Optogenetic therapy restores retinal activity in primate for at least a year following photoreceptor ablation

Juliette E McGregor; Karteek Kunala; Zhengyang Xu; Peter J Murphy; Tyler Godat; Jennifer M Strazzeri; Brittany A Bateman; William S Fischer; Keith Parkins; Colin J Chu; Teresa Puthussery; David R Williams; William H Merigan

doi:10.1016/j.ymthe.2021.09.014

Optogenetic therapy restores retinal activity in primate for at least a year following photoreceptor ablation

Juliette E McGregor
, Karteek Kunala
, Zhengyang Xu
, Peter J Murphy
, Tyler Godat
, Jennifer M Strazzeri
, Brittany A Bateman
, William S Fischer
, Keith Parkins
, Colin J Chu
, Teresa Puthussery
, David R Williams
, William H Merigan

Research output: Contribution to journal › Article (Academic Journal) › peer-review

18 Citations (Scopus)

117 Downloads (Pure)

Abstract

All retina-based vision restoration approaches rely on the assumption that photoreceptor loss does not preclude reactivation of the remaining retinal architecture. Whether extended periods of vision loss limit the efficacy of restorative therapies at the retinal level is unknown. We examined long-term changes in optogenetic responsivity of foveal retinal ganglion cells (RGCs) in non-human primates following localized photoreceptor ablation by high intensity laser exposure. By performing fluorescence adaptive optics scanning light ophthalmoscopy (AOSLO) of RGCs expressing both the calcium indicator GCaMP6s and the optogenetic actuator ChrimsonR, it was possible to track optogenetic mediated calcium responses in deafferented RGCs over time. Fluorescence fundus photography revealed a 40% reduction in ChrimsonR fluorescence in RGCs lacking photoreceptor input over the three weeks following photoreceptor ablation. Despite this, in vivo imaging revealed good cellular preservation of RGCs 3 months after the loss of photoreceptor input, and histology confirmed good structural preservation at 2 years. Optogenetic responses of RGCs in primate persisted for at least one year after the loss of photoreceptor input, with a sensitivity index similar to optogenetic responses recorded in intact retina. These results are promising for all potential therapeutic approaches to vision restoration that rely on preservation and reactivation of RGCs.

Original language	English
Journal	Molecular Therapy
Early online date	20 Sept 2021
DOIs	https://doi.org/10.1016/j.ymthe.2021.09.014
Publication status	E-pub ahead of print - 20 Sept 2021

Bibliographical note

Funding Information:
The authors wish to thank Amber Walker for assistance with animal care and anesthesia, Thurma McDaniel for tissue preparation for immuno-histochemistry, and Qiang Yang, who developed image acquisition, stabilization, and registration software. We thank Kamal Dhakal for providing training and Jennifer Hunter and Khang Huynh for accommodating access to the 2P AOSLO. We thank Michael Giocomelli, Krystel Huxlin, and Louis DiVincenti for helpful discussions and Ron Plotnik and David DiLoreto, Jr. for providing clinical advice. We thank Erin Tepesch and Ying Kai Chan for assistance with characterizing inflammation. We thank the vector core at the Perelman School of Medicine, University of Pennsylvania and the Genetically-Encoded Neuronal Indicator and Effector (GENIE) Project and the Janelia Research Campus of the Howard Hughes Medical Institute, specifically Vivek Jayaraman, PhD, Douglas S. Kim, PhD, Loren L. Looger, PhD, and Karel Svoboda, PhD. This work was supported by the National Eye Institute ( https://nei.nih.gov/about ) through grants NIH EY021166 (to W.H.M), NIH EY025497 (to D.R.W.), and NIH EY024265 (to T.P.) and P30 Core Grants NIH EY001319 (University of Rochester) and NIH EY003176 (UC Berkeley) and through an unrestricted grant to the Flaum Eye Institute from Research to Prevent Blindness ( https://www.rpbusa.org/rpb/grants-and-research/grants/rpb-unrestricted-grants/rpb-unrestricted-grants/ ). Confocal imaging was conducted in at the CRL Molecular Imaging Center at UC Berkeley (RRID: SCR_017852 ), supported by the Helen Wills Neuroscience Institute. We thank Holly Aaron and Feather Ives for microscopy assistance

