CHOLINERGIC-INDUCED PRODUCTION OF REACTIVE OXYGEN SPECIES IN HUMAN NEUROBLASTOMA-CELLS

Stimulation of human SH-SY5Y neuroblastoma cells by a muscarinic recep tor agonist, carbachol (CCh; 1 mM), elevated levels of free intracellu lar calcium and subsequently increased the production of reactive oxyg en species (ROS). Quinuclidinylbenzilate (QNB) binding increased at 1 h after CCh, but returned back to the control level at 3 h. Production of ROS increased, however, during the 3 h time period. CCh also incre ased the translocation of protein kinase C (PKC) to the membrane. ROS production was completely blocked by atropine and a PKC inhibitor, Po 31-8220. These results show that increased ROS production was a result of muscarinic receptor stimulation, and that PKC had an active role i n this cellular stimulation. ROS production upon cellular stimulation by CCh was completely inhibited also by superoxide dismutase, and part ially by catalase, indicating that the formation of superoxide anion d ominated in cholinergic-induced generation of ROS in human neuroblasto ma cells. These results also show that muscarinic stimulation causes s ustained ROS production in human neuroblastoma cells. The slow increas e in ROS production by CCh suggest a stepwise cascade of events leadin g to oxidative stress with a triggering role of cholinergic muscarinic receptors in this process.