Visualization of Synapses in Larval Stages of Drosophila melanogaster Using the GRASP Technique

The flow of information within the nervous system occurs via precise connections between synaptic partners. In recent years, the development of various methods to visualize synaptic contacts has helped elucidate the connectivity within complex neuronal networks. One such method is the GRASP (GFP Reconstitution Across Synaptic Partners) technique that consists of the expression of a portion of the green fluorescent protein (GFP) at each side of the synapse, allowing the reconstitution of green fluorescence depending on the proximity of the cells expressing such tools. In Drosophila, various studies have shown the successful application of GRASP in adult flies to identify synaptic partners, whereas its use at earlier stages such as in first instar larval stages remains less common. Therefore, we provide here a detailed protocol for the visualization of GRASP-based neuronal contacts within previously established synaptic partners in first and third instar larvae.