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

The α2-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 α2-adrenoceptors in order to know if they could happen at the cellular level. In a first step, incubation with morphine (10 μM) or the δ opioid agonist DPDPE (1 μM) for 6 h was shown to decrease the reduction of (4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium (MTT) by NG108-15 neuroblastoma × glioma hybrid cells in a naloxone-sensitive manner, thus showing that the opioids affect the redox status of the cells in a δ receptor-mediated way. Further experiments with 2–24 h incubation periods were subsequently performed with morphine 0.1 μM, 10 μM 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 μM and 10 μM) 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 μM, 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 δ receptors and also sensitive to α2-adrenoceptor blockade; therefore, the interactions previously described in vivo between opioid and α2-adrenoceptor ligands do not necessarily require the presence of functional neuronal networks and they could happen at the cellular level.