Neuronal firing modulation by a membrane-targeted photoswitch


Optical technologies allowing modulation of neuronal activity at high spatio-temporal resolution are becoming paramount in neuroscience. In this respect, azobenzene-based photoswitches are promising nanoscale tools for neuronal photostimulation. Here we engineered a light-sensitive azobenzene compound (Ziapin2) that stably partitions into the plasma membrane and causes its thinning through trans-dimerization in the dark, resulting in an increased membrane capacitance at steady state. We demonstrated that in neurons loaded with the compound, millisecond pulses of visible light induce a transient hyperpolarization followed by a delayed depolarization that triggers action potential firing. These effects are persistent and can be evoked in vivo up to 7 days, proving the potential of Ziapin2 for the modulation of membrane capacitance in the millisecond timescale, without directly affecting ion channels or local temperature.

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.


All prices are NET prices.

Fig. 1: Transcis isomerization of Ziapin2 in various environments.
Fig. 2: Ziapin2 distributes to the plasma membrane and lipid rafts in neurons.
Fig. 3: Ziapin2 reversibly modifies membrane thickness in artificial membranes, cell lines and neurons.
Fig. 4: Light-evoked membrane voltage modulation by Ziapin2 in primary neurons.
Fig. 5: Light-evoked firing activity in primary neurons loaded with Ziapin2.
Fig. 6: Light-evoked cortical responses in vivo in mice loaded with Ziapin2 in the somatosensory cortex.

Data availability

The datasets generated and analysed during the current study are available from the corresponding author on reasonable request.


  1. 1.

    Paoletti, P., Ellis-Davies, G. C. R. & Mourot, A. Optical control of neuronal ion channels and receptors. Nat. Rev. Neurosci. 20, 514–532 (2019).

    CAS  Google Scholar 

  2. 2.

    Rivnay, J., Wang, H., Fenno, L., Deisseroth, K. & Malliaras, G. Next-generation probes, particles, and proteins for neural interfacing. Sci. Adv. 3, e1601649 (2017).

    Google Scholar 

  3. 3.

    Tønnesen, J. Optogenetic cell control in experimental models of neurological disorders. Behav. Brain Res. 255, 35–43 (2013).

    Google Scholar 

  4. 4.

    Deisseroth, K. Optogenetics: 10 years of microbial opsins in neuroscience. Nat. Neurosci. 18, 1213–1225 (2015).

    CAS  Google Scholar 

  5. 5.

    Zhang, J. J., Wang, J. X. & Tia, H. Taking orders from light: progress in photochromic bio-materials. Mater. Horiz. 1, 169–184 (2014).

    CAS  Google Scholar 

  6. 6.

    Fujiwara, H. & Yonezawa, Y. Photoelectric response of a black lipid membrane containing an amphiphilic azobenzene derivative. Nature 351, 724–726 (1991).

    CAS  Google Scholar 

  7. 7.

    Yonezawa, Y., Fujiwara, H. & Sato, T. Photoelectric response of black lipid membranes incorporating an amphiphilic azobenzene derivative. Thin Solid Films 210/211, 736–738 (1992).

    Google Scholar 

  8. 8.

    Tanaka, M. & Yonezawa, Y. Photochemical regulation of ion transport through “quasi-channels” embedded in black lipid membrane. Mat. Sci. Eng. C. 4, 297–301 (1997).

    Google Scholar 

  9. 9.

    Garner, L. E. et al. Modification of the optoelectronic properties of membranes via insertion of amphiphilic phenylenevinylene oligoelectrolytes. JACS 132, 10042–10052 (2010).

    CAS  Google Scholar 

  10. 10.

    Hinks, J. et al. Modeling cell membrane perturbation by molecules designed for transmembrane electron transfer. Langmuir 30, 2429–2440 (2014).

    CAS  Google Scholar 

  11. 11.

    Gorostiza, P. & Isacoff, E. Optical switches and triggers for the manipulation of ion channels and pores. Mol. Biosyst. 3, 686–704 (2007).

