Effect of agmatine on the development of morphine dependence in rats: potential role of cAMP system

Abstract

Agmatine is an endogenous amine derived from arginine that potentiates morphine analgesia and blocks symptoms of naloxone-precipitated morphine withdrawal in rats. In this study, we sought to determine whether treatment with agmatine during the development of morphine dependence inhibits the withdrawal symptoms and that the effect is mediated by cAMP system. Exposure of rats to morphine for 7 days resulted in marked naloxone-induced withdrawal symptoms and agmatine treatment along with morphine significantly decreasing the withdrawal symptoms. The levels of cAMP were markedly increased in morphine-treated rat brain slices when incubated with naloxone and this increase was significantly reduced in rats treated with morphine and agmatine. The induction of tyrosine hydroxylase after morphine exposure was also reduced in locus coeruleus when agmatine was administered along with morphine. We conclude that agmatine reduces the development of dependence to morphine and that this effect is probably mediated by the inhibition of cAMP signaling pathway during chronic morphine exposure.

Fig. 1

The effects of agmatine (Agm) on naloxone precipitated morphine withdrawal. Rats were implanted with placebo (P) or one morphine (M) pellet (75 mg morphine base) on day 1 and two more pellets on day 3. After 7 days, withdrawal was induced on the eighth day by injecting naloxone (2 mg/kg, i.p.). Agmatine (10 mg/kg, i.p.) was injected twice daily for 7 days and no agmatine was injected on the eighth day, the day of inducing withdrawal. Panels A through E indicate mean withdrawal frequencies induced by naloxone that was observed for 15 min for each group (n=6). *p<0.05 compared to morphine group as determined by analysis of variance. Panel F illustrates the global withdrawal scores of the three groups of rats. This score consists of graded signs (based on the frequency) and checked signs (independent of the frequency) and was calculated using all symptoms of withdrawal. *p<0.05 compared to morphine group.

Fig. 2

Effect of agmatine on cAMP production in morphine-treated rat brain. Rats were exposed to morphine pellets (75 mg) for 7 days (one pellet on day 1 and two more pellets on day 3) and sacrificed on the eighth day. Brain cortical slices were prepared and cAMP production was measured in vitro. The levels of cAMP was measured in cortical slices (300 μm) in vitro after incubating with agmatine, norepinephrine or naloxone (100 μM each) for 15 min. The values are from 6 rats in each group and in vitro incubations were done in triplicates. *p<0.001 compared to placebo group; **p<0.001 compared to morphine group.

Fig. 3

Effect of agmatine on TH expression in morphine-treated rat brain. Rats were treated with placebo, morphine (one 75 mg pellet on day 1 and two more pellets on day 3) or morphine+agmatine (10 mg/kg, i.p., twice daily for 7 days) and brain regions were analyzed by immunoblot for the expression of TH. Lanes: 1, placebo; 2, morphine; 3, morphine+agmatine. The same samples were also used to determine the expression of β-actin as the control housekeeping protein. This is a representative data from one set of rats that was replicated in four animals in each group.