Leitinger_2000_J.Comp.Neurol_416_345

Three different cytochemical methods were used to detect acetylcholine in large, second-order neurons of locust ocelli (L-neurons). The first method used polyclonal antibodies raised against choline cleaved from acetylcholine and then conjugated with native protein, and this revealed strong staining for acetylcholine in axons whose number, size, and location indicated that they were of L-neurons. A corresponding staining pattern was found using the second method with a polyclonal antiserum against choline acetyltransferase (ChAT). The third method was the histochemical detection at the electron microscope level of acetylcholinesterase, the enzyme responsible for the breakdown of acetylcholine. We found that this enzyme is located in synaptic clefts of L-neurons in both of the brain regions where L-neurons are known to make excitatory and inhibitory output synapses. Acetylcholinesterase was confined to synaptic sites, which is consistent with a role in synaptic transmission at these synapses. Taken together, the findings suggest that L-neurons use acetylcholine as a neurotransmitter.