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. 2020 Nov 6;9:e62297. doi: 10.7554/eLife.62297

Figure 3. Adult flight phenotypes arise from late pupal expression of the IP3R and mAChR.

(A) Box plots of flight bout durations in adults after temporal expression of IP3RDN in THD’ neurons (THD’; TubGAL80ts > ItprDN) by transferring them to 29°C at the indicated stages of development. For adult expression, flies were transferred to 29°C immediately after eclosion and tested for flight after 3 days. (B) Box plots of flight bout durations after expressing the IP3R (Itpr+) in dopaminergic cells of itpr mutants at the indicated stages of development. For stage specific expression a TH; TubGAL80ts strain was used and the progeny transferred to 29°C at appropriate time developmental points. (C) Levels of the IP3R increase in late pupae between 72-96 hrs APF and plateau between 96hrs APF and adults (3 days). A representative western blot from lysates of dissected central nervous systems of Canton S probed with anti-IP3R and anti-spectrin is shown (top). Three independent lysates and blots were quantified (below). **p< 0.05, t-test. (D) Box plots with flight bout durations of adult flies after knockdown of the Muscarinic acetylcholine receptor (mAChR) in THD’ neurons (THD’;TubGAL80ts > dsmAChR) at the indicated stages of pupal development and in adults.Box plot symbols are as described in methods, n≥30, ***p< 0.005 *p< 0.05, n.s., not significant by Kruskal-Wallis test (for A and D) and n≥23, ***p< 0.005, at p < 0.05 by Mann-Whitney U test (for B). All comparisons for significance were with the control values for A and C, with itpr mutants for B.

Figure 3—source data 1. Flight duration of flies upon overexpression of IP3RDN at different stages of development.
Figure 3—source data 2. Flight duration of itpr mutant flies after overexpressing a wild type itpr transgene at different stages of development.
Figure 3—source data 3. Western blots of IP3R.
Figure 3—source data 4. Flight duration of flies upon knockdown of mAChR in dopaminergic neurons at different stages of development.
Figure 3—source data 5. Flight duration of flies upon knockdown of mAChR in PPL1-γ2α′1 neurons.

Figure 3.

Figure 3—figure supplement 1. Expression of IP3RDN and mAChR RNAi in THD’ and MB296B marked neurons.

Figure 3—figure supplement 1.

(A) Expression of IP3RDN does not change the number of PPL1 neurons. Cells were quantified from both hemi-segments of N=6 brains. (B) PPL1- γ2α′1 neurons marked by MB296BGAL4 are shown (white arrows) with their pattern of innervation to the MB neuropil as visualized using membrane-localized GFP. Expression of IP3RDN (lower panel) did not affect the pattern of MB innervation. (C) Box plots of flight bout durations in control flies and flies with knockdown of mAChR in MB296B cells. n≥ 30, **p< 0.05 by Kruskal-Wallis test.