Funding Information:
The authors wish to thank Amber Walker for assistance with animal care and anesthesia, Thurma McDaniel for tissue preparation for immuno-histochemistry, and Qiang Yang, who developed image acquisition, stabilization, and registration software. We thank Kamal Dhakal for providing training and Jennifer Hunter and Khang Huynh for accommodating access to the 2P AOSLO. We thank Michael Giocomelli, Krystel Huxlin, and Louis DiVincenti for helpful discussions and Ron Plotnik and David DiLoreto, Jr. for providing clinical advice. We thank Erin Tepesch and Ying Kai Chan for assistance with characterizing inflammation. We thank the vector core at the Perelman School of Medicine, University of Pennsylvania and the Genetically-Encoded Neuronal Indicator and Effector (GENIE) Project and the Janelia Research Campus of the Howard Hughes Medical Institute, specifically Vivek Jayaraman, PhD, Douglas S. Kim, PhD, Loren L. Looger, PhD, and Karel Svoboda, PhD. This work was supported by the National Eye Institute (https://nei.nih.gov/about) through grants NIH EY021166 (to W.H.M), NIH EY025497 (to D.R.W.), and NIH EY024265 (to T.P.) and P30 Core Grants NIH EY001319 (University of Rochester) and NIH EY003176 (UC Berkeley) and through an unrestricted grant to the Flaum Eye Institute from Research to Prevent Blindness (https://www.rpbusa.org/rpb/grants-and-research/grants/rpb-unrestricted-grants/rpb-unrestricted-grants/). Confocal imaging was conducted in at the CRL Molecular Imaging Center at UC Berkeley (RRID:SCR_017852), supported by the Helen Wills Neuroscience Institute. We thank Holly Aaron and Feather Ives for microscopy assistance, J.E.M. designed and performed the experiments, developed and performed data analysis, interpreted results, managed the project, presented data, and wrote the manuscript. K.K. performed ultrafast laser exposures. Z.X. assisted with data analysis. P.J.M and T.G. assisted with data collection and analysis. K.P developed analysis routines. J.M.S. provided animal support. B.A.B. performed fluorescence fundus, OCT, and cSLO imaging. W.S.F. customized the fundus camera and performed OCT and cSLO. C.J.C. assessed intraocular inflammation. T.P. designed and performed immuno histochemistry and microscopy, interpreted results, and revised the manuscript. D.R.W. secured funding, interpreted results, and revised the manuscript. W.H.M. conceived the study, secured funding, interpreted results, performed intravitreal injections, revised the manuscript, and supervised the project. D.R.W. has received income in the form of royalties in excess of $10,000 from licenses on patents upon which he is an inventor, held by the University of Rochester. These patents have been awarded for wavefront sensing and adaptive optics technology, both of which were used to carry out the research described in this paper. He has also received research support in excess of $50,000 in the prior fiscal year from a company in a health care field. The nature of this research is completely unrelated to the subject matter or methods of the present publication. All other authors declare no competing interests.

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© 2021 The Authors

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McGregor, J. E., Kunala, K., Xu, Z., Murphy, P. J., Godat, T., Strazzeri, J. M., Bateman, B. A., Fischer, W. S., Parkins, K., Chu, C. J., Puthussery, T., Williams, D. R., & Merigan, W. H. (2021). Optogenetic therapy restores retinal activity in primate for at least a year following photoreceptor ablation. Molecular Therapy. Advance online publication. https://doi.org/10.1016/j.ymthe.2021.09.014

@article{b7f83287fd6444d7958a71f54e02142c,

title = "Optogenetic therapy restores retinal activity in primate for at least a year following photoreceptor ablation",

abstract = "All retina-based vision restoration approaches rely on the assumption that photoreceptor loss does not preclude reactivation of the remaining retinal architecture. Whether extended periods of vision loss limit the efficacy of restorative therapies at the retinal level is unknown. We examined long-term changes in optogenetic responsivity of foveal retinal ganglion cells (RGCs) in non-human primates following localized photoreceptor ablation by high intensity laser exposure. By performing fluorescence adaptive optics scanning light ophthalmoscopy (AOSLO) of RGCs expressing both the calcium indicator GCaMP6s and the optogenetic actuator ChrimsonR, it was possible to track optogenetic mediated calcium responses in deafferented RGCs over time. Fluorescence fundus photography revealed a 40\% reduction in ChrimsonR fluorescence in RGCs lacking photoreceptor input over the three weeks following photoreceptor ablation. Despite this, in vivo imaging revealed good cellular preservation of RGCs 3 months after the loss of photoreceptor input, and histology confirmed good structural preservation at 2 years. Optogenetic responses of RGCs in primate persisted for at least one year after the loss of photoreceptor input, with a sensitivity index similar to optogenetic responses recorded in intact retina. These results are promising for all potential therapeutic approaches to vision restoration that rely on preservation and reactivation of RGCs.",

