Figure 1.
Forward Appetitive Conditioning Produces a Cell-Wide, Calcium-Based Memory Trace in the DPM Neuron Processes.
(A) Diagram illustrating the appetitive olfactory conditioning protocols used in this study. Starved flies were trained using forward, unpaired or CS-only conditioning. Naïve animals were used as control. The DPM neuron responses were subsequently recorded by presenting each odor for 3 sec separated by 3 min.
(B) Image of the DPM neuron innervation of the MB neuropil indicating the regions of interest subjected to imaging. The left panel illustrates the lobes of the MBs with the planes of imaging outlined. The right panels illustrate the regions of interest as viewed from the dorsal perspective.
(C) Time course of the fluorescence response of G-CaMP to Oct and Ben in the vertical branch of the DPM neuron calculated as the %ΔF/Fo. The shaded area around each line indicates the standard error of the mean across time.
(D) Bar graph of the response ratio of the CS+/CS− as a function of time after appetitive conditioning measured in the DPM vertical branch neurons after forward (f-Oct+, f-Ben+), unpaired, CS-only, and naïve conditioning. A robust increase in calcium influx was detected in the vertical branch at 30–50 min after forward conditioning with either CS+ odor. The enhanced response to the trained odor remained significant at 50–70 min, 70–90 min and 120–150 min after conditioning. The increased was not observed after unpaired or CS-only conditioning, or in naïve flies. Asterisks indicate a statistically significant difference as determined by Kruskal-Wallis analysis followed by Mann-Whitney pairwise comparisons (p≤0.02). n=8–12 for all groups.
(E) Bar graph of the response ratio of the CS+/CS− after appetitive conditioning measured horizontal branch of the DPM neurons. An increase in calcium was detected in the horizontal branch at 30–50, 50–70, 70–90 and 120–150 min after forward conditioning. Asterisks indicate a statistically significant difference as assessed by Kruskal-Wallis analysis followed by Mann-Whitney pairwise comparisons (p≤0.0031). n=6–10 for all groups.
(F) The DPM neuron vertical branch exhibited an increased response to the trained odor after appetitive and aversive conditioning. This increased response was observed with both training protocols at 30–50 and 50–70 min after training. In contrast with the aversive memory trace, the appetitive memory trace persisted in the vertical lobes during the 70–90 and 120–150 min time windows and became undetectable by 180–210 min. Asterisks indicate statistically significant differences as determined with Kruskal-Wallis analysis followed by Mann-Whitney pairwise comparisons (p≤0.04). n=8–9 for all groups.
(G) The DPM neuron horizontal branch exhibited an increased response to the trained odor after appetitive but not aversive conditioning. Appetitive conditioning induced an increased response to the trained odor in the horizontal branch at 30–50, 50–70, 70–90, and 120–150 min after conditioning. In contrast, aversive olfactory conditioning failed to induce a memory trace in the DPM horizontal branch. Asterisks indicate statistically significant differences as determined with Kruskal-Wallis analysis followed by Mann-Whitney pairwise comparisons (p≤0.009). n=6–15 for all groups. For all panels, error bars indicate the standard error of the mean. See also Figure S1.