Acute ocular hypertension in the living human eye: Model description and initial cellular responses to elevated intraocular pressure.

This initial methods study presents the initial immunohistochemical and transcriptomic changes in the optic nerve head and retina from three research-consented brain-dead organ donors following prolonged and transient intraocular pressure (IOP) elevation. In this initial study, research-consented brain-dead organ donors were exposed to unilateral elevation of IOP for 7.5 h (Donor 1), 30 h (Donor 2), and 1 h (Donor 3) prior to organ procurement. Optic nerve tissue and retinal tissue was obtained following organ procurement for immunohistological and transcriptomic analysis. Optic nerve sections in Donor 1 exposed to 7.5-hours of unilateral sub-ischemic IOP elevation demonstrated higher levels of protein expression of the astrocytic marker, glial fibrillary acidic protein (GFAP), within the lamina cribrosa with greatest expression inferior temporally in the treated eye compared to control. Spatial transcriptomic analysis performed on optic nerve head tissues from Donor 2 exposed to 30 h of unilateral IOP elevation demonstrated differential transcription of mRNA across laminar and scleral regions. Immunohistochemistry of retinal sections from Donor 2 exhibited higher GFAP and IBA1 expression in the treated eye compared with control, but this was not observed in Donor 3, which was exposed to only 1-hour of IOP elevation. While there were no differences in GFAP protein expression in the retina following the 1-hour IOP elevation in Donor 3, there were higher levels of transcription of GFAP in the inner nuclear layer, and CD44 in the retinal ganglion cell layer, indicative of astrocytic and Müller glial reactivity as well as an early inflammatory response, respectively. We found that transcriptomic differences can be observed across treated and control eyes following unilateral elevation of IOP in brain dead organ donors. The continued development of this model affords the unique opportunity to define the acute mechanotranscriptomic response of the optic nerve head, evaluate the injury and repair mechanisms in the retina in response to IOP elevation, and enable correlation of in vivo imaging and functional testing with ex vivo cellular responses for the first time in the living human eye.

Frmpd1 Facilitates Trafficking of G-Protein Transducin and Modulates Synaptic Function in Rod Photoreceptors of Mammalian Retina.

Trafficking of transducin (Gαt) in rod photoreceptors is critical for adaptive and modulatory responses of the retina to varying light intensities. In addition to fine-tuning phototransduction gain in rod outer segments (OSs), light-induced translocation of Gαt to the rod synapse enhances rod to rod bipolar synaptic transmission. Here, we show that the rod-specific loss of Frmpd1 (FERM and PDZ domain containing 1), in the retina of both female and male mice, results in delayed return of Gαt from the synapse back to outer segments in the dark, compromising the capacity of rods to recover from light adaptation. Frmpd1 directly interacts with Gpsm2 (G-protein signaling modulator 2), and the two proteins are required for appropriate sensitization of rod-rod bipolar signaling under saturating light conditions. These studies provide insight into how the trafficking and function of Gαt is modulated to optimize the photoresponse and synaptic transmission of rod photoreceptors in a light-dependent manner.

Remodeling of the Lamina Cribrosa: Mechanisms and Potential Therapeutic Approaches for Glaucoma.

Glaucomatous optic neuropathy is the leading cause of irreversible blindness in the world. The chronic disease is characterized by optic nerve degeneration and vision field loss. The reduction of intraocular pressure remains the only proven glaucoma treatment, but it does not prevent further neurodegeneration. There are three major classes of cells in the human optic nerve head (ONH): lamina cribrosa (LC) cells, glial cells, and scleral fibroblasts. These cells provide support for the LC which is essential to maintain healthy retinal ganglion cell (RGC) axons. All these cells demonstrate responses to glaucomatous conditions through extracellular matrix remodeling. Therefore, investigations into alternative therapies that alter the characteristic remodeling response of the ONH to enhance the survival of RGC axons are prevalent. Understanding major remodeling pathways in the ONH may be key to developing targeted therapies that reduce deleterious remodeling.

Melanopsin modulates refractive development and myopia.

Myopia, or nearsightedness, is the most common form of refractive abnormality and is characterized by excessive ocular elongation in relation to ocular power. Retinal neurotransmitter signaling, including dopamine, is implicated in myopic ocular growth, but the visual pathways that initiate and sustain myopia remain unclear. Melanopsin-expressing retinal ganglion cells (mRGCs), which detect light, are important for visual function, and have connections with retinal dopamine cells. Here, we investigated how mRGCs influence normal and myopic refractive development using two mutant mouse models: Opn4-/- mice that lack functional melanopsin photopigments and intrinsic mRGC responses but still receive other photoreceptor-mediated input to these cells; and Opn4DTA/DTA mice that lack intrinsic and photoreceptor-mediated mRGC responses due to mRGC cell death. In mice with intact vision or form-deprivation, we measured refractive error, ocular properties including axial length and corneal curvature, and the levels of retinal dopamine and its primary metabolite, L-3,4-dihydroxyphenylalanine (DOPAC). Myopia was measured as a myopic shift, or the difference in refractive error between the form-deprived and contralateral eyes. We found that Opn4-/- mice had altered normal refractive development compared to Opn4+/+ wildtype mice, starting ∼4D more myopic but developing ∼2D greater hyperopia by 16 weeks of age. Consistent with hyperopia at older ages, 16 week-old Opn4-/- mice also had shorter eyes compared to Opn4+/+ mice (3.34 vs 3.42 mm). Opn4DTA/DTA mice, however, were more hyperopic than both Opn4+/+ and Opn4-/- mice across development ending with even shorter axial lengths. Despite these differences, both Opn4-/- and Opn4DTA/DTA mice had ∼2D greater myopic shifts in response to form-deprivation compared to Opn4+/+ mice. Furthermore, when vision was intact, dopamine and DOPAC levels were similar between Opn4-/- and Opn4+/+ mice, but higher in Opn4DTA/DTA mice, which differed with age. However, form-deprivation reduced retinal dopamine and DOAPC by ∼20% in Opn4-/- compared to Opn4+/+ mice but did not affect retinal dopamine and DOPAC in Opn4DTA/DTA mice. Lastly, systemically treating Opn4-/- mice with the dopamine precursor L-DOPA reduced their form-deprivation myopia by half compared to non-treated mice. Collectively our findings show that disruption of retinal melanopsin signaling alters the rate and magnitude of normal refractive development, yields greater susceptibility to form-deprivation myopia, and changes dopamine signaling. Our results suggest that mRGCs participate in the eye's response to myopigenic stimuli, acting partly through dopaminergic mechanisms, and provide a potential therapeutic target underling myopia progression. We conclude that proper mRGC function is necessary for correct refractive development and protection from myopia progression.