author = "McGregor, \{Juliette E\} and Karteek Kunala and Zhengyang Xu and Murphy, \{Peter J\} and Tyler Godat and Strazzeri, \{Jennifer M\} and Bateman, \{Brittany A\} and Fischer, \{William S\} and Keith Parkins and Chu, \{Colin J\} and Teresa Puthussery and Williams, \{David R\} and Merigan, \{William H\}",

note = "Funding Information: The authors wish to thank Amber Walker for assistance with animal care and anesthesia, Thurma McDaniel for tissue preparation for immuno-histochemistry, and Qiang Yang, who developed image acquisition, stabilization, and registration software. We thank Kamal Dhakal for providing training and Jennifer Hunter and Khang Huynh for accommodating access to the 2P AOSLO. We thank Michael Giocomelli, Krystel Huxlin, and Louis DiVincenti for helpful discussions and Ron Plotnik and David DiLoreto, Jr. for providing clinical advice. We thank Erin Tepesch and Ying Kai Chan for assistance with characterizing inflammation. We thank the vector core at the Perelman School of Medicine, University of Pennsylvania and the Genetically-Encoded Neuronal Indicator and Effector (GENIE) Project and the Janelia Research Campus of the Howard Hughes Medical Institute, specifically Vivek Jayaraman, PhD, Douglas S. Kim, PhD, Loren L. Looger, PhD, and Karel Svoboda, PhD. This work was supported by the National Eye Institute ( https://nei.nih.gov/about ) through grants NIH EY021166 (to W.H.M), NIH EY025497 (to D.R.W.), and NIH EY024265 (to T.P.) and P30 Core Grants NIH EY001319 (University of Rochester) and NIH EY003176 (UC Berkeley) and through an unrestricted grant to the Flaum Eye Institute from Research to Prevent Blindness ( https://www.rpbusa.org/rpb/grants-and-research/grants/rpb-unrestricted-grants/rpb-unrestricted-grants/ ). Confocal imaging was conducted in at the CRL Molecular Imaging Center at UC Berkeley (RRID: SCR\_017852 ), supported by the Helen Wills Neuroscience Institute. We thank Holly Aaron and Feather Ives for microscopy assistance Funding Information: The authors wish to thank Amber Walker for assistance with animal care and anesthesia, Thurma McDaniel for tissue preparation for immuno-histochemistry, and Qiang Yang, who developed image acquisition, stabilization, and registration software. We thank Kamal Dhakal for providing training and Jennifer Hunter and Khang Huynh for accommodating access to the 2P AOSLO. We thank Michael Giocomelli, Krystel Huxlin, and Louis DiVincenti for helpful discussions and Ron Plotnik and David DiLoreto, Jr. for providing clinical advice. We thank Erin Tepesch and Ying Kai Chan for assistance with characterizing inflammation. We thank the vector core at the Perelman School of Medicine, University of Pennsylvania and the Genetically-Encoded Neuronal Indicator and Effector (GENIE) Project and the Janelia Research Campus of the Howard Hughes Medical Institute, specifically Vivek Jayaraman, PhD, Douglas S. Kim, PhD, Loren L. Looger, PhD, and Karel Svoboda, PhD. This work was supported by the National Eye Institute (https://nei.nih.gov/about) through grants NIH EY021166 (to W.H.M), NIH EY025497 (to D.R.W.), and NIH EY024265 (to T.P.) and P30 Core Grants NIH EY001319 (University of Rochester) and NIH EY003176 (UC Berkeley) and through an unrestricted grant to the Flaum Eye Institute from Research to Prevent Blindness (https://www.rpbusa.org/rpb/grants-and-research/grants/rpb-unrestricted-grants/rpb-unrestricted-grants/). Confocal imaging was conducted in at the CRL Molecular Imaging Center at UC Berkeley (RRID:SCR\_017852), supported by the Helen Wills Neuroscience Institute. We thank Holly Aaron and Feather Ives for microscopy assistance, J.E.M. designed and performed the experiments, developed and performed data analysis, interpreted results, managed the project, presented data, and wrote the manuscript. K.K. performed ultrafast laser exposures. Z.X. assisted with data analysis. P.J.M and T.G. assisted with data collection and analysis. K.P developed analysis routines. J.M.S. provided animal support. B.A.B. performed fluorescence fundus, OCT, and cSLO imaging. W.S.F. customized the fundus camera and performed OCT and cSLO. C.J.C. assessed intraocular inflammation. T.P. designed and performed immuno histochemistry and microscopy, interpreted results, and revised the manuscript. D.R.W. secured funding, interpreted results, and revised the manuscript. W.H.M. conceived the study, secured funding, interpreted results, performed intravitreal injections, revised the manuscript, and supervised the project. D.R.W. has received income in the form of royalties in excess of \$10,000 from licenses on patents upon which he is an inventor, held by the University of Rochester. These patents have been awarded for wavefront sensing and adaptive optics technology, both of which were used to carry out the research described in this paper. He has also received research support in excess of \$50,000 in the prior fiscal year from a company in a health care field. The nature of this research is completely unrelated to the subject matter or methods of the present publication. All other authors declare no competing interests. Publisher Copyright: {\textcopyright} 2021 The Authors",