    CAS  Google Scholar 

  12. 12.

    Fortin, D. L. et al. Photochemical control of endogenous ion channels and cellular excitability. Nat. Methods 5, 331–338 (2008).

    CAS  Google Scholar 

  13. 13.

    Kramer, R. H., Mourot, A. & Adesnik, H. Optogenetic pharmacology for control of native neuronal signaling proteins. Nat. Neurosci. 16, 816–823 (2013).

    Google Scholar 

  14. 14.

    Tochitsky, I., Kienzler, M. A., Isacoff, E. & Kramer, R. H. Restoring vision to the blind with chemical photoswitches. Chem. Rev. 118, 10748–10773 (2018).

    CAS  Google Scholar 

  15. 15.

    Laprell, L. et al. Restoring light sensitivity in blind retinae using a photochromic AMPA receptor agonist. ACS Chem. Neurosci. 7, 15–20 (2016).

    CAS  Google Scholar 

  16. 16.

    Laprell, L. et al. Photopharmacological control of bipolar cells restores visual function in blind mice. J. Clin. Invest. 127, 2598–2611 (2017).

    Google Scholar 

  17. 17.

    Ghezzi, D. et al. A hybrid bioorganic interface for neuronal photoactivation. Nat. Commun. 2, 166 (2011).

    Google Scholar 

  18. 18.

    Ghezzi, D. et al. A polymer optoelectronic interface restores light sensitivity in blind rat retinas. Nat. Photonics 7, 400–406 (2013).

    CAS  Google Scholar 

  19. 19.

    Feyen, P. et al. Light-evoked hyperpolarization and silencing of neurons by conjugated polymers. Sci. Rep. 6, 22718 (2016).

    CAS  Google Scholar 

  20. 20.

    Rand, D. et al. Direct electrical neurostimulation with organic pigment photocapacitors. Adv. Mater. 30, e1707292 (2018).

    Google Scholar 

  21. 21.

    Maya-Vetencourt, J. F. et al. A fully organic retinal prosthesis restores vision in a rat model of degenerative blindness. Nat. Mater. 16, 681–689 (2017).

    CAS  Google Scholar 

  22. 22.

    Martino, N. et al. Photothermal cellular stimulation in functional bio-polymer interfaces. Sci. Rep. 5, 8911 (2015).

    CAS  Google Scholar 

  23. 23.

    Lodola, F., Martino, N., Tullii, G., Lanzani, G. & Antognazza, M. R. Conjugated polymers mediate effective activation of the mammalian ion channel transient receptor potential vanilloid 1. Sci. Rep. 7, 8477 (2017).

    CAS  Google Scholar 

  24. 24.

    Shapiro, M. G., Homma, K., Villarreal, S., Richter, C. P. & Bezanilla, F. Infrared light excites cells by changing their electrical capacitance. Nat. Commun. 3, 736 (2012).

    Google Scholar 

  25. 25.

    Carvalho-de-Souza, J. L. et al. Photosensitivity of neurons enabled by cell-targeted gold nanoparticles. Neuron 86, 207–217 (2015).

    CAS  Google Scholar 

  26. 26.

    Bandara, H. M. & Burdette, S. C. Photoisomerization in different classes of azobenzene. Chem. Soc. Rev. 41, 1809–1825 (2012).

    CAS  Google Scholar 

  27. 27.

    Hartley, G. S. The Cis-form of azobenzene. Nature 140, 281–281 (1937).

    CAS  Google Scholar 

  28. 28.

    Rau, H. Spectroscopic properties of organic azo compounds. Angew. Chem. 12, 224–235 (1973).

    Google Scholar 

  29. 29.

    Tang B. Z. & Qin A. Aggregation-Induced Emission: Fundamentals (Wiley, 2013).

  30. 30.

    Sierocki, P. et al. Photoisomerization of azobenzene derivatives in nanostructured silica. J. Phys. Chem. B 110, 24390–24398 (2006).