Mechanisms of retinal ganglion cell injury following acute increases in intraocular pressure.

The maintenance of intraocular pressure (IOP) is critical to preserving the pristine optics required for vision. Disturbances in IOP can directly impact the optic nerve and retina, and inner retinal injury can occur following acute and chronic IOP elevation. There are a variety of animal models that have been developed to study the effects of acute and chronic elevation of IOP on the retina, retinal ganglion cell (RGC) morphology, intracellular signaling, gene expression changes, and survival. Acute IOP models induce injury that allows for the study of RGC response to well characterized injury and potential recovery. This review will focus on the initial impact of acute IOP elevation on RGC injury and recovery as these early responses may be the best targets for potential therapeutic interventions to promote RGC survival in glaucoma.

Ambient Light Regulates Retinal Dopamine Signaling and Myopia Susceptibility.

Purpose: Exposure to high-intensity or outdoor lighting has been shown to decrease the severity of myopia in both human epidemiological studies and animal models. Currently, it is not fully understood how light interacts with visual signaling to impact myopia. Previous work performed in the mouse retina has demonstrated that functional rod photoreceptors are needed to develop experimentally-induced myopia, alluding to an essential role for rod signaling in refractive development. Methods: To determine whether dim rod-dominated illuminance levels influence myopia susceptibility, we housed male C57BL/6J mice under 12:12 light/dark cycles with scotopic (1.6 × 10-3 candela/m2), mesopic (1.6 × 101 cd/m2), or photopic (4.7 × 103 cd/m2) lighting from post-natal day 23 (P23) to P38. Half the mice received monocular exposure to -10 diopter (D) lens defocus from P28-38. Molecular assays to measure expression and content of DA-related genes and protein were conducted to determine how illuminance and lens defocus alter dopamine (DA) synthesis, storage, uptake, and degradation and affect myopia susceptibility in mice. Results: We found that mice exposed to either scotopic or photopic lighting developed significantly less severe myopic refractive shifts (lens treated eye minus contralateral eye; -1.62 ± 0.37D and -1.74 ± 0.44D, respectively) than mice exposed to mesopic lighting (-3.61 ± 0.50D; P < 0.005). The 3,4-dihydroxyphenylacetic acid /DA ratio, indicating DA activity, was highest under photopic light regardless of lens defocus treatment (controls: 0.09 ± 0.011 pg/mg, lens defocus: 0.08 ± 0.008 pg/mg). Conclusions: Lens defocus interacted with ambient conditions to differentially alter myopia susceptibility and DA-related genes and proteins. Collectively, these results show that scotopic and photopic lighting protect against lens-induced myopia, potentially indicating that a broad range of light levels are important in refractive development.

Short-Wavelength (Violet) Light Protects Mice From Myopia Through Cone Signaling.

Purpose: Exposure to short-wavelength light influences refractive development and inhibits myopic development in many animal models. Retinal mechanisms underlying this response remain unknown. This study used a mouse model of lens-induced myopia to evaluate the effect of different wavelength light on refractive development and dopamine levels in the retina. A possible retinal pathway is tested using a mutant mouse with dysfunctional cones. Methods: Wild-type C57BL/6J (WT) and ALS/LtJ/Gnat2cpfl3 (Gnat2-/-) mice were exposed to one of three different light conditions beginning at postnatal day 28: broad-spectrum "white" (420-680 nm), medium wavelength "green" (525 ± 40 nm), and short wavelength "violet" (400 ± 20 nm). One-half of the mice received hyperopic lens defocus. All mice were exposed to the light for 4 weeks; animals were measured weekly for refractive error and axial parameters. Retinal dopamine and the dopamine metabolite 3,4-dihydroxyphenylacetic acid were measured by HPLC. Results: In WT mice, short-wavelength violet light induced hyperopia and violet light inhibited lens-induced myopia when compared with mice exposed to white light. Hyperopia could be attributed to shallower vitreous chambers in WT animals. There were no changes in the levels of dopamine or its metabolite. In Gnat2-/- mice, violet light did not induce hyperopia or inhibit lens-induced myopia. Conclusions: These findings show that short-wavelength light slows refractive eye growth, producing hyperopic responses in mice and inhibiting lens-induced myopia. The lack of inhibition in mice with dysfunctional cones suggests that cone signaling plays a role in the hyperopic response to short-wavelength (violet) light.