year = "2021",

month = sep,

day = "20",

doi = "10.1016/j.ymthe.2021.09.014",

language = "English",

journal = "Molecular Therapy",

issn = "1525-0016",

publisher = "Cell Press",

}

TY - JOUR

T1 - Optogenetic therapy restores retinal activity in primate for at least a year following photoreceptor ablation

AU - McGregor, Juliette E

AU - Kunala, Karteek

AU - Xu, Zhengyang

AU - Murphy, Peter J

AU - Godat, Tyler

AU - Strazzeri, Jennifer M

AU - Bateman, Brittany A

AU - Fischer, William S

AU - Parkins, Keith

AU - Chu, Colin J

AU - Puthussery, Teresa

AU - Williams, David R

AU - Merigan, William H

N1 - Funding Information: The authors wish to thank Amber Walker for assistance with animal care and anesthesia, Thurma McDaniel for tissue preparation for immuno-histochemistry, and Qiang Yang, who developed image acquisition, stabilization, and registration software. We thank Kamal Dhakal for providing training and Jennifer Hunter and Khang Huynh for accommodating access to the 2P AOSLO. We thank Michael Giocomelli, Krystel Huxlin, and Louis DiVincenti for helpful discussions and Ron Plotnik and David DiLoreto, Jr. for providing clinical advice. We thank Erin Tepesch and Ying Kai Chan for assistance with characterizing inflammation. We thank the vector core at the Perelman School of Medicine, University of Pennsylvania and the Genetically-Encoded Neuronal Indicator and Effector (GENIE) Project and the Janelia Research Campus of the Howard Hughes Medical Institute, specifically Vivek Jayaraman, PhD, Douglas S. Kim, PhD, Loren L. Looger, PhD, and Karel Svoboda, PhD. This work was supported by the National Eye Institute ( https://nei.nih.gov/about ) through grants NIH EY021166 (to W.H.M), NIH EY025497 (to D.R.W.), and NIH EY024265 (to T.P.) and P30 Core Grants NIH EY001319 (University of Rochester) and NIH EY003176 (UC Berkeley) and through an unrestricted grant to the Flaum Eye Institute from Research to Prevent Blindness ( https://www.rpbusa.org/rpb/grants-and-research/grants/rpb-unrestricted-grants/rpb-unrestricted-grants/ ). Confocal imaging was conducted in at the CRL Molecular Imaging Center at UC Berkeley (RRID: SCR_017852 ), supported by the Helen Wills Neuroscience Institute. We thank Holly Aaron and Feather Ives for microscopy assistance Funding Information: The authors wish to thank Amber Walker for assistance with animal care and anesthesia, Thurma McDaniel for tissue preparation for immuno-histochemistry, and Qiang Yang, who developed image acquisition, stabilization, and registration software. We thank Kamal Dhakal for providing training and Jennifer Hunter and Khang Huynh for accommodating access to the 2P AOSLO. We thank Michael Giocomelli, Krystel Huxlin, and Louis DiVincenti for helpful discussions and Ron Plotnik and David DiLoreto, Jr. for providing clinical advice. We thank Erin Tepesch and Ying Kai Chan for assistance with characterizing inflammation. We thank the vector core at the Perelman School of Medicine, University of Pennsylvania and the Genetically-Encoded Neuronal Indicator and Effector (GENIE) Project and the Janelia Research Campus of the Howard Hughes Medical Institute, specifically Vivek Jayaraman, PhD, Douglas S. Kim, PhD, Loren L. Looger, PhD, and Karel Svoboda, PhD. This work was supported by the National Eye Institute (https://nei.nih.gov/about) through grants NIH EY021166 (to W.H.M), NIH EY025497 (to D.R.W.), and NIH EY024265 (to T.P.) and P30 Core Grants NIH EY001319 (University of Rochester) and NIH EY003176 (UC Berkeley) and through an unrestricted grant to the Flaum Eye Institute from Research to Prevent Blindness (https://www.rpbusa.org/rpb/grants-and-research/grants/rpb-unrestricted-grants/rpb-unrestricted-grants/). Confocal imaging was conducted in at the CRL Molecular Imaging Center at UC Berkeley (RRID:SCR_017852), supported by the Helen Wills Neuroscience Institute. We thank Holly Aaron and Feather Ives for microscopy assistance, J.E.M. designed and performed the experiments, developed and performed data analysis, interpreted results, managed the project, presented data, and wrote the manuscript. K.K. performed ultrafast laser exposures. Z.X. assisted with data analysis. P.J.M and T.G. assisted with data collection and analysis. K.P developed analysis routines. J.M.S. provided animal support. B.A.B. performed fluorescence fundus, OCT, and cSLO imaging. W.S.F. customized the fundus camera and performed OCT and cSLO. C.J.C. assessed intraocular inflammation. T.P. designed and performed immuno histochemistry and microscopy, interpreted results, and revised the manuscript. D.R.W. secured funding, interpreted results, and revised the manuscript. W.H.M. conceived the study, secured funding, interpreted results, performed intravitreal injections, revised the manuscript, and supervised the project. D.R.W. has received income in the form of royalties in excess of $10,000 from licenses on patents upon which he is an inventor, held by the University of Rochester. These patents have been awarded for wavefront sensing and adaptive optics technology, both of which were used to carry out the research described in this paper. He has also received research support in excess of $50,000 in the prior fiscal year from a company in a health care field. The nature of this research is completely unrelated to the subject matter or methods of the present publication. All other authors declare no competing interests. Publisher Copyright: © 2021 The Authors