    CAS  Google Scholar 

  31. 31.

    Fendler, J. H. Surfactant vesicles as membrane mimetic agents: characterization and utilization. Acc. Chem. Res. 13, 7–13 (1980).

    CAS  Google Scholar 

  32. 32.

    Head, B. P., Patel, H. H. & Insel, P. A. Interaction of membrane/lipid rafts with the cytoskeleton: impact on signaling and function: membrane/lipid rafts, mediators of cytoskeletal arrangement and cell signaling. Biochim. Biophys. Acta 1838, 532–545 (2014).

    CAS  Google Scholar 

  33. 33.

    Fruscione, F. et al. PRRT2 controls neuronal excitability by negatively modulating Na+ channel 1.2/1.6 activity. Brain 141, 1000–1016 (2018).

    Google Scholar 

  34. 34.

    Thalhammer, A. et al. Alternative splicing of P/Q-type Ca2+ channels shapes presynaptic plasticity. Cell Rep. 20, 333–343 (2017).

    CAS  Google Scholar 

  35. 35.

    Zhang, Q. & Bazuin, C. G. Liquid crystallinity and other properties in complexes of cationic azo-containing surfactomesogens with poly(styrenesulfonate). Macromolecules 42, 4775–4786 (2009).

    CAS  Google Scholar 

  36. 36.

    Peddie, V., Anderson, J., Harvey, J. E., Smith, G. J. & Kay, A. Synthesis and solution aggregation studies of a suite of mixed neutral and zwitterionic chromophores for second-order nonlinear optics. J. Org. Chem. 79, 10153–10169 (2014).

    CAS  Google Scholar 

  37. 37.

    Allen, J. A., Halverson-Tamboli, R. A. & Rasenick, M. M. Lipid raft microdomains and neurotransmitter signalling. Nat. Rev. Neurosci. 8, 128–140 (2007).

    CAS  Google Scholar 

  38. 38.

    Frank, J. A., Franquelim, H. G., Schwille, P. & Trauner, D. Optical control of lipid rafts with photoswitchable ceramides. J. Am. Chem. Soc. 138, 12981–12986 (2016).

    CAS  Google Scholar 

  39. 39.

    Hodgkin, Al & Huxley, A. F. A quantitative description of membrane current and its application to conduction and excitation in nerve. J. Physiol. 117, 500–544 (1952).

    CAS  Google Scholar 

  40. 40.

    Tremere, L. A., Pinaud, R., Irwin, R. P. & Allen, C. N. Postinhibitory rebound spikes are modulated by the history of membrane hyperpolarization in the SCN. Eur. J. Neurosci. 28, 1127–1135 (2008).

    Google Scholar 

  41. 41.

    Jo, S., Kim, T., Iyer, V. G. & Im, W. CHARMM-GUI: a Web-based graphical user interface for CHARMM. J. Comput. Chem. 29, 1859–1865 (2008).

    CAS  Google Scholar 

  42. 42.

    Phillips, J. C. et al. Scalable molecular dynamics with NAMD +. J. Comput. Chem. 26, 1781–1802 (2005).

    CAS  Google Scholar 

  43. 43.

    Huang, J. & MacKerell, A. D. Jr. CHARMM36 all-atom additive protein force field: validation based on comparison to NMR data. J. Comput. Chem. 34, 2135–2145 (2013).

    CAS  Google Scholar 

  44. 44.

    Bennett, W. F. & Tieleman, D. P. Molecular simulation of rapid translocation of cholesterol, diacylglycerol, and ceramide in model raft and nonraft membranes. J. Lipid Res. 53, 421–429 (2012).

    CAS  Google Scholar 

  45. 45.

    Maragliano, L. & Vanden-Eijnden, E. A temperature accelerated method for sampling free energy and determining reaction pathways in rare events simulations. Chem. Phys. Lett. 426, 168–175 (2006).

    CAS  Google Scholar 

  46. 46.

