Joseph F. Rizzo - US grants

Human retinal ganglion cell sub-populations have not been systematically studied with modern techniques. There is compelling evidence in animal species that ganglion cell shape is correlated with function. We propose to create a morphologic taxonomy of the human ganglion cells by applying new techniques that have been developed for staining ganglion cells and filling their dendrites with marker molecules. Specifically, we will utilize fluorescent dyes and single cell injections to characterize these cells. These fluorescent dyes have the ability to selectively stain sub-populations of cells, thus permitting rapid identification of cell types over wide areas of the whole-mount retina. Single-cell injections are used to definitively demonstrate the cell shape. This technique has the special advantage of being applicable to post-mortem tissue, thereby making it possible to study human retina. Clinical experience suggests that optic neuropathies represent a heterogenous group of disorders. For example, in a disease such as glaucoma, the early neuropathy may be due to a selective vulnerability of certain sub-classes of ganglion cells. If this is so, there may be (in this condition and others) sub-clinical visual pathology not detected by the usual visual tests because only a few functional types of retinal ganglion cells are damaged. As an initial approach to the question, we will follow the loss of identified ganglion cell types in rabbits after experimental injury to the optic nerve. Better characterization of human ganglion cell types is of interest in its own right, and also for comparison with the well characterized ganglion cell populations in animals. If it is true that diseases of the optic nerve may preferentially affect selected types of retinal ganglion cells, this knowledge may lead to the development of more specific tests of visual function and the detection of more subtle disease states.

One of the great challenges of neuroscience is to understand how nerve cells act synergistically to create perception, thought, and emotion. The PI has for the last 16 years directed the Boston Retinal Implant Project, the goal of which is to develop a prosthesis to restore vision to the blind. An extrapolation of this work strongly suggests that even a perfectly engineered device will not succeed in producing high quality vision. The missing factor is the knowledge of how to communicate with the central nervous system, of which the retina is a part. The neural code is complex; developing insights into its properties will occur gradually, and over a much longer period of time than this grant will cover. The PI's goals for this phase of the project are to develop platform technologies to advance the study of neural coding, and to perform learning experiments that will provide new information about the natural coding of the visual system and how artificial stimuli can be delivered to emulate natural responses. Using the existing strengths of his research group, the PI will build devices that will make it possible to record and wirelessly transmit neural responses from large regions of the retina or brain, with the ultimate goal of capturing these responses in awake animals as they roam freely within a test environment. Computational strategies will be used to compare neural responses that are generated by stimulating the retina with light to those generated by stimulating the retina with electricity delivered in the same geometric patterns. Learning algorithms will be used to adjust the patterns of electrical stimulation to emulate the natural light-induced responses. Collectively, these studies will provide new insights into properties of neural coding and how the natural responses of the brain can be emulated to create vision, and take us a few steps further along the path toward restoration of function to disabled patients.

Broader Impacts: This work will increase the probability that a retinal prosthesis will one day restore useful vision to blind patients. Many millions of patients with age-related macular degeneration and another 1.6 million with retinitis pigmentosa suffer from blindness because of a relatively selective loss of photoreceptors. A retinal prosthesis could restore vision to such patients by directly stimulating the nerve cells that connect the eye to the brain. This hope will be realized only if proper communication strategies to deliver visual information to the brain are learned. This research will also train students in engineering and biomedical disciplines who will contribute to the emerging fields of neural coding, brain-machine interfaces and the science of learning.

