with amphetamine probably explains the reduced place conditioning in CCK2 receptor deficient mice

Rimonabant attenuates amphetamine sensitisation in a CCK2 receptor-dependent manner

In this behavioural and pharmacological study in male CCK2 receptor-deficient mice (CCK2−/−), we evaluated the role of the interaction of endocannabinoids (eCBs) and cholecystokinin (CCK) on the regulation of anxiety-related and motor behaviours. Repeated treatment with amphetamine (2 mg/kg daily for four days) induced slightly weaker motor sensitisation in CCK2−/− mice compared to their wild-type (CCK2+/+) littermates. Co-administration of rimonabant (1 mg/kg) with amphetamine antagonised the development of motor sensitisation in CCK2+/+ mice. However, we did not find a similar effect of rimonabant in CCK2−/− mice. We did not find any differences between the behaviour of CCK2+/+ and CCK2−/− mice in models designed to assess emotional behaviours (dark/light exploration, marble burying and conditioned place aversion). This study supports the hypothesis that eCBs play a role in the development of amphetamine-induced sensitisation. Moreover, we have demonstrated that intact CCK2 receptors are necessary for the development of eCB-mediated sensitisation to amphetamine

Altered pain sensitivity and morphine-induced anti-nociception in mice lacking CCK2 receptors

Rationale Cholecystokinin (CCK) interacts with the endopioid system in the regulation of various physiological functions, including the control of pain sensitivity, motor activity and emotional behaviour. Objective The aim of the present work was to study the pain sensitivity, morphine-induced antinociception and density of opioid receptors in mice lacking CCK2 receptors. Methods Plantar analgesia and hotplate tests were used to evaluate pain sensitivity and morphine-induced antinociception. The parameters of opioid receptors were analysed by using [3H]-diprenorphine binding. Results In the plantar analgesia test the latency of hind paw withdrawal was significantly increased in CCK2 receptor deficient mice compared to wild-type (+/+) littermates. The treatment with saline reversed the reduced pain sensitivity in heterozygous (+/−) and homozygous (−/−) mice. The administration of morphine (1 mg/kg) induced a significantly stronger antinociceptive effect in homozygous (−/−) mice compared with wild-type (+/+) animals. In the hotplate test, only homozygous (−/−) mutant mice displayed the delayed latency of hind paw licking/shaking in comparison with wild-type (+/+) mice. The injection of saline and isolation of mice for 30 min reversed the delayed response in homozygous (−/−) mice. However, in this test, the anti-nociceptive action of morphine (5–10 mg/kg) in mutant mice did not differ from that in wild-type (+/+) littermates. By contrast, the jump latency was decreased in both homozygous (−/−) and heterozygous (+/−) mice in the hotplate test. The increased density of opioid receptors was established in the striatum of homozygous (−/−) mice. Conclusion It is apparent that the targeted mutagenesis of the CCK2 receptor gene has different effects on the sensitivity of opioid receptors in various brain structures. This is a probable reason for the altered pain sensitivity and morphine-induced antinociception in mutant mice compared to wild-type (+/+) littermates

Distinct changes in the behavioural effects of morphine and naloxone in CCK2 receptor-deficient mice

The effects of morphine, μ-opioid receptor agonist, and naloxone, a non-selective opioid receptor antagonist, in the locomotor activity and place conditioning tests were studied in the CCK2 receptor-deficient male mice. The exposure of mice to the motility boxes for 3 consecutive days induced a significant inhibition of locomotor activity in the wild-type (+/+) mice compared to homozygous (−/−) animals. The administration of naloxone (10 mg/kg i.p.) to animals, adapted to the motility boxes, induced a significant reduction of locomotor activity in the homozygous (−/−), but not in the wild-type (+/+) mice. Treatment of habituated mice with morphine (10 mg/kg i.p.) caused a stronger increase of locomotor activity in the wild-type (+/+) mice compared to the homozygous (−/−) littermates. In the place preference test the pairing of the preferred side with naloxone (1 and 10 mg/kg i.p.) induced a dose-dependent place aversion in the wild-type (+/+) mice. The treatment with naloxone was less effective in the homozygous (−/−) mice, because the high dose of naloxone (10 mg/kg) tended to shift the preference. The pairing of morphine (3 mg/kg i.p.) injections with the non-preferred side induced a significant place preference both in the wild-type (+/+) and homozygous (−/−) mice. The increased density of opioid receptors was established in the striatum of homozygous (−/−) mice, but not in the other forebrain structures. In conclusion, the targeted invalidation of CCK2 receptors induces a dissociation of behavioural effects of morphine and naloxone. Morphine-induced place preference remained unchanged, whereas hyper-locomotion was less pronounced in the mutant mice compared to the wild-type (+/+) littermates. By contrast, naloxone-induced place aversion was weaker, but naloxone caused a stronger inhibition of locomotor activity in the homozygous (−/−) mice than in the wild-type (+/+) animals. These behavioural alterations can be explained in the light of data that the targeted mutation of CCK2 receptors induces distinct changes in the properties of opioid receptors in various brain structures

Targeted mutation of CCK2 receptor gene modifies the behavioural effects of diazepam in female mice

