Alterations of Inner Retinal Physiology in Retinal Degenerations, Gene Therapy, and Neurologic Disorders

National Eye Institute

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Abstract

PURPOSE: During this first year at NEI, with 60% effort devoted to developing additional clinical skills and involvement in clinical research projects (reported elsewhere), much of Dr. Stasheffs remaining effort in laboratory research has been devoted to establishing the infrastructure to support retinal electrophysiology experiments conducted with multielectrode techniques (as detailed in the following sections). Additional activities have included further analysis and reporting of prior project areas to be continued at NEI, and determining project priorities for the coming fiscal year. DEVELOPMENT OF ANIMAL RESOURCES: A double-transgenic mouse strain useful for identification of specific retinal ganglion cell types in a widely-studied model of hereditary retinal degeneration had been partly developed during my term as a faculty member at the University of Iowa, but had not yet been bred to a fully-homozygous state. This rd10/Thy1-GFP strain has been re-derived at NEI over the course of the fiscal year, with confirmation of genotype, and the first litter produced from parents putatively homozygous for both rd10 and Thy1-GFP genotypes now awaits confirmatory genotyping. ACQUISITION AND CONFIGURATION OF EXPERIMENTAL RESOURCES: Laboratory space was identified and additional equipment required to construct a multi-electrode recording rig purchased and set up. Placement and adjustment of hardware for direct stimulation of the retina with complex images and videos is nearing completion, together with adaptation of operating system and software to drive such stimulation. Specialized and co-compatible image intensifiers and dissecting microscope were purchased and now are in use for dissection of retinas under infrared illumination. ESTABLISHMENT OF COLLABORATIVE RELATIONSHIPS: A mutually beneficial relationship has been established with NIH Principal Investigators Drs. Jeff Diamond and Tudor Badea and key members of their laboratories as well as Dr. Wei Lis, for mutual exchange of experience and information on multielectrode recording techniques and data analysis methods. We recently formalized this further with the establishment of a special interest group among our laboratories that will meet periodically to optimize a shared system for data analysis with improved efficiency, as well as direct correspondence to in vivo electrophysiologic measures (forms of electroretinography) and behavioral assessment of visual function (such as optokinetic responses OKR). This will be useful for the study of normal retinal physiology (Dr. Diamonds group), the role of specific genetically-identified/controlled ganglion cell types (Dr. Badeas lab), and both retinal degenerations and traumatic brain injury (TBI) (our laboratory). A productive collaboration continues with Drs. Matthew Harper and Laura Dutca of the University of Iowa and the Veterans Affairs Center for Excellence in Vision Loss and Restoration, for research on TBI. A major publication resulting from this work appeared at the end of 2014 in Investigative Ophthalmology and Visual Science, and it also has served as the core for a growing relationship with the Center for Neuroscience and Regenerative Medicine (a consortium between NIH, Walter Reed Military Medical Center, and the Uniformed Services University of the Health Sciences dedicated to TBI research). A major grant application was solicited by the CNRM and currently is under review. SECURING ADDITIONAL RESEARCH FUNDING: At the time of relocating to NIH from the University of Iowa, Dr. Stasheff was completing the second year of a three-year Research Grant from the March of Dimes to support the laboratorys ongoing research into two animal models of childhood-onset retinal degeneration (Lebers congenital amaurosis LCA). Following some effort to confirm mutual agreement on the legal terms, the remaining funds were successfully transferred to NEI. A Letter of Intent for competitive renewal of this March of Dimes funding was submitted and currently is under review. As noted above, an application for a Research Grant from the CNRM currently is under review as well. SECURING PERSONNEL: A part time data analyst with highly specialized experience in precisely the data analysis employed in our laboratory was identified and hired. Potential applicants for a post-baccalaureate experimental assistant/data analyst are being reviewed presently. SUMMER INTERN: A senior undergraduate summer intern was supervised under the Diversity in Vision Research and Ophthalmology (DIVRO) program of NEI. This was of mutual benefit to the intern, NEI in promoting its missions, and the laboratorys progress in completing data analysis required for revising a manuscript for publication describing a new mode of neural encoding in the retina and its degradation in two widely-investigated mouse models of retinal degeneration, rd1 and rd10. DATA ANALYSIS & PREPARATION OF MANUSCRIPTS FOR PUBLICATION: With the establishment of the above resources, necessary data analysis has been nearly completed to allow revision or first submission of manuscripts reporting the following: pharmacologic dissection of mechanisms driving spontaneous hyperactivity in rd1 and rd10 models of retinal degeneration changes in quantitative relationships describing the response of rd1 ganglion cells to graded stimuli a new mode of neural encoding in retinal ganglion cells, its quantification, and evidence that it is disrupted in rd1 and rd10 mice In addition, Dr. Stasheff is the invited Associate Editor for a Research Topic published by Frontiers in Cellular Neuroscience, a series to include 14 publications on disruptions of visual processing accompanying retinal degenerations, including his own Perspective on the clinical implications of basic science findings in this discipline. PRESENTATIONS AT NATIONAL MEETINGS: Oct 2014 Invited Speaker, The Eye and the Chip 2014, Detroit Institute of Ophthalmology Feb 2015 Annual Meeting of the North American Neuro-Ophthalmology Society (NANOS) May 2015 Annual Meeting of the Association for Research in Vision and Ophthalmology (ARVO)