We propose to develop and improve a novel minimally-invasive retinal prosthesis design. The goal is to restore a limited but useful level of vision to patients blind with retinitis pigmentosa or macular degeneration. The implant will be driven wirelessly, with almost the entire bulk of the implant attached to the outer wall (sclera) of the eye. Only a thin microelectrode array will penetrate the sclera to electrically stimulate the retina from beneath. This minimally invasive design avoids intrusive vitreal surgery, the need for tacks or glue for attachment to the retina, heating of the retina by intraocular electronics, and motion-induced retinal stress from the implant. It can also be removed without major difficulty if needed. We will develop our existing design and prototype in the following three major areas for eventual human use: 1) We will develop a high-feedthrough hermetic micropackage to protect the implant electronics from bodily fluids. It will be thin, contoured to the curvature of the eye, surgically convenient to implant and biocompatible. This is the only method that will protect the electronics for the ten year minimum required by the FDA. The initial design will allow for 200 electrically conducting pins to pass through the case to stimulate almost 200 electrodes, over 3 times as many as any other hermetically sealed design currently available. We will also further develop techniques for surgical implantation. 2) For the thin microelectrode array that penetrates the sclera, we will develop a waterproof silicon carbide encapsulation with a biocompatible polymer coating to prevent dense cellular overgrowth that can hinder electrical stimulation. In preventing cellular overgrowth, the polymer coating also enables surgical removal of the device, if that were to become necessary months or years after implantation. The coating will be covalently attached for firm adhesion, sufficiently dense to prevent proteins or cells from approaching the surface of the array, and capable of holding and releasing anti-inflammatory agents and other drugs. 3) We will make the implanted electronics resistant to electrical noise and interference, add a system to control power transmission from outside to increase battery life, and increase the voltage swing of the electrode driver circuits to enable stimulation of the retina with larger, shorter current pulses. We will carry out a number of implantation experiments in the eye of the Yucatan minipig to test the design for correct contour, surgical convenience and long-term biocompatibility. An outside vendor laboratory will conduct cytotoxicity tests on device materials, implant prototypes and candidate polymer coatings for biocompatibility. Please Note: In this revision, which NIH requested under the American Recovery and Reinvestment Act (ARRA) of 2009, we have been asked to reduce the proposal to a two-year duration. The additional research assistant and research scientist we have requested will make it possible to complete all the work outlined in the revised project summary above in two years. The reductions from the original three-year proposal are: (i) under Area 1), we will not be able to perform the third year's surgical trials of the hermetic package, under Area 2) we will be able to begin but not complete the proposed work on accelerated in-vitro testing of the multilayered electrode arrays, and we will not be able to synthesize coatings based on triblock polymers or compare the drug-release kinetics of covalently-bonded vs physically adhered micelles, and (iii) the animal implantation experiments will be limited to two years and16 Yucatan mini-pigs rather than the three years and 24 mini-pigs originally proposed.

This project represents an ongoing collaboration between teams at two institutions. As people live longer, blindness caused by degenerative diseases of the retina such as macular degeneration or retinitis pigmentosa is today a major disability among the aging in the developed world. These types of "neural" blindness cannot currently be medically treated in any satisfactory manner. There is now compelling experimental evidence in humans that even when such diseases cause a loss of photoreceptors (i.e., rod and cone cells in the retina), electrical stimulation of the remaining retinal neurons that survive this loss can be used to bypass the damaged tissue and deliver visual information to the brain. This is essentially the same concept that supported the development of the cochlear prosthesis, which has been a fabulous success, restoring hearing to many tens of thousands of deaf patients. The PIs and their respective teams have been working for over 20 years toward the goal of developing a retinal prosthesis to restore truly useful vision to patients in an analogous manner. With prior funding from a number of agencies including NSF, they have created enabling technology for a miniaturized high-density implantable wirelessly-driven neuro-prosthesis package with over 200 individually-addressable channels, which is over three times the inputs and outputs in any current commercially available neurostimulator. The field's ability to create complex integrated circuitry for neurostimulation and/or recording has outpaced the development of long-term implantable packaging, microelectrode array, and assembly technology. If optimized, those technologies would make possible new devices that interface with hundreds of neural tissue sites simultaneously. This is the PI's aim in the current project. The funding will complement existing grants to the PIs and their collaborators, and will allow them to complete development of a new 200+ channel co-fired ceramic signal feed-through disc, to optimize the micro-fabrication process for high-density microelectrode arrays that interface with neural tissue, and to improve the bonding and interconnection processes required to assemble the implant package.

