Modulation of agmatine on calcium signal in morphine-dependent CHO cells by activation of IRAS, a candidate for imidazoline I1 receptor

The present study investigated the effects of agmatine action on imidazoline I1 receptor antisera-selected protein (IRAS), a candidate for imidazoline I1 receptor, on prolonged morphine-induced adaptations of calcium signal and long-lasting alterations in gene expression to further elucidate the role of IRAS in opioid dependence. Two cell lines, Chinese hamster ovary cells expressing μ opioid receptor alone (CHO-μ) and expressing μ opioid receptor and IRAS together (CHO-μ/IRAS), were used. After chronic treatment with morphine for 48 h, naloxone induced a significant elevation of intracellular calcium concentration ([Ca2+]i) in CHO-μ and CHO-μ/IRAS cells. Agmatine (0.01–3 μM) concentration-dependently inhibited the naloxone-precipitated [Ca2+]i elevation when co-pretreated with morphine in CHO-μ/IRAS, but not in CHO-μ. Efaroxan, an imidazoline I1 receptor-preferential antagonist, completely reversed the effect of agmatine in CHO-μ/IRAS. Agmatine (1–10 μM) administration after chronic morphine exposure for 48 h partially decreased the [Ca2+]i elevation in CHO-μ/IRAS which was entirely antagonized by efaroxan, but not in CHO-μ. In addition, agmatine (1 μM) co-pretreated with morphine attenuated the naloxone-precipitated increases of cAMP-responsive element binding protein and extracellular signal-regulated kinase 1/2 phosphorylations and c-Fos expression in CHO-μ/IRAS. These effects were blocked by efaroxan as well. Taken together, these results indicate that the agmatine-IRAS action system attenuates the up-regulations of Ca2+ signal and its downstream gene expression in morphine-dependent model in vitro, providing additional evidence to support the contribution of IRAS to opioid dependence.