Guillaume Chenegros

Enhancing retinal images by nonlinear registration

Being able to image the human retina in high resolution opens a new era in many important fields, such as pharmacological research for retinal diseases, researches in human cognition, nervous system, metabolism and blood stream, to name a few. In this paper, we propose to share the knowledge acquired in the fields of optics and imaging in solar astrophysics in order to improve the retinal imaging in the perspective to perform a medical diagnosis. The main purpose would be to assist health care practitioners by enhancing the spatial resolution of the retinal images and increase the level of confidence of the abnormal feature detection. We apply a nonlinear registration method using local correlation tracking to increase the field of view and follow structure evolutions using correlation techniques borrowed from solar astronomy technique expertise. Another purpose is to define the tracer of movements after analyzing local correlations to follow the proper motions of an image from one moment to another, such as changes in optical flows that would be of high interest in a medical diagnosis.

A stimulation platform for optogenetic and bionic vision restoration

Optogenetic therapy holds the promise to restore visual function in patients affected by retinal degenerative diseases. However, the light-sensitivity of the molecule mediating light responses is much less than the one of healthy retinal cells so that no photo-stimulation is expected under natural environmental conditions. In this work, we present a platform set up to stimulate optogenetically-engineered retinal cells, and the algorithms associated with different types of stimulation. The system consists of a neuromorphic silicon retina as a visual frontend, a projecting device capable of delivering fast and precise light stimulation and a computing platform implementing the stimulation algorithms. We describe different strategies, varying depending on the type of cells transfected. The silicon retina provides a natural front-end for an artificial visual system, complying with the information encoding principles, timing properties and dynamic range of either photoreceptors or RGCs. The encoding of the visual information is performed with sub-millisecond accuracy, respecting the temporal characteristics of the neural system. The platform and algorithms hereby presented provide a basis for medical devices matching the requirements of optogenetic therapeutic use. An embedded version of this platform will be used in the forthcoming clinical trials of the GS030 vision restoration therapy.

Full-field OCT technique for high speed event-based optical flow and particle tracking

This article introduces a method to extract the speed and density of microparticles in real time at several kHz using an asynchronous event-based camera mounted on a full-field optical coherence tomography (FF-OCT) setup. These cameras detect significant amplitude changes, allowing scene-driven acquisitions. They are composed of an array of autonomously operating pixels. Events are triggered when an illuminance change at the pixel level is significant at 1μs time precision. The event-driven FF-OCT algorithm relies on a time-based optical flow computation to operate directly on incoming events and updates the estimation of velocity, direction and density while reducing both computation and data load. We show that for fast moving microparticles in a range of 0.4 - 6.5mm/s, the method performs faster and more efficiently than existing techniques in real time. The target application of this work is to evaluate erythrocyte dynamics at the microvascular level in vivo with a high temporal resolution.

268. Optogenetic Engineering of Retinal Ganglion Cells with AAV2.7m8-ChrimsonR-tdTomato (GS030-DP) Is Well Tolerated and Induces Functional Responses to Light in Non-Human Primates

Introduction: Expression of a light-sensitive opsin in retinal ganglion cells (RGCs) is an attractive strategy to restore vision. We evaluated the ability of ChrimsonR-tdTomato (ChrR-tdT), derived from the algal light-gated cation channel ChrimsonR (Ed Boyden, MIT), to convert light insensitive RGCs into photoactivatable cells in normal macaques. A photostimulation device (GS030-MD) is developed in parallel to complement the biologics. This GS030 combination treatment is intended to treat blindness caused by retinal degenerative diseases such as retinitis pigmentosa. Methods: Cynomolgus macaques were injected intravitreally with the AAV2.7m8 vector encoding ChrR-tdT under the control of the CAG promoter (GS030-DP; 5×1011 vg/eye). Electrophysiological measurements by microelectrode array (MEA) and patch clamp as well as expression of the ChrR-tdT protein by immunofluorescence were assessed on explanted retinas 2 months after injection. Local tolerance was evaluated by ophthalmic examination and histology at 2 and 6 months post administration. Results: ChR-tdT was essentially expressed in RGCs and its expression restricted to the perifoveal area. MEA recordings showed light responses in all treated retinas, with 3 out of 4 retinas displaying high amplitudes of electrical responses to light stimulation (up to 360 Hz). One retina was less responsive (50 Hz). In patch clamp experiments, conducted by targeting tdT-expressing RGCs, large photocurrents were recorded in 3 out of 4 retinas in response to illumination, and according to the expected action spectrum for ChR-tdT. An exploratory study was conducted in parallel in monkeys, which received a single bilateral intravitreal administration of GS030-DP (5×1011 vg/eye) to assess local tolerance, systemic toxicity and immunogenicity at 2 and 6 months. No clinical signs indicative of systemic toxicity or local intolerance were observed. No adverse effects were seen by ophthalmic or histological examinations, especially no retinal structural modifications, inflammation or necrosis. Anti-AAV2 neutralizing antibodies (NAbs) measured in serum at 2, 3 and 6 months slightly increased at month 2 (≤ 1:100) and then returned to baseline levels at month 6. No NAbs were detected in aqueous humor at necropsy (at 2 or 6 months). In parallel, in preparation of a first-in-man clinical trial, a complete prototype of the photostimulation device (“goggles”) was developed with a full functional optical chain, an electronic subsystem, and firmware and software architecture. These goggles (GS030-MD) capture external scenes through an event-based camera and deliver visual stimuli onto the transduced retina at irradiances shown to activate ChrR-tdT expressing RGCs in monkey retinas. Conclusion: GS030 vector (GS030-DP) transduced efficiently and safely RGCs in vivo after intravitreal administration and induced light responses in normal monkey retinas under pharmacological block of endogenous phototransducion. The GS030 treatment combining the AAV2.7m8-ChrR-tdT vector and the photo stimulation goggles represents a valuable treatment for vision restoration in retinitis pigmentosa.

Live demonstration: A stimulation platform for optogenetic and bionic vision restoration

Optogenetics can be used to restore light responses in patients affected by retinal degenerative diseases. The light-sensitivity of the molecule introduced by genetic therapy is however very limited in terms of wavelength and irradiance needed to activate a useful neural response, and thus needs an external device to be correctly stimulated. Moreover, the visual signal needs to be encoded so as to respect the code used by the target cells (e.g. retinal ganglion cells). In this demonstration we present a platform that can be used to stimulate optogenetically-treated retinal cells, and the algorithms associated with different types of stimulations. We show different stimulation strategies, varying accordingly to the type of cells transfected.

Adding the third dimension on adaptive optics retina imager thanks to full-field optical coherence tomography

Retinal pathologies, like ARMD or glaucoma, need to be early detected, requiring imaging instruments with resolution at a cellular scale. However, in vivo retinal cells studies and early diagnoses are severely limited by the lack of resolution on eye-fundus images from classical ophthalmologic instruments. We built a 2D retina imager using Adaptive Optics to improve lateral resolution. This imager is currently used in clinical environment. We are currently developing a time domain full-field optical coherence tomograph. The first step was to conceive the images reconstruction algorithms and validation was realized on non-biological samples. Ex vivo retina are currently being imaged. The final step will consist in coupling both setups to acquire high resolution retina cross-sections.