Functional Imaging of Antennal Lobe Neurons in Drosophila with Synapto-pHluorin

A method for imaging the synaptic activity of antennal lobe neurons in the Drosophila brain was developed to visualize and study cellular memory traces. Cellular memory traces are defined as any change in the activity of a neuron that is induced by learning, which subsequently alters the processing and response of the nervous system to the sensory information that is learned. Synapto-pHluorin (spH), a protein fusion of a pH-sensitive mutant of green fluorescent protein with synaptobrevin, was expressed in the projection neurons of flies. This protein, by virtue of its pH sensitivity, registers synaptic activity with increased fluorescence due to its movement from the lumen side of the synaptic vesicle to the synaptic cleft upon vesicle release. The steps required to measure synaptic activity using this fluorescent reporter in living flies in response to sensory cues and learning include: (McGuire et al., Prog in Neurobiol 76:328–347, 2005) microsurgery of transgenic flies to visualize spH fluorescence, (Davis, Annu Rev Neurosci 28:275–302, 2005) application of odor and electric shock cues to the fly being imaged, (Keene and Waddell, Nat Rev Neurosci 8:341–354, 2007) microscopic monitoring of the fluorescence before, during, and after the presentation of sensory cues, and [Miesenb�ck, Nature 394:192–195, 1998] analyzing the changes in fluorescence across time and space. One example of how this methodology has revealed insights into learning processes is discussed. The pairing of an odor (conditioned stimulus, CS) with electric shock pulses (unconditioned stimulus, US) produced a change in the synaptic response of PNs to the trained odor.