PY - 2021/9/20

Y1 - 2021/9/20

N2 - All retina-based vision restoration approaches rely on the assumption that photoreceptor loss does not preclude reactivation of the remaining retinal architecture. Whether extended periods of vision loss limit the efficacy of restorative therapies at the retinal level is unknown. We examined long-term changes in optogenetic responsivity of foveal retinal ganglion cells (RGCs) in non-human primates following localized photoreceptor ablation by high intensity laser exposure. By performing fluorescence adaptive optics scanning light ophthalmoscopy (AOSLO) of RGCs expressing both the calcium indicator GCaMP6s and the optogenetic actuator ChrimsonR, it was possible to track optogenetic mediated calcium responses in deafferented RGCs over time. Fluorescence fundus photography revealed a 40% reduction in ChrimsonR fluorescence in RGCs lacking photoreceptor input over the three weeks following photoreceptor ablation. Despite this, in vivo imaging revealed good cellular preservation of RGCs 3 months after the loss of photoreceptor input, and histology confirmed good structural preservation at 2 years. Optogenetic responses of RGCs in primate persisted for at least one year after the loss of photoreceptor input, with a sensitivity index similar to optogenetic responses recorded in intact retina. These results are promising for all potential therapeutic approaches to vision restoration that rely on preservation and reactivation of RGCs.

AB - All retina-based vision restoration approaches rely on the assumption that photoreceptor loss does not preclude reactivation of the remaining retinal architecture. Whether extended periods of vision loss limit the efficacy of restorative therapies at the retinal level is unknown. We examined long-term changes in optogenetic responsivity of foveal retinal ganglion cells (RGCs) in non-human primates following localized photoreceptor ablation by high intensity laser exposure. By performing fluorescence adaptive optics scanning light ophthalmoscopy (AOSLO) of RGCs expressing both the calcium indicator GCaMP6s and the optogenetic actuator ChrimsonR, it was possible to track optogenetic mediated calcium responses in deafferented RGCs over time. Fluorescence fundus photography revealed a 40% reduction in ChrimsonR fluorescence in RGCs lacking photoreceptor input over the three weeks following photoreceptor ablation. Despite this, in vivo imaging revealed good cellular preservation of RGCs 3 months after the loss of photoreceptor input, and histology confirmed good structural preservation at 2 years. Optogenetic responses of RGCs in primate persisted for at least one year after the loss of photoreceptor input, with a sensitivity index similar to optogenetic responses recorded in intact retina. These results are promising for all potential therapeutic approaches to vision restoration that rely on preservation and reactivation of RGCs.

U2 - 10.1016/j.ymthe.2021.09.014

DO - 10.1016/j.ymthe.2021.09.014

M3 - Article (Academic Journal)

C2 - 34547460

SN - 1525-0016

JO - Molecular Therapy

JF - Molecular Therapy

ER -

Optogenetic therapy restores retinal activity in primate for at least a year following photoreceptor ablation

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