    Dalla Serra, M. & Menestrina, G. Liposomes in study of pore-forming toxins. Meth Enzymol. 372, 99–124 (2003).

    CAS  Google Scholar 

  47. 47.

    Dalla Serra, M. & Menestrina, G. Characterization of molecular properties of pore-forming toxins with planar lipid bilayers. Meth Mol. Biol. 145, 171–188 (2000).

    CAS  Google Scholar 

  48. 48.

    Schmitt, B. M. & Koepsell, H. An improved method for real-time monitoring of membrane capacitance in Xenopus laevis oocytes. Biophys. J. 82, 1345–1357 (2002).

    CAS  Google Scholar 

  49. 49.

    Pusch, M. & Neher, E. Rates of diffusional exchange between small cells and a measuring patch pipette. Pflug. Arch. 411, 204–211 (1988).

    CAS  Google Scholar 

  50. 50.

    Gillis, K. D. Admittance-based measurement of membrane capacitance using the EPC-9 patch-clamp amplifier. Pflug. Arch. 439, 655–664 (2000).

    CAS  Google Scholar 

Download references


We thank F. Fruscione and F. Zara (Giannina Gaslini Institute, Genova, Italy) for help in preparing iPSC-derived human neurons; P. Bianchini, M. Oneto and M. Scotto (Center for Nanoscopy and Nikon Imaging Center, Istituto Italiano di Tecnologia, Genova, Italy); L. Cingolani (Center for Synaptic Neuroscience and Technology, Istituto Italiano di Tecnologia, Genova, Italy) for providing the ChETA-encoding lentiviral vectors; A. Mehilli and G. Mantero (Center for Synaptic Neuroscience and Technology, Istituto Italiano di Tecnologia, Genova, Italy) for precious help in primary cultures and in vivo elelectrophysiology, respectively. This work was supported by the Italian Ministry of Health (project RF-2013-02358313 to G.P., G.L. and F.B.) and Istituto Italiano di Tecnologia (pre-startup project to G.L. and F.B.). The support of the Ra.Mo. Foundation (Milano, Italy), Fondazione 13 Marzo (Parma, Italy), Rare Partners srl (Milano, Italy) and Fondazione Cariplo (project 2018-0505) to G.L. and F.B. is also acknowledged.

Author information




C.B. designed and engineered Ziapin2. S.C., L.C. and F.O. performed the synthesis and characterization of Ziapin2. D.F. calculated the atomic charges and optimized coordinates. G.M.P. performed the spectroscopic characterization. L.M. performed molecular dynamics simulations. M.D.S., L.L. and M.M. performed planar lipid membrane and AFM studies. M.B., G.G. and E.C. studied the in vitro and in vivo distribution of the Ziapin compounds in neurons. M.L.D.F., P.B., E.C. and F.L. performed the in vitro patch-clamp experiments and analysed the data. V.V. elaborated the numerical RC model. J.F.M-V., E.C., D.S. and C.G.E. performed and analysed the in vivo experiments. M.L.D.F., E.C., P.B., G.M.P. and F.L. contributed to paper writing. G.L., C.B. and F.B. conceived the work. G.L. and F.B. planned the experiments, analysed the data and wrote the manuscript.

Corresponding authors

Correspondence to Guglielmo Lanzani or Fabio Benfenati.

Ethics declarations

Competing interests

The authors declare no competing interests.

Additional information

Peer review information Nature Nanotechnology thanks Or Shemesh, Joao L. Carvalho-de-Souza and the other, anonymous, reviewer(s) for their contribution to the peer review of this work.

Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Supplementary information

Supplementary Information

Supplementary information and Figs. 1–18.

Reporting Summary

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

DiFrancesco, M.L., Lodola, F., Colombo, E. et al. Neuronal firing modulation by a membrane-targeted photoswitch. Nat. Nanotechnol. 15, 296–306 (2020).

Download citation

Further reading


Quick links

Find nanotechnology articles, nanomaterial data and patents all in one place. Visit Nano by Nature Research