Drosophila Neuromuscular Junction (NMJ) Quantification: A Method to Assess Synaptic Morphology and Function

- First, immunostain Drosophila larvae with markers that highlight different aspects of NMJs or neuromuscular junctions. This is the region where the ending of a motor neuron axon contacts a muscle, and its morphology is used as a readout of synaptic function.

The axon terminates in several branches that form bulges called boutons. Neurotransmitters are released from boutons and cause muscle contraction. Areas of neurotransmitter release are called active zones. Here one marker is specific for a synaptic protein that outlines the NMJ, and the other marker is for a protein present in active zones.

Next, acquire NMJ images using microscopy and quantitatively assess their morphology with semi-automated image analysis software. Apply a preset series of software commands to the images. The output files contain analyzed NMJ images and morphology results. The features that can be measured include number and surface area of boutons, NMJ length and branch number, number of unconnected NMJ compartments, and number of active zones.

In the following example, we will analyze the morphology of Drosophila NMJs stained with DLG1, a postsynaptic marker, and BRP, an active zone marker.

- For this pr