Ex Vivo Calcium Imaging to Study Drosophila Brain Responses to Neuropeptides

Begin with an imaging chamber containing an immobilized Drosophila brain explant submerged in a buffer to maintain cellular function.

The neurons in the brain express cytoplasmic fluorescent calcium indicators, which bind to calcium ions and fluoresce.

Place the imaging chamber under a fluorescence microscope equipped with a water-immersion lens.

Lower the lens, and under bright-field illumination, focus on the brain.

Switch to fluorescence mode and capture the image.

Next, introduce a neuropeptide into the buffer and initiate time-lapse recording.

The neuropeptide binds to its receptor on neurons, triggering intracellular signaling. This opens the calcium channels and allows the calcium ion influx.

These calcium ions bind to the indicators and increase fluorescence intensity.

Recapture the image after neuropeptide stimulation.

Compare the images before and after neuropeptide stimulation.

An increase in fluorescence intensity after neuropeptide stimulation reflects the Drosophila brain's response to neuropeptide signaling.