Rationale Evidence suggests that GABA and CCK have opposite roles in the regulation of anxiety. Objective The aim of the present work was to study diazepam-induced anxiolytic-like action and impairment of motor co-ordination, and the parameters of benzodiazepine receptors in mice lacking CCK2 receptors. Methods The action of diazepam (0.5–3 mg/kg IP) was studied in the elevated plus-maze model of anxiety and rotarod test using mice lacking CCK2 receptors. The parameters of benzodiazepine receptors were analysed using [3H]-flunitrazepam binding. Results In the plus-maze test, the exploratory activity of the homozygous (−/−) mice was significantly higher compared to their wild-type (+/+) littermates. However, the wild-type (+/+) mice displayed higher sensitivity to the anxiolytic-like action of diazepam. Even the lowest dose of diazepam (0.5 mg/kg) induced a significant increase of open arm entries in the wild-type (+/+) mice. A similar effect in the homozygous (−/−) mice was established after the administration of diazepam 1 mg/kg. The highest dose of diazepam (3 mg/kg) caused a prominent anxiolytic-like effect in the wild-type (+/+) mice, whereas in the homozygous (−/−) animals suppression of locomotor activity was evident. The performance of the homozygous (−/−) mice in the rotarod test did not differ from that of the wild-type (+/+) littermates. However, a difference between the wild-type (+/+) and homozygous (−/−) animals became evident after treatment with diazepam. Diazepam (0.5 and 3 mg/kg) induced significantly stronger impairment of motor co-ordination in the homozygous (−/−) mice compared to their wild-type (+/+) littermates. The density of benzodiazepine binding sites was increased in the cerebellum, but not in the cerebral cortex and hippocampus, of the homozygous (−/−) mice. Conclusions Female mice lacking CCK2 receptors are less anxious than their wild-type (+/+) littermates. The reduced anxiety in homozygous (−/−) mice probably explains why the administration of a higher dose of diazepam is necessary to induce an anxiolytic-like action in these animals. The highest dose of diazepam (3 mg/kg) induced significantly stronger suppression of locomotor activity and impairment of motor co-ordination in the homozygous (−/−) mice compared to the wild-type (+/+) littermates. The increase in the action of diazepam is probably related to the elevated density of benzodiazepine receptors in the cerebellum of homozygous (−/−) mice. The present study seems to be in favour of increased tone of the GABAergic system in mice without CCK2 receptors

Cholecystokinin 2 receptor-deficient mice display altered function of brain dopaminergic system

Rationale: Cholecystokinin (CCK) has been shown to coexist and interact with dopamine in the regulation of behaviour. Two different CCK receptors (CCK1 and CCK2) have an opposite influence on the activity of dopamine neurons. Stimulation of CCK2 receptors decreases the release of dopamine and that receptor could mediate the neuroleptic-like effect of CCK. Objective: To investigate the activity of the dopaminergic system in pharmacological experiments on CCK2 receptor (CCK2R)-deficient mice. Methods: We used age- and sex-matched littermates in all our experiments. To evaluate the behavioural differences, we performed the rotarod test and measured the locomotor activity of animals using computer-connected photoelectric motility boxes. Amphetamine and apomorphine, two dopaminergic drugs with different pharmacodynamic properties, were used to influence the activity of the dopaminergic system in the brain. Neurochemical differences related to the different genotype were analysed by means of high-performance liquid chromatography and radioligand binding studies. Results: Motor co-ordination was significantly impaired in the rotarod test of CCK2R receptor-deficient mice. Moreover, the locomotor activity of heterozygous (+/–) and homozygous (–/–) CCK2R receptor-deficient mice was somewhat reduced. A low dose of apomorphine (0.1 mg/kg), an unselective agonist of dopamine receptors, suppressed locomotor activity significantly more in homozygous (–/–) and heterozygous (+/–) mutant mice than in their wild-type (+/+) littermates. Amphetamine (3–6 mg/kg), increasing release of dopamine from the presynaptic terminals, caused a dose-dependent motor stimulation in wild-type (+/+) mice. In heterozygous (+/–) and homozygous (–/–) mice, a lower dose of amphetamine (3 mg/kg) did not alter the locomotor activity, whereas the higher dose of (6 mg/kg) induced a significantly stronger increase in locomotor activity in homozygous (–/–) mice than in their heterozygous (+/–) and wild-type (+/+) littermates. Despite the changes in the action of apomorphine and amphetamine in homozygous (–/–) mice, we did not find any significant differences in the concentration of dopamine and their metabolites in the striatum or cortex. However, the density of dopamine D2 receptors was significantly increased in the striatum of homozygous (–/–) animals compared with wild-type (+/+) mice. Conclusions: The targeted mutation of the CCK2 receptor gene induced gene dose-dependent changes in the activity of the dopaminergic system. The sensitivity of presynaptic dopamine receptors was increased in heterozygous (+/–) and homozygous (–/–) animals, whereas the increase in sensitivity of postsynaptic dopamine receptors was apparent only in homozygous (–/–) mice. Targeted mutagenesis Wild-type Heterozygous Homozygous Dopamine Serotonin Receptor Cholecystokinin Cholecystokinin2 receptor Motor co-ordination Rotarod test Motility boxes Locomotor activit