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Neuronal firing modulation by a membrane-targeted photoswitch

Nature Nanotechnology volume 15, pages 296–306 (2020)Cite this article

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Abstract

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.

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Fig. 1: Trans→cis 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.

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Data availability

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

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Acknowledgements

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.

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Author notes

  1. Mattia Bramini

    Present address: Mattia Bramini, Department of Applied Physics, Faculty of Sciences, University of Granada, Granada, Spain

  2. Cyril Giles Eleftheriou

    Present address: Cyril Giles Eleftheriou, Departments of Ophtalmology and Neurology, Burke Medical Research Institute, Weil Medical College of Cornell University, White Plains, NY, USA

  3. José Fernando Maya-Vetencourt

    Present address: José Fernando Maya-Vetencourt, Department of Biology, University of Pisa, Pisa, Italy

  4. These authors contributed equally: Mattia Lorenzo DiFrancesco, Francesco Lodola, Elisabetta Colombo, Chiara Bertarelli, Guglielmo Lanzani, Fabio Benfenati.

Authors and Affiliations

  1. Center for Synaptic Neuroscience, Istituto Italiano di Tecnologia, Genoa, Italy

    Mattia Lorenzo DiFrancesco, Elisabetta Colombo, Luca Maragliano, Mattia Bramini, Giorgio Grasselli, Cyril Giles Eleftheriou, Dmytro Shmal, José Fernando Maya-Vetencourt & Fabio Benfenati

  2. IRCCS Ospedale Policlinico San Martino, Genoa, Italy

    Mattia Lorenzo DiFrancesco, Elisabetta Colombo, Luca Maragliano, Mattia Bramini, Pietro Baldelli, Giorgio Grasselli, Cyril Giles Eleftheriou, Dmytro Shmal, José Fernando Maya-Vetencourt & Fabio Benfenati

  3. Center for Nano Science and Technology, Istituto Italiano di Tecnologia, Milan, Italy

    Francesco Lodola, Giuseppe Maria Paternò, Simone Cimò, Vito Vurro, Chiara Bertarelli & Guglielmo Lanzani

  4. Department of Experimental Medicine, University of Genova, Genoa, Italy

    Pietro Baldelli

  5. Istituto di Biofisica, Consiglio Nazionale delle Ricerche, Trento, Italy

    Mauro Dalla Serra, Lorenzo Lunelli & Marta Marchioretto

  6. Laboratory of Biomarker Studies and Structure Analysis for Health, Fondazione Bruno Kessler, Trento, Italy

    Mauro Dalla Serra, Lorenzo Lunelli & Marta Marchioretto

  7. Dipartimento di Chimica, Materiali e Ingegneria Chimica ‘Giulio Natta’, Politecnico di Milano, Milan, Italy

    Simone Cimò, Letizia Colella & Chiara Bertarelli

  8. Department of Chemistry, Institut für Physikalische Chemie, University of Cologne, Cologne, Germany

    Daniele Fazzi

  9. Department of Chemistry, Biology and Biotechnology, Università degli Studi di Perugia, Perugia, Italy

    Fausto Ortica

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  1. Mattia Lorenzo DiFrancesco
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Contributions

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.

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Correspondence to Guglielmo Lanzani or Fabio Benfenati.

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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.

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DiFrancesco, M.L., Lodola, F., Colombo, E. et al. Neuronal firing modulation by a membrane-targeted photoswitch. Nat. Nanotechnol. 15, 296–306 (2020). https://doi.org/10.1038/s41565-019-0632-6

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