Yohimbine prevents the effect of morphine on the redox status of neuroblastoma x glioma NG108-15 cells

The alpha(2)-adrenoceptor antagonist yohimbine is known to interact with the effects of opioid receptor agonists in vivo, and thus could modulate the action of morphine-like analgesics. The focus of the present work was to further study these interactions in a cell culture endowed with opioid and alpha(2)-adrenoceptors in order to know if they could happen at the cellular level. In a first step. incubation with morphine (10 mu M) or the delta opioid agonist DPDPE (1 mu M) for 6 h was shown to decrease the reduction of (4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium (MTT) by NG108-15 neuroblastoma x glioma hybrid cells in a naloxone-sensitive manner, thus showing that the opioids affect the redox status of the cells in a delta receptor-mediated way. Further experiments with 2-24h incubation periods were subsequently performed with morphine 0.1 mu M, 10 mu M and 1 mM and several tests to confirm the effects on metabolism (MTT, Alamar Blue tests) to examine the potential toxic consequences (neutral red test, trypan blue exclusion assay, LDH test, caspase 3/7 activity) and to study the potential effect of yohimbine on morphine toxicity. These studies confirmed that incubation with morphine (0.1 mu M and 10 mu M) affected to a similar extent the redox status of the cells, an effect that did not translated into significant cell death and was transient since completely disappeared after 24 h of incubation. Morphine 1 mM was much more toxic than the lower concentrations. Yohimbine effectively prevented the effects of the lower concentrations of morphine when added to the incubation medium at 10 mu M, a concentration devoid of significant toxicity. It seems that the exposure to pharmacologically relevant concentrations of morphine gives rise to short-term metabolic alterations of NG108-15 cells mediated by delta receptors and also sensitive to alpha(2)-adrenoceptor blockade; therefore, the interactions previously described in vivo between opioid and alpha(2)-adrenoceptor ligands do not necessarily require the presence of functional neuronal networks and they could happen at the cellular level. (C) 2009 Elsevier Ireland Ltd. All rights reserved.