Abstract
The fruit fly Drosophila melanogaster, an insect 4 mm long, has served as the experimental subject in a wide range of biological research, including neuroscience. In this chapter, we briefly introduce optogenetic applications in Drosophila neuroscience research. First, we describe the development of Drosophila from egg to adult. In fly neuroscience, temperature-controlled perturbation of neural activity, sometimes called “thermogenetics,” has been an invaluable tool that predates the advent of optogenetics. After briefly introducing this perturbation technique, we describe the process of generating transgenic flies that express optogenetic probes in a specific group of cells. Transgenic techniques are crucial in the application of optogenetics in Drosophila neuroscience; here we introduce the transposon P-elements, ϕC31 integrase, and CRISPR-Cas9 methods. As for cell-specific gene expression techniques, the binary expression systems utilizing Gal4-UAS, LexA-lexAop, and Q-system are described. We also present a short and basic optogenetic experiment with Drosophila larvae as a practical example. Finally, we review a few recent studies in Drosophila neuroscience that made use of optogenetics. In this overview of fly development, transgenic methods, and applications of optogenetics, we present an introductory background to optogenetics in Drosophila.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
Similar content being viewed by others
Abbreviations
- ATR:
-
All-Trans Retinal
- attB:
-
the Bacterial attachment site
- attP:
-
the Phage attachment site
- Cas9:
-
CRISPR Associated protein 9
- ChR2:
-
Channelrhodopsin 2
- CRISPR:
-
Clustered Regularly Interspaced Short Palindromic Repeats
- FRT:
-
Flippase Recognition Target
- GDLs:
-
GABAergic DorsoLateral neurons
- GFP:
-
Green Fluorescent Protein
- PMSIs:
-
Period-positive Median Segmental Interneurons
- TRP:
-
Transient Receptor Potential
- UAS:
-
the Upstream Activation Sequence
References
Aberle H, Haghighi AP, Fetter RD, McCabe BD, Magalhães TR, Goodman CS (2002) Wishful thinking encodes a BMP type II receptor that regulates synaptic growth in drosophila. Neuron 33(4):545–558. https://doi.org/10.1016/S0896-6273(02)00589-5
Bellen HJ, Levis RW, He Y, Carlson JW, Evans-Holm M, Bae E, Kim J, Metaxakis A, Savakis C, Schulze KL, Hoskins RA, Spradling AC (2011) The drosophila gene disruption project: progress using transposons with distinctive site specificities. Genetics 188(3):731–743. https://doi.org/10.1534/genetics.111.126995
Bellen HJ, Tong C, Tsuda H (2010) A history lesson for the future. Neuroscience 11(juLy):514–522. https://doi.org/10.1038/nrn2839
Brand AH, Perrimon N (1993) Targeted gene expression as a means of altering cell fates and generating dominant phenotypes. Development 118(2):401–415
Chen M, Obar R, Schroeder C (1991) Multiple forms of dynamin are encoded by shibire, a Drosophila gene involved in endocytosis. Nature 351(6327):583–586. https://www.nature.com/articles/382731a0.pdf
Fushiki A, Zwart MF, Kohsaka H, Fetter RD, Cardona A, Nose A (2016) A circuit mechanism for the propagation of waves of muscle contraction in Drosophila. elife 5:1–23. https://doi.org/10.7554/elife.13253
Gepner R, Skanata MM, Bernat NM, Kaplow M, Gershow M (2015) Computations underlying Drosophila photo- taxis, odor-taxis, and multi-sensory integration. elife 4:1–21. https://doi.org/10.7554/eLife.06229
Gratz SJ, Cummings AM, Nguyen JN, Hamm DC, Donohue LK, Harrison MM, Wildonger J, O’connor-Giles KM (2013) Genome engineering of Drosophila with the CRISPR RNA-guided Cas9 nuclease. Genetics 194(4):1029–1035. https://doi.org/10.1534/genetics.113.152710
Grigliatti T, Hall L, Rosenbluth R, Suzuki D (1973) Temperature-sensitive mutations in Drosophila melanogaster. Mol Gen Genomics 120:107–114. https://doi.org/10.1126/science.170.3959.695
Groth AC, Fish M, Nusse R, Calos MP (2004) Construction of transgenic drosophila by using the site-specific integrase from phage φC31. Genetics 166(4):1775–1782. https://doi.org/10.1534/genetics.166.4.1775
