Effects of exogenous cholecystokinin octapeptide on acquisition of naloxone precipitated withdrawal induced conditioned place aversion in rats

Abstract

Cholecystokinin octapeptide (CCK-8), a gut-brain peptide, regulates a variety of physiological behavioral processes. Previously, we reported that exogenous CCK-8 attenuated morphine-induced conditioned place preference, but the possible effects of CCK-8 on aversively motivated drug seeking remained unclear. To investigate the effects of endogenous and exogenous CCK on negative components of morphine withdrawal, we evaluated the effects of CCK receptor antagonists and CCK-8 on the naloxone-precipitated withdrawal-induced conditioned place aversion (CPA). The results showed that CCK2 receptor antagonist (LY-288,513, 10 µg, i.c.v.), but not CCK1 receptor antagonist (L-364,718, 10 µg, i.c.v.), inhibited the acquisition of CPA when given prior to naloxone (0.3 mg/kg) administration in morphine-dependent rats. Similarly, CCK-8 (0.1-1 µg, i.c.v.) significantly attenuated naloxone-precipitated withdrawal-induced CPA, and this inhibitory function was blocked by co-injection with L-364,718. Microinjection of L-364,718, LY-288,513 or CCK-8 to saline pretreated rats produced neither a conditioned preference nor aversion, and the induction of CPA by CCK-8 itself after morphine pretreatments was not significant. Our study identifies a different role of CCK1 and CCK2 receptors in negative affective components of morphine abstinence and an inhibitory effect of exogenous CCK-8 on naloxone-precipitated withdrawal-induced CPA via CCK1 receptor.

Figure 1

A: Experimental procedure of naloxone-precipitated withdrawal-induced CPA. The preference score (CPA score) defined as the time spent in the naloxone-paired compartment on day 7 (test phase) for 15 min minus the average time spent in the same compartment on day 4 and day 5 (pre-conditioning phase). Sal, saline; Mor, morphine; Nal, naloxone; B: Conditioned place aversion (CPA) was induced by naloxone (0.3 mg/kg, i.p.)-precipitated withdrawal in morphine (10 mg/kg, s.c.) pretreated rats. Analysis of the data obtained from CPA testing with two-way ANOVA revealed a significant morphine and naloxone effect on the CPA scores. The data are expressed as the mean ± S.D. (n = 8, 9, 8, 8). **P<0.01, compared to Sal+Sal group.

Figure 2. Effects of CCK receptor antagonists on the development of naloxone-precipitated withdrawal-induced CPA in morphine dependent rats.

Animals were pretreated with either saline or morphine. They were subsequently microinjected with vehicle (1 µl, i.c.v.), CCK1 receptor antagonist (L-364,718, 10 µg, i.c.v.) and CCK2 receptor antagonist (LY-288,513, 10 µg, i.c.v.) 15 min prior to naloxone precipitation and CPA conditioning. Data of CPA scores are expressed as means ± S.D. (n = 10, 9, 7, 9 and 9, 8, 10, 8 in saline and morphine pretreated rats, respectively) **P<0.01 compared with the morphine pretreated and saline microinjected group by Bonferroni's post hoc test.

Figure 3. Effects of exogenous CCK-8 on the development of naloxone-precipitated withdrawal-induced CPA in morphine dependent rats.

A: The induction of CPA by CCK-8 (0.01–1 µg, i.c.v.) itself after morphine pretreatment in rats. Animals were pretreated with morphine, and then microinjected with CCK-8 instead of naloxone for precipitation before conditioning. Data of CPA scores are expressed as means ± S.D. (n = 7, 8, 8, 9). B: Animals were pretreated with either saline or morphine, and then microinjected with CCK-8 (0.01, 0.1 and 1 µg, i.c.v.) 15 min before naloxone precipitation and CPA conditioning. Data of CPA scores are expressed as means ± S.D. (n = 8, 9, 8, 7 and 8, 7, 9, 8 in saline and morphine pretreated rats respectively) **P<0.01 compared with the morphine pretreated and saline microinjected group by Bonferroni's post hoc test.

Figure 4. Effect of selective CCK receptor antagonists on CCK-8 inhibiting the development of naloxone-precipitated withdrawal-induced CPA in morphine dependent rats.

Animals were pretreated with morphine and then co-microinjected with CCK-8 (0.01, 0.1 and 1 µg, i.c.v.) and saline, vehicle, CCK1 receptor antagonist (L-364,718, 10 µg, i.c.v.) or CCK2 receptor antagonist (LY-288,513, 10 µg, i.c.v.) 15 min before naloxone precipitation and CPA conditioning. Data of CPA scores are expressed as means ± S.D. (n = 7, 8, 9, 8, 8, respectively) **P<0.01, as compared with morphine-pretreated and saline-microinjected control, ^# P<0.05, as compared with CCK-8 plus saline co-microinjected group. Statistical analysis was conducted by one-way ANOVA followed by Bonferroni's test.