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The identification of neurotransmitter type used by a neuron is important for the functional dissection of neuronal circuits. In the model organism Drosophila melanogaster , several methods for discerning the neurotransmitter systems are available. Here, we expanded the toolbox for the identification of cholinergic neurons by generating a new line FRT-STOP-FRT-VAChT::HA that is a conditional tagged knock-in of the vesicular acetylcholine transporter ( VAChT ) gene in its endogenous locus. Importantly, in comparison to already available tools for the detection of cholinergic neurons, the FRT-STOP-FRT-VAChT::HA allele also allows for identification of the subcellular localization of the cholinergic presynaptic release sites in a cell-specific manner. We used the newly generated FRT-STOP-FRT-VAChT::HA line to characterize the Mi1 and Tm3 neurons in the fly visual system and found that VAChT is present in the axons of both cell types, suggesting that Mi1 and Tm3 neurons provide cholinergic input to the elementary motion detectors, the T4 neurons.

Transgenic line for the identification of cholinergic release sites inDrosophila melanogaster