Hamada FN, Rosenzweig M, Kang K, Pulver SR, Ghezzi A, Jegla TJ, Garrity PA (2008) An internal thermal sensor controlling temperature preference in Drosophila. Nature 454(7201):217–220. https://doi.org/10.1038/nature07001
Hernandez-Nunez L, Belina J, Klein M, Si G, Claus L, Carlson JR, Samuel ADT (2015) Reverse-correlation analysis of navigation dynamics in Drosophila larva using optogenetics. elife 4:1–16. https://doi.org/10.7554/eLife.06225
Katow H, Takahashi T, Saito K, Tanimoto H, Kondo S (2019) Tango knock-ins visualize endogenous activity of G protein-coupled receptors in Drosophila. J Neurogenet 33(2):44–51. https://doi.org/10.1080/01677063.2019.1611806
Kitamoto T (2001) Conditional modification of behavior in drosophila by targeted expression of a temperature-sensitive shibire allele in defined neurons. J Neurobiol 47(2):81–92. https://doi.org/10.1002/neu.1018
Klapoetke NC, Murata Y, Kim SS, Pulver SR, Birdsey-Benson A, Cho YK, Morimoto TK, Chuong AS, Carpenter EJ, Tian Z, Wang J, **e Y, Yan Z, Zhang Y, Chow BY, Surek B, Melkonian M, Jayaraman V, Constantine-Paton M et al (2014) Independent optical excitation of distinct neural populations. Nat Methods 11(3):338–346. https://doi.org/10.1038/nmeth.2836
Kohsaka H, Okusawa S, Itakura Y, Fushiki A, Nose A (2012) Development of larval motor circuits in Drosophila. Dev Growth Differ 54(3):408–419. https://doi.org/10.1111/j.1440-169X.2012.01347.x
Kohsaka H, Takasu E, Morimoto T, Nose A (2014) A group of segmental premotor interneurons regulates the speed of axial locomotion in drosophila larvae. Curr Biol 24(22):2632–2642. https://doi.org/10.1016/j.cub.2014.09.026
Kondo S, Ueda R (2013) Highly improved gene targeting by germline-specific Cas9 expression in Drosophila. Genetics 195(3):715–721. https://doi.org/10.1534/genetics.113.156737
Kosaka T, Ikeda K (1983) Possible temperature-dependent blockage of synaptic vesicle recycling induced by a single gene mutation in drosophila. J Neurobiol 14(3):207–225. https://doi.org/10.1002/neu.480140305
Lai SL, Lee T (2006) Genetic mosaic with dual binary transcriptional systems in drosophila. Nat Neurosci 9(5):703–709. https://doi.org/10.1038/nn1681
Levis R, Hazelrigg T, Rubin GM (1985) Effects of genomic position on the expression of transduced copies of the white gene of Drosophila. Science 229(4713):558–561. https://doi.org/10.1126/science.2992080
Li-Kroeger D, Kanca O, Lee PT, Cowan S, Lee MT, Jaiswal M, Salazar JL, He Y, Zuo Z, Bellen HJ (2018) An expanded toolkit for gene tagging based on MiMIC and scarless CRISPR tagging in Drosophila. elife 7:1–27. https://doi.org/10.7554/eLife.38709
Lin JY, Knutsen PM, Muller A, Kleinfeld D, Tsien RY (2013) ReaChR: a red-shifted variant of channelrhodopsin enables deep transcranial optogenetic excitation. Nat Neurosci 16(10):1499–1508. https://doi.org/10.1038/nn.3502
Markstein M, Pitsouli C, Villalta C, Celniker SE, Perrimon N (2008) Exploiting position effects and the gypsy retrovirus insulator to engineer precisely expressed transgenes. Nat Genet 40(4):476–483. https://doi.org/10.1038/ng.101
Matsunaga T, Fushiki A, Nose A, Kohsaka H (2013) Optogenetic perturbation of neural activity with laser illumination in semi-intact drosophila larvae in motion. J Vis Exp 77:1–5. https://doi.org/10.3791/50513
McKemy DD, Neuhausser WM, Julius D (2002) Identification of a cold receptor reveals a general role for TRP channels in thermosensation. Nature 416(6876):52–58. https://doi.org/10.1038/nature719
Mohr SE, Hu Y, Ewen-Campen B, Housden BE, Viswanatha R, Perrimon N (2016) CRISPR guide RNA design for research applications. FEBS J 283:3232–3238. https://doi.org/10.1111/febs.13777
Moreira JM, Itskov PM, Goldschmidt D, Baltazar C, Steck K, Tastekin I, Walker SJ, Ribeiro C (2019) optoPAD, a closed-loop optogenetics system to study the circuit basis of feeding behaviors. elife 8:1–19. https://doi.org/10.7554/eLife.43924
Peabody NC, Pohl JB, Diao F, Vreede AP, Sandstrom DJ, Wang H, Zelensky PK, White BH (2009) Characterization of the decision network for wing expansion in drosophila using targeted expression of the TRPM8 channel. J Neurosci 29(11):3343–3353. https://doi.org/10.1523/JNEUROSCI.4241-08.2009
