The effects of a muscarinic receptor agonist, carbachol (CCh), on the production of reactive oxygen species (ROS) and the levels of the intracellular glutathione (GSH) were studied in human SH-SY5Y neuroblastoma cell line. ROS and GSH were measured by using fluorescent probes, dichlorofluorescin (DCF) and monochlorobimane (MBCL), respectively. The translocation of protein kinase C (PKC) to the cell membrane was measured by phorbol dibutyrate (PDBu) binding and muscarinic receptor number was assayed by using quinuclidinylbenzilate (QNB) binding in intact SH-SY5Y cells. One millimolar CCh increased ROS production 1.87 fold at 60 min, 2.37 fold at 120 min, and 3.0 fold at 180 min as compared to the control values. However, CCh, at a concentration of 500 μM, caused only a slight increase in ROS production, and values obtained with 100 μM CCh remained at control levels. Intracellular GSH levels decreased by about 20% when the cells were incubated for 120 min with 1 mM CCh. However, GSH levels returned back to the control level after 180 min incubation. PDBu binding (B M A X ) increased by 73% after 20 min incubation with 1 mM CCh as compared to the control cells. Also 500 μM CCh increased PDBu binding by 33-54% as compared to corresponding controls, i.e. PKC translocation to the membrane was enhanced. QNB binding (B M A X ) increased by 166% after 60 min incubation with 1 mM CCh, and returned back to the control level after 180 min incubation. We conclude that muscarinic receptor stimulation with CCh may cause oxidative burst in human neuroblastoma cells which may be partially routed through PKC, because increase in PDBu binding is a prerequisite for the production of ROM. However, there may be other effector mechanisms that have to be activated prior to the production of ROS.