Our research will investigate how critical axonal microtubules are in the pathogenesis of glaucoma. The axons of the retinal ganglion cells (RGCs) are gradually lost in glaucoma, but little is known about the mechanistic link between ocular hypertension, which is a major risk factor, and the loss of RGC axons. Our overarching hypothesis is that RGC microtubules are involved in a reversible stage of the disease and may provide a molecular substrate for the RGC’s sensitivity to the elevated intraocular pressure triggering the pathogenic pathway. Recently, we demonstrated that axonal microtubules degrade more rapidly than the loss of RGC axons, resulting in microtubule deficit, and microtubule deficit is spatially correlated with axonal atrophy. It suggests that microtubule deficit is a new pathology of early glaucoma, which shares a common pathogenic origin as the loss of RGC axons. Here we will investigate microtubule deficit in depth. The potential mechanism and therapeutic significance will be studied using DBA mice as a model of inherited glaucoma. In aim 1, we will investigate the possible relationship between the posttranslational modifications of axonal microtubules and microtubule deficit. Also known as the tubulin code, various posttranslational modifications modulate the interaction with microtubule-associated proteins to regulate the stability and function of microtubules. We will examine how the tubulin code is altered during glaucoma and how it is correlated with other glaucoma pathologies such as microtubule deficit. In aim 2, we will test microtubule deficit as an endpoint to evaluate the efficacy of therapeutic treatments. Microtubule deficit will be measured of the DBA retinas receiving an increased level of nicotinamide adenine dinucleotide, which is known to protect the RGC axons and promote the RGC viability. The recovery of microtubules will suggest a crucial causal relationship with metabolic stress. Upon completion, our research will yield new insights into the mechanism of microtubule deficit toward improved glaucoma therapy. The notion that the tubulin codes might play a role in glaucoma pathogenesis is innovative. Also, new types of data will be obtained due to a novel retinal imaging, i.e., intrinsic second-harmonic generation (SHG) microscopy. Consequently, our research will open a new field of questions and can be paradigm-shifting in glaucoma therapy.