PEGylated cholecystokinin is more potent in inducing anorexia than conditioned taste aversion in rats

Abstract

Background and purpose: The physiological involvement of endogenous cholecystokinin (CCK) in the termination of feeding has been challenged by evidence of aversive effects of exogenous CCK8. We previously prolonged the anorectic effect of CCK by conjugation to polyethylene glycol (PEGylation) to produce PEG-CCK9. In this study, we investigated the ability of different doses of PEG-CCK9 to induce conditioned taste aversion (CTA) and satiety and identified the receptors involved in CTA induction.

Experimental approach: Induction of CTA, measured by the saccharin preference ratio determined in a two-bottle CTA procedure, and of satiety in adult male Wistar rats after intraperitoneal (i.p.) injection of different doses of PEG-CCK9 (1, 2, 4, 8, 16 or 32 microg kg(-1)) was compared. Devazepide (100 microg kg(-1)) and 2-NAP (3 mg kg(-1)), two selective CCK1-receptor antagonists, were co-administered i.p. with PEG-CCK9 (8 microg kg(-1)) and the CTA effects monitored.

Key results: PEG-CCK9 dose-dependently induced CTA, with a minimal effective dose of 8 microg kg(-1), whereas the minimal effective dose to induce satiety was 1 microg kg(-1). The CTA effects of PEG-CCK9 were completely abolished by i.p. administration of devazepide prior to PEG-CCK9 treatment and only partially abolished by administration of 2-NAP.

Conclusions and implications: Although PEG-CCK9-induced satiety and PEG-CCK9-induced CTA both increased with dose, the conjugate was more potent in inducing satiety, suggesting that the anorexia could not be completely attributed to the aversiveness of the drug. As observed with induction of satiety, PEG-CCK9-induced CTA was mediated by CCK1-receptors.

Figure 1

Dose-dependency of conditioned taste aversion (CTA) effects in rats treated with PEG-CCK₉ (1, 2, 4, 8, 16 and 32 μg kg⁻¹), determined using a two-bottle test procedure. Graph (a) shows the saccharin preference ratios of the PEG-CCK₉-treated rats, presented as a percentage of their own control saccharin preference ratio (mean+s.e.mean; n=6). ANOVA analysis (F=9,136, d.f.=(6, 38)) with the post hoc Tukey test revealed significant different results compared with controls (dose 0 μg kg⁻¹). ^*P<0.05; ^**P<0.001. Graph (b) depicts the PEG-CCK₉-induced saccharin preference ratio data presented as a percentage of their own control saccharin preference ratio, log-transformed and fitted to a standard dose–response model with constraint; bottom=0 (mean±s.e.mean; n=6). The fitted dose–response relationship revealed an EC₅₀ of 14.3 μg kg⁻¹ with a 95% confidence interval of 8.9–22.9.

Figure 2

Relationship between PEG-CCK₉-induced food intake reduction and the corresponding conditioned taste aversion (CTA). The dose-related effects of PEG-CCK₉ (1, 2, 4, 8 and 16 μg kg⁻¹) on food intake and on CTA, both presented as a percentage of control (mean±s.e.mean; food intake data: n=10; CTA data: n=6), were plotted on the same graph and approximated with linear regression to determine the relationship between the two behavioural measures. The 95% confidence intervals of the two regression slopes overlap (slope 95% confidence interval for the food intake dose–response curve: −5.7 to −1.2; slope 95% confidence interval for the CTA dose–response curve: −4.7 to −1.6), indicating that the two functions are parallel and thus the food intake reduction and CTA induced by the same doses of PEG-CCK₉ are functionally related. ANOVA analysis of the food intake data (F=13,75, d.f.=(5, 124)) and the CTA-data (F=9,136, d.f.=(6, 38)) with the post hoc Tukey test revealed significant different results compared with controls (dose 0 μg kg⁻¹). The minimal effective dose (MED) for the two parameters is indicated by an asterisk.

Figure 3

Effect of devazepide (a) or 2-NAP (b) on PEG-CCK₉-induced conditioned taste aversion (CTA). The saccharin preference ratios, determined in a two-bottle CTA procedure, are presented as a percentage of their own control (mean±s.e.mean; n=8). The letters above the bars indicate statistically significant differences among treatment groups (P<0.05) (ANOVA followed by Tukey's multiple comparison test: devazepide; F=17,97, d.f.=(3, 28), 2-NAP; F=7,567, d.f.=(3, 27)).