In Vivo Small Molecule Delivery to the Optic Nerve in a Rodent Model

Abstract

Small molecule delivery to the optic nerve would allow for exploration of molecular and cellular pathways involved in normal physiology and optic neuropathies such as glaucoma, and provide a tool for screening therapeutics in animal models. We report a novel surgical method for small molecule drug delivery to the optic nerve head (ONH) in a rodent model. In proof-of-principle experiments, we delivered cytochalasin D (Cyt D; a filamentous actin inhibitor) to the junction of the superior optic nerve and globe in rats to target the actin-rich astrocytic cytoskeleton of the ONH. Cyt D delivery was quantified by liquid chromatography and mass spectrometry of isolated optic nerve tissue. One day after Cyt D delivery, anterior ONH filamentous actin bundle content was significantly reduced as assessed by fluorescent-tagged phalloidin labeling, relative to sham delivery. Anterior ONH nuclear counts and axon-specific beta-3 tubulin levels, as well as peripapillary retinal ganglion cell layer nuclear counts were not significantly altered after Cyt D delivery relative to sham delivery. Lastly, the surgical delivery technique caused minimal observable axon degeneration up to 10 days post-surgery. This small molecule delivery technique provides a new approach to studying optic neuropathies in in vivo rodent models.

Optic Nerve Head Astrocytes Display Axon-Dependent and -Independent Reactivity in Response to Acutely Elevated Intraocular Pressure

“…2A) that delineate astrocyte extensions, with high concordance with astrocyte-specific markers such as glial fibrillary acidic protein (GFAP) and aquaporin 4 10. As previously reported,10,21,37 control ONH astrocytes exhibit highly arranged, actin-based extensions perpendicular (84.3° ± 1.0°) to the anterior-posterior and axonal microtubule axes (Fig. 2B, top row; Fig.…”

“…The eye was then infero-ducted by grasping the remaining superior rectus muscle insertion using 0.12-mm toothed forceps. The junction of the superior optic nerve and the globe was exposed using a two-pronged instrument modified from a Lewis Lens Loop,37 with care to avoid any trauma or excess stretch to the optic nerve. Next, a 25-G microvitreoretinal blade was used to carefully puncture the superior optic nerve sheath, as far away from the globe as possible.…”

The effects of PEGylation on LNP based mRNA delivery to the eye

“…Specific to the TM, researchers have been investigating and developing gene therapy approaches that reduce ocular pressure [ 37 ]. As glaucoma is a multifaceted disease, researchers have also developed a method of small molecule delivery to the ONH of rats that allows for exploration of molecular and cellular pathways involved in neuropathies and treatment screening [ 38 ]. In future studies our LNPs could be used to target the Müller glia, the ONH and the TM to aid in treatment development for retinal degeneration and glaucoma.…”

“…The TM is most likely transduced as the particles get circulated out of the vitreous chamber. The TM and the ONH are commonly under investigation to better understand and treat glaucoma [36][37][38]. Specific to the TM, researchers have been investigating and developing gene therapy approaches that reduce ocular pressure [37].…”

Factors affecting optic nerve head biomechanics in a rat model of glaucoma

“…Further, the rat has benefits for biomechanics studies over the mouse due to its larger eye size, which makes its tissues easier to handle for biomechanical testing. In addition, rat ocular hypertensive models present with important characteristics of human glaucoma, and rat models of both acute and chronic IOP elevation exist, allowing early and late events in the disease process to be studied [7][8][9].…”