Pfeiffer B, Jenett A, Hammonds A, Ngo T, Misra S, Murphy C, Scully A, Carlson J, Wan K, Laverty T, Mungall C, Svirskas R, Kadonaga J, Doe C, Eisen M, Rubin G, Janelia (2008) Tools for neuroanatomy and neurogenetics in drosophila. Proc Natl Acad Sci 105(28):9715–9720. https://doi.org/10.5694/j.1326-5377.1975.tb111372.x
Pfeiffer BD, Ngo TTB, Hibbard KL, Murphy C, Jenett A, Truman JW, Rubin GM (2010) Refinement of tools for targeted gene expression in drosophila. Genetics 186(2):735–755. https://doi.org/10.1534/genetics.110.119917
Potter CJ, Tasic B, Russler EV, Liang L, Luo L (2010) The Q system: a repressible binary system for transgene expression, lineage tracing, and mosaic analysis. Cell 141(3):536–548. https://doi.org/10.1016/j.cell.2010.02.025
Pulver SR, Pashkovski SL, Hornstein NJ, Garrity PA, Griffith LC (2009) Temporal dynamics of neuronal activation by channelrhodopsin-2 and TRPA1 determine behavioral output in Drosophila larvae. J Neurophysiol 101(6):3075–3088. https://doi.org/10.1152/jn.00071.2009
Riabinina O, Luginbuhl D, Marr E, Liu S, Wu MN, Luo L, Potter CJ (2015) Improved and expanded Q-system reagents for genetic manipulations. Nat Methods 12(3):219–222. https://doi.org/10.1038/nmeth.3250
Riabinina O, Vernon SW, Dickson BJ, Baines RA (2019) Split-QF system for fine-tuned transgene expression in Drosophila. Genetics 212(1):53–63. https://doi.org/10.1534/genetics.119.302034
Rubin GM, Spradling AC (1982) Genetic transformation of Drosophila with transposable element vectors. Science 218(4570):348–353. https://doi.org/10.1126/science.6289436
Shearin HK, Dvarishkis AR, Kozeluh CD, Stowers RS (2013) Expansion of the Gateway MultiSite Recombination Cloning Toolkit. PLoS One 8(10):1–14. https://doi.org/10.1371/journal.pone.0077724
Simpson JH, Looger LL (2018) Functional imaging and optogenetics in Drosophila. Genetics 208:1291–1309
Thum A, Knapek S, Rister J, Dierichs-Schmitt E, Heisenberg M, Tanimoto H (2006) Differential potencies of effector genes in adult Drosophila. Comp Gen Pharmacol 498:194–2003. https://doi.org/10.1002/cne
Ting CY, Gu S, Guttikonda S, Lin TY, White BH, Lee CH (2011) Focusing transgene expression in drosophila by coupling Gal4 with a Novel Split-Lex A expression system. Genetics 188(1):229–233. https://doi.org/10.1534/genetics.110.126193
Tomasiunaite U, Widmann A, Thum AS (2018) Maggot instructor: semi-automated analysis of learning and memory in Drosophila larvae. Front Psychol 9:1–18. https://doi.org/10.3389/fpsyg.2018.01010
Van Der Bliek AM, Meyerowrtz EM (1991) Dynamin-like protein encoded by the Drosophila shibire gene associated with vesicular traffic. Nature 351(6325):411–414. https://doi.org/10.1038/351411a0
Venken KJT, Simpson JH, Bellen HJ (2011) Genetic manipulation of genes and cells in the nervous system of the fruit fly. Neuron 72(2):202–230. https://doi.org/10.1016/j.neuron.2011.09.021
Wiedenheft B, Sternberg SH, Doudna JA (2012) RNA-guided genetic silencing systems in bacteria and archaea. Nature 482(7385):331–338. https://doi.org/10.1038/nature10886
**ang Y, Yuan Q, Vogt N, Looger LL, Jan LY, Jan YN (2010) Light-avoidance-mediating photoreceptors tile the Drosophila larval body wall. Nature 468(7326):921–926. https://doi.org/10.1038/nature09576
Acknowledgments
We thank Ms. Kasumi Shibahara for help with preparing the figures. This work was supported by MEXT/JSPS KAKENHI grants (22115002, 15H04255, 221S0003 to A.N. and 26430004, 17 K07042, 20 K06908 to H.K.).
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2021 Springer Nature Singapore Pte Ltd.
About this chapter
Cite this chapter
Kohsaka, H., Nose, A. (2021). Optogenetics in Drosophila. In: Yawo, H., Kandori, H., Koizumi, A., Kageyama, R. (eds) Optogenetics. Advances in Experimental Medicine and Biology, vol 1293. Springer, Singapore. https://doi.org/10.1007/978-981-15-8763-4_19
Download citation
DOI: https://doi.org/10.1007/978-981-15-8763-4_19
Published:
Publisher Name: Springer, Singapore
Print ISBN: 978-981-15-8762-7
Online ISBN: 978-981-15-8763-4
eBook Packages: Biomedical and Life SciencesBiomedical and Life Sciences (R0)