Hyperglycaemia has been observed with exposure to organophosphate insecticides. This study was designed to compare the effects of calcium channel blockers, alpha-adrenergic, beta-adrenergic, and muscarinic receptor blockers, and of free radical scavengers on insulin secretion from diazinon-treated islets of Langerhans isolated from the pancreas of rats using standard collagenase digestion, separation by centrifugation, and hand-picking technique. The islets were then cultured in an incubator at 37 °C and 5 % CO2. In each experimental set 1 mL of 8 mmol L-1 glucose plus 125 μg mL-1 or 625 μg mL-1 of diazinon were added, except for the control group, which received 8 mmol L-1 glucose alone. The cultures were then treated with one of the following: 30 μmol L-1 atropine, 100 μmol L-1 ACh + 10 μmol L-1 neostigmine, 0.1 μmol L-1 propranolol, 2 μmol L-1 nifedipine, 50 μmol L-1 phenoxybenzamine, or 10 μmol L-1 alphatocopherol. In all experiments, diazinon significantly reduced glucose-stimulated insulin secretion at both doses, showing no dose dependency, as the average inhibition for the lower dose was 62.20 % and for the higher dose 64.38 %. Acetylcholine and alpha-tocopherol restored, whereas atropine potentiated diazinon-induced hyposecretion of insulin. Alpha-, beta- and calcium channel blockers did not change diazinon-induced effects. These findings suggest that diazinon affects insulin secretion mainly by disturbing the balance between free radicals and antioxidants in the islets of Langerhans and by inducing toxic stress.