Optogenetics pp 133-147 | Cite as

Optogenetic Manipulation and Probing

  • Masamichi OhkuraEmail author
  • Junko Sadakari
  • Junichi Nakai


Controlling and monitoring the activities of defined cell populations should provide powerful methodologies for better understanding individual cellular functions in vivo. To enable the control and monitoring of cellular activities, ‘photo-actuator molecules’ and ‘fluorescent probe molecules’ have been generated, respectively. Photo-actuators are the motor molecules that can trigger cellular activities by photo-activation of specific intracellular molecules, and fluorescent probes are the molecules utilized to detect cellular activities by emitting fluorescence upon binding to their specific target structures of intracellular molecules. These actuators and probes are also known as ‘optogenetic tools’, and they can be expressed in specific cells or specific organelles for a long period, because they are genetically encoded. In recent years, the development and improvement of optogenetic tools has progressed rapidly. Researchers can now choose optogenetic tools that better suit their needs. In this review, we describe the history, species, and development of optogenetic tools, and future issues, limiting the definition of optogenetic tools to those based on proteins.


Optogenetic tools Photo-actuator molecules Fluorescent probes Channelrhodopsin (ChR) variants Halorhodopsin (NpHR/halo)/archaerhodopsin (Arch) variants Light-driven G-protein-coupled receptor (opto-XR) variants Ca2+ probes Voltage probes H+ probes Cl probes 