Broader Impacts: The 200+ channel wirelessly-driven implant that will constitute the primary project outcome will have over three times the number of individually-addressable stimulating electrodes now available from any group. This funding will further allow the PI to ready devices for later pre-clinical testing (with anticipated follow-on support from the VA). Project results will be widely disseminated in publications, and by distributing sample devices within the rehabilitation R&D community. The device which is the focus of this project will also be useful in a myriad other future chronically implantable prosthetics, palliative devices, and human-computer interface devices.

DESCRIPTION (provided by applicant): This proposal seeks to conduct FDA-required pre-clinical testing of a retinal prosthesis designed and assembled by our team at Harvard Medical School and the Massachusetts Institute of Technology. Our retinal prosthesis is designed to restore vision to patients who are blind from either retinitis pigmentosa (RP) or age-related macular degeneration (AMD). There is no treatment to restore lost vision for either disease, although promising research in other fields also offers some hope of being able to help these patients. RP is the leading cause of inherited blindness in the world and is particularly devastating because it causes a slowly progressive loss of vision across the entire visual field of both eyes. AMD, which is the leading cause of blindness in the industrialized world, causes loss of central vision that robs a patient of being able to read or easily recognize faces. Our prosthetic system is composed of two components. The first component is a modified pair of glasses that contains an ultra-small camera that captures details of a visual scene; the glasses are connected to a cell phone-like device that processes the visual images. The digital visual signal and operating power are sent wirelessly to the second component of the prosthetic system, which is implanted around the back of the eye. The implanted component receives the visual signal and a custom-designed computer chip uses the incoming operating power to distribute electrical pulses to the retina. These electrical pulses will initiate nerve impulses t the brain via the optic nerve. The primary goal of this prosthesis is to improve the quality-of-lie of severely blind patients by allowing them to navigate in unfamiliar environments. Once achieved, the device also has the potential to facilitate other activities of daily living, like reding street signs, addresses on buildings, bathroom designations (i.e. male vs. female), and finding and reaching accurately for glasses, plates, and utensils, among other things. The quality of vision that can be restored depends substantially upon the number of stimulation channels, or pixels. Our device has the largest number (256) of individually-controllable stimulation channels of any neural prosthetic device in the world. This relatively large number of stimulation channels should provide higher quality vision compared to the other devices. Following completion of the work described in this grant proposal, our team would hope to receive an FDA approval to conduct a subsequent Phase I safety study of our device.

? DESCRIPTION (provided by applicant): This proposal seeks to conduct FDA-required pre-clinical testing of a retinal prosthesis with penetrating electrodes designed and assembled by our team at Harvard Medical School and the Massachusetts Institute of Technology. Our retinal prosthesis is designed to restore vision to patients who are blind from either retinitis pigmentosa (RP) or age-related macular degeneration (AMD). There is no treatment to restore lost vision for either disease, although promising research in other fields also offers some hope of being able to help these patients. RP is the leading cause of inherited blindness in the world and is particularly devastating because it causes a slowly progressive loss of vision across the entire visual field of both eyes. AMD, which is the leading cause of blindness in the industrialized world, causes loss of central vision that robs a patient of being able to read or easily recognize faces. Our prosthetic system is composed of two components. The first component is a modified pair of glasses that contains an ultra-small camera that captures details of a visual scene; the glasses are connected to a cell phone-like device that processes the visual images. The digital visual signal and operating power are sent wirelessly to the second component of the prosthetic system, which is implanted around the back of the eye. The implanted component receives the visual signal and a custom-designed computer chip uses the incoming operating power to distribute electrical pulses to the retina. These electrical pulses will initiate nerve impulses tothe brain via the optic nerve. The primary goal of this prosthesis is to improve the quality-of-life of severely blind patients by enabling navigation in unfamiliar environments. Once achieved, the device also has the potential to facilitate other activities of daily living, like reading street sgns, addresses on buildings, bathroom designations (i.e. male vs. female), and finding and reaching accurately for glasses, plates, and utensils, among other things. The quality of vision that can be restored depends substantially upon the number of stimulation channels, or pixels. Our device has the largest number (256) of individually-controllable stimulation channels of any neural prosthetic device in the world. This relatively large number of stimulation channels should provide higher quality vision compared to the other devices. The penetrating electrodes should increase the safety of electrical stimulation and improve the quality of vision over what has been obtained with retinal prosthetic devices that use planar electrodes. Following completion of the work described in this grant proposal, our team would hope to receive an FDA approval to conduct a subsequent Phase I safety study of our retinal prosthesis with penetrating electrodes.

