Denise M. Inman, Ph.D. - US grants

? DESCRIPTION (provided by applicant): Retinal ganglion cell axons undergo degeneration prior to cell body loss in the pathogenesis of glaucoma. Prior to degeneration, retinal ganglion cell axons demonstrate metabolic dysfunction, including ATP decreases that correlate with high intraocular pressure. This metabolic dysfunction could be a result of changes in axonal mitochondria or alterations in the provision of energy substrates from glia to axons. A critical question for therapeutic development is whether the metabolic dysfunction is intrinsic to the retinal ganglion cell axon. The long-term goal of this work is to determine the onset, progression and mechanisms of retinal ganglion axon dysfunction so that crucial intervention points for developing glaucoma therapies can be defined and prioritized. The overall objective of this proposal is to examine the causes of energy availability failure in early glaucoma and place those causes in context with structural pathology. The central hypothesis of this work is that the mechanism of metabolic dysfunction is shared by the RGC axon mitochondria and optic nerve glia. This hypothesis has been formulated from preliminary data gathered in the applicant's laboratory that suggests deficits in mitochondrial recycling, low glycogen stores and lactate transporters in glaucomatous optic nerve. The rationale for this proposed work is that investigating the source of metabolic dysfunction in glaucomatous optic nerve will identify new therapeutic targets in a disease that has no mechanism-based interventions. Guided by strong preliminary data, this hypothesis will be tested by the following specific aims: 1) determine whether mitochondria underlie the metabolic vulnerability of glaucoma by investigating ROS production and mitochondria quality control; 2) by establishing the relationship between mitochondrial respiration and degeneration; and 3) by determining how glia drive metabolic vulnerability through energy store levels and mobilization, lactate transporter expression and distribution, and metabolic enzyme activity. Overall, this work will determine the contribution of energy availability to glaucoma pathogenesis by analyzing how mitochondrial and glial-related energy substrates and delivery mechanisms are altered in optic neuropathy. Confirmation will occur through manipulation of mitochondrial quality control, reactive oxygen species levels, and energy stores to the nerve, and quantifying the resultant decrease in axon degeneration. The approach is innovative because it departs from the general perspective that energy availability and utilization are tangential to optic neuropathy. Critical intervention points to halt optic neuropathy will emerge from this analysis. For this reason, the proposed research is significant, but also because it will provide insight into the role of energy in axon degeneration generally.

Project Summary Retinal ganglion cell (RGC) axons degenerate after functional decline in glaucoma, a circumstance potentially caused, but certainly exacerbated by, energy deficit, indicating that metabolic dysfunction contributes to glaucomatous degeneration. High intraocular pressure (IOP) in glaucoma results in hypoxia that promotes glycolysis through upregulation of glycolysis-associated genes, but also mitochondrial recycling, and downregulation of genes encoding for oxidative phosphorylation (OxPhos). A critical question is whether RGC metabolic reprogramming from OxPhos to glycolysis is initiated by IOP-associated hypoxia, and whether repeated reprogramming and development of pseudohypoxia underlies the significant energy stress and dysfunction observed in the glaucomatous retina. The long-term goal of this work is to leverage our understanding of retinal metabolism to maintain visual function despite the stressors inherent in glaucomatous neurodegeneration. The overall objective of this proposal is to determine the impact of IOP-mediated damage on retinal and optic nerve head metabolic resilience. Our central hypothesis is that ocular hypertension (OHT)-associated hypoxia transitions into pseudohypoxia, destabilizing neural and glial metabolism and mitochondrial homeostasis, with downstream negative impact on RGCs in the retina and ONH. This hypothesis derives from our recent analysis indicating that mitochondria and availability of energy substrates, both targets of hypoxia-associated regulation, contribute to the metabolic challenges of RGCs. The rationale for the work is that investigating metabolic cooperation and dysfunction in glaucomatous retina will identify new therapeutic targets for a disease that has no mechanism-based interventions. Guided by strong preliminary data, the aims of this proposal will address a critical element of the retinal energy management in glaucoma by revealing 1) whether OHT is the impetus for metabolic reprogramming, followed by adaptation as pseudohypoxia, in the glaucomatous retina; 2) how metabolic coupling is managed among neurons and glia in the normal and ocular hypertensive retina; 3) how RGCs, Müller glia, and ONH astrocytes respond to and manage metabolic challenge on a cell-by-cell basis. Overall, this work will enable us to determine how RGCs, Müller glia, and ONH astrocytes respond metabolically to IOP increase, whether those changes persist long-term, and how we might promote metabolic resilience. The approach is innovative by taking the position that insight will only come from investigating the metabolic dependencies of the primary cell types impacted during the development of glaucoma pathology. In addition, we will apply new cutting-edge approaches to the investigation of mitochondria such as cell- specific mitochondrial isolation followed by metabolomics and co-detection of protein and gene expression through spatial transcriptomics. This proposal is significant because we will reveal mitochondrial-specific metabolic dysregulation amenable to therapeutic intervention.