  1. Airan RD, Thompson KR, Fenno LE et al (2009) Temporally precise in vivo control of intracellular signalling. Nature 458:1025–1029PubMedCrossRefGoogle Scholar
  2. Akerboom J, Chen TW, Wardill TJ et al (2012) Optimization of a GCaMP calcium indicator for neural activity imaging. J Neurosci 32:13819–13840PubMedCentralPubMedCrossRefGoogle Scholar
  3. Ataka K, Pieribone VA (2002) A genetically targetable fluorescent probe of channel gating with rapid kinetics. Biophys J 82:509–516PubMedCentralPubMedCrossRefGoogle Scholar
  4. Baird GS, Zacharias DA, Tsien RY (1999) Circular permutation and receptor insertion within green fluorescent proteins. Proc Natl Acad Sci U S A 96:11241–11246PubMedCentralPubMedCrossRefGoogle Scholar
  5. Berndt A, Yizhar O, Gunaydin LA et al (2009) Bi-stable neural state switches. Nat Neurosci 12:229–234PubMedCrossRefGoogle Scholar
  6. Boyden ES, Zhang F, Bamberg E et al (2005) Millisecond-timescale, genetically targeted optical control of neural activity. Nat Neurosci 8:1263–1268PubMedCrossRefGoogle Scholar
  7. Chen X, Leischner U, Rochefort NL et al (2011) Functional mapping of single spines in cortical neurons in vivo. Nature 475:501–505PubMedCrossRefGoogle Scholar
  8. Chow BY, Han X, Dobry AS et al (2010) High-performance genetically targetable optical neural silencing by light-driven proton pumps. Nature 463:98–102PubMedCentralPubMedCrossRefGoogle Scholar
  9. Han X, Boyden ES (2007) Multiple-color optical activation, silencing, and desynchronization of neural activity, with single-spike temporal resolution. PLoS One 2:e299. doi: 10.1371/journal.pone.0000299 PubMedCentralPubMedCrossRefGoogle Scholar
  10. Heim N, Griesbeck O (2004) Genetically encoded indicators of cellular calcium dynamics based on troponin C and green fluorescent protein. J Biol Chem 279:14280–14286PubMedCrossRefGoogle Scholar
  11. Hochbaum DR, Zhao Y, Farhi SL et al (2014) All-optical electrophysiology in mammalian neurons using engineered microbial rhodopsins. Nat Methods 11:825–833PubMedCentralPubMedCrossRefGoogle Scholar
  12. Horikawa K, Yamada Y, Matsuda T et al (2010) Spontaneous network activity visualized by ultrasensitive Ca2+ indicators, yellow Cameleon-Nano. Nat Methods 7:729–732PubMedCrossRefGoogle Scholar
  13. Inoue M, Takeuchi A, Horigane S et al (2015) Rational design of a high-affinity, fast, red calcium indicator R-CaMP2. Nat Methods 12:64–70Google Scholar
  14. Kralj JM, Douglass AD, Hochbaum DR et al (2011) Optical recording of action potentials in mammalian neurons using a microbial rhodopsin. Nat Methods 9:90–95PubMedCentralPubMedCrossRefGoogle Scholar
  15. Kuner T, Augustine GJ (2000) A genetically encoded ratiometric indicator for chloride: capturing chloride transients in cultured hippocampal neurons. Neuron 27:447–459PubMedCrossRefGoogle Scholar
  16. Li H, Foss SM, Dobryy YL et al (2011) Concurrent imaging of synaptic vesicle recycling and calcium dynamics. Front Mol Neurosci 4:34. doi: 10.3389/fnmol.2011.00034 PubMedCentralPubMedCrossRefGoogle Scholar
  17. Miesenböck G, De Angelis DA, Rothman JE (1998) Visualizing secretion and synaptic transmission with pH-sensitive green fluorescent proteins. Nature 394:192–195PubMedCrossRefGoogle Scholar
  18. Miyawaki A, Llopis J, Heim R et al (1997) Fluorescent indicators for Ca2+ based on green fluorescent proteins and calmodulin. Nature 388:882–887PubMedCrossRefGoogle Scholar
  19. Murata Y, Iwasaki H, Sasaki M et al (2005) Phosphoinositide phosphatase activity coupled to an intrinsic voltage sensor. Nature 435:1239–1243PubMedCrossRefGoogle Scholar
  20. Mutoh H, Akemann W, Knöpfel T (2012) Genetically engineered fluorescent voltage reporters. ACS Chem Neurosci 3:585–592PubMedCentralPubMedCrossRefGoogle Scholar
  21. Nagai T, Sawano A, Park ES et al (2001) Circularly permuted green fluorescent proteins engineered to sense Ca2+. Proc Natl Acad Sci U S A 98:3197–3202PubMedCentralPubMedCrossRefGoogle Scholar
  22. Nagel G, Szellas T, Huhn W et al (2003) Channelrhodopsin-2, a directly light-gated cation-selective membrane channel. Proc Natl Acad Sci U S A 100:13940–13945PubMedCentralPubMedCrossRefGoogle Scholar
  23. Nakai J, Ohkura M, Imoto K (2001) A high signal-to-noise Ca2+ probe composed of a single green fluorescent protein. Nat Biotechnol 19:137–141PubMedCrossRefGoogle Scholar
  24. Oh E, Maejima T, Liu C et al (2010) Substitution of 5-HT1A receptor signaling by a light-activated G protein-coupled receptor. J Biol Chem 285:30825–30836Google Scholar
  25. Ohkura M, Matsuzaki M, Kasai H et al (2005) Genetically encoded bright Ca2+ probe applicable for dynamic Ca2+ imaging of dendritic spines. Anal Chem 77:5861–5869PubMedCrossRefGoogle Scholar
  26. Ohkura M, Sasaki T, Kobayashi C et al (2012a) An improved genetically encoded red fluorescent Ca2+ indicator for detecting optically evoked action potentials. PLoS One 7:e39933. doi: 10.1371/journal.pone.0039933 PubMedCentralPubMedCrossRefGoogle Scholar
  27. Ohkura M, Sasaki T, Sadakari J et al (2012b) Genetically encoded green fluorescent Ca2+ indicators with improved detectability for neuronal Ca2+ signals. PLoS One 7:e51286. doi: 10.1371/journal.pone.0051286 PubMedCentralPubMedCrossRefGoogle Scholar
  28. Sakai R, Repunte-Canonigo V, Raj CD et al (2001) Design and characterization of a DNA-encoded, voltage-sensitive fluorescent protein. Eur J Neurosci 13:2314–2318PubMedCrossRefGoogle Scholar
  29. Sankaranarayanan S, Ryan TA (2001) Calcium accelerates endocytosis of vSNAREs at hippocampal synapses. Nat Neurosci 4:129–136PubMedCrossRefGoogle Scholar
  30. Siegel MS, Isacoff EY (1997) A genetically encoded optical probe of membrane voltage. Neuron 19:735–741PubMedCrossRefGoogle Scholar
  31. Takahashi N, Kitamura K, Matsuo N et al (2012) Locally synchronized synaptic inputs. Science 335:353–356PubMedCrossRefGoogle Scholar
  32. Tallini YN, Ohkura M, Choi BR et al (2006) Imaging cellular signals in the heart in vivo: cardiac expression of the high-signal Ca2+ indicator GCaMP2. Proc Natl Acad Sci U S A 103:4753–4758PubMedCentralPubMedCrossRefGoogle Scholar
  33. Tian L, Hires SA, Mao T et al (2009) Imaging neural activity in worms, flies and mice with improved GCaMP calcium indicators. Nat Methods 6:875–881PubMedCentralPubMedCrossRefGoogle Scholar
  34. Zhang F, Wang LP, Brauner M et al (2007) Multimodal fast optical interrogation of neural circuitry. Nature 446:633–639PubMedCrossRefGoogle Scholar
  35. Zhao Y, Araki S, Wu J et al (2011) An expanded palette of genetically encoded Ca2+ indicators. Science 333:1888–1891PubMedCentralPubMedCrossRefGoogle Scholar
  36. Zoltowski BD, Schwerdtfeger C, Widom J et al (2007) Conformational switching in the fungal light sensor Vivid. Science 316:1054–1057PubMedCentralPubMedCrossRefGoogle Scholar

Copyright information

© Springer Japan 2015

Authors and Affiliations

  • Masamichi Ohkura
    • 1
    Email author
  • Junko Sadakari
    • 1
  • Junichi Nakai
    • 1
  1. 1.Saitama University Brain Science InstituteSaitamaJapan

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