Detecting Physiological Responses from Sensory Neurons in Drosophila

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Samenvatting

Source: Shankar, S., et al. Measuring Physiological Responses of Drosophila Sensory Neurons to Lipid Pheromones Using Live Calcium Imaging. J. Vis. Exp. (2016).

This video demonstrates a method for detecting sensory neuron responses in Drosophila using ligand binding and fluorescence imaging. Ligands bind to neuron receptors, causing calcium influx. The calcium binds to indicator complexes, leading to fluorescence emission, which confirms the neuronal response to the ligand.

Protocol

1. Sample Preparation

Cross the Gal4 driver line to UAS-GCaMP5G Drosophila melanogaster flies (w;P{20XUAS-IVS-GCaMP5G}attP40). Allow the cross to grow at 25 °C.
NOTE: Generating flies with multiple copies of the Gal4 or UAS-GCaMP transgenes can help to enhance the fluorescent signal intensity. An earlier version of the UAS-GCaMP transgene (UAS-GCaMP3) showed very weak fluorescence intensity when expressed under the control of the Gr68a-Gal4 driver.
1. Collect and separate by sex newly eclosed adult flies and raise at 25 °C.

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Resultaten

Fruit fly restraint techniques; legs on tape; microscopy of tarsal segments; pipette interaction.
Figure 1: Mounting a live fly for imaging of the foreleg or proboscis neurons. (A) Mounting the foreleg: a 3.2X image of a live male fly, showing the forelegs fastened to the coverslip with tape. (B) A 40X image of the fly shows each of the tarsal segments.......