Under an existing funded grant, Dr. Rizzo (Principal Investigator, from Massachusetts Eye and Ear Infirmary) and his subcontractor Bionic Eye Technologies (Ithaca, NY) have worked to develop vision prostheses to restore functional sight to people with profound vision impairment. As part of this work, we have optimized a 324-channel wirelessly transmitter/receiver that provides power and data to an implanted high-density stimulator circuit (ASIC chip) behind the eye. This device represents a significant increase in the independently-controllable channel count over any commercially available neural stimulator. During the course of preparing units for a program of ?pre-clinical? testing for the device that has been vetted by the FDA, unanticipated problems were discovered with the ASIC chip?s wireless communication module. Our team has developed an effective a work- around solution, and this supplemental proposal seeks additional funds to validate the performance of our low- power, programmable, gate array-based wireless transceiver circuit. Successful validation of our ASIC + work- around solution will provide highly sophisticated circuitry that could be incorporated into a wide array of low- power implantable neuroprosthetic devices. Sub-awardee Bionic Eye will seek to take commercial advantage of this circuitry by offering licensing opportunities to other companies to disseminate this technology beyond what could be realized within the field of sight restoration alone.

SUMMARY Vascular dysfunction, with or without elevated intraocular pressure (IOP), is believed to be an important risk factor in glaucoma and other neuropathies. However, the link between vascular dysfunction and the mechanisms leading to the characteristic visual field defects in glaucoma is not fully understood. This is partly due to the lack of a solid quantitative understanding of the 3D architecture of the vasculature of the optic nerve head (ONH), its anatomical relationship with the load-bearing connective tissues, and how it is affected by IOP. Our overarching hypothesis is that features of the vasculature and its relationship with the connective tissues predispose certain ONH regions to compromised perfusion and that this susceptibility is amplified by elevated IOP. To test this hypothesis, we will sequentially collect in vivo, ex vivo, and histological 3D morphological and biomechanical data on vascular and connective tissues of the ONH in normal eyes and in eyes with experimental glaucoma (EG). We will focus on the critical lamina cribrosa (LC) region in three species: human, monkey (closest model to human, collagenous LC), and mouse (most used model, no collagenous LC). In Aim 1, we will map in 3D the vasculature and connective tissues of the ONHs of humans, monkeys, and mice, and analyze these maps quantitatively including by watershed analysis. We predict that zones of visual loss in early glaucoma will correspond to regions with the most vulnerable vascular supply, e.g., sparse capillaries with low connectivity and low perfusion redundancy. We postulate that, in primates, not all LC beams have a capillary, and conversely, that some capillaries are not within a robust collagen-rich beam. We will also address the clinically important question to which extent in vivo OCT angiography visualizes the smaller or deeper vessels inside the ONH. In Aim 2, we will perform ex vivo inflation tests on monkey and mouse eyes to quantify the effects of acute IOP elevation on vessel perfusion and biomechanics, and the LC beams support. Our preliminary data suggests that ?unprotected? vessels may be particularly vulnerable to mechanical distortion, which could, in turn, affect blood flow. In Aim 3, we will characterize the effects of chronic IOP elevation on vessels and beams. Specifically, we will compare eyes before and after chronic IOP elevation (EG), and with the contralateral control. This will allow us to discern characteristics that pre-dispose an eye to glaucoma from those that are the result of the disease. We will test the hypothesis that the patterns of vessel sensitivity to elevated IOP in mice (that have only a glial lamina) are different from those in primates. Combining multiple imaging modalities across the same ONHs in three species will provide cross-verification of the techniques, and deeper insights into the role of LC collagenous beams supporting the ONH vasculature under normal and elevated IOP. These experiments will help identifying ONH features that predict susceptibility to neural injury and vision loss.