Brillouin Microscopic Assessment of Retinal Stiffness During Reactive Müller Gliosis

Increased retinal stiffness can indicate reactive gliosis, fibrosis, or ECM remodeling – all hallmarks of early retinal damage due to diseases such as age-related macular degeneration (AMD), glaucoma, and diabetic retinopathy. Here, we demonstrate that Brillouin microscopy can detect increased stiffness within the damaged mouse retina coincident with reactive Müller gliosis. Overall design: ROSA26R-Ai9+/ki and Glast-CreERT2+/tg mice (both on a C57BL6/J background). Glast-CreERT2+/tg; ROSA26R-Ai9+/ki mice were induced with intraperitoneal (IP) injections of tamoxifen (Sigma, T5648) (75 mg/kg body weight) were given for five consecutive days. For intravitreal NMDA injections, mice were anesthetized using Isoflurane. One drop of Proparacaine Hydrochloride (0.5%) was placed on to the eye to be injected. The anesthetized mouse was positioned on its side under a dissecting microscope while on a warming plate and a sterile 28G hypodermic needle in inserted in the post-limbus region, with the bevel of the needle facing upwards to avoid the lens. The puncture needle was then carefully removed, and a small amount of vitreous fluid was wicked away from the puncture site using a twisted Kimwipe. Then, using a Hamilton syringe, a sterile blunt needle (33G) was gently inserted into the hole, pass the lens, and into the vitreous chamber where 2 microliters of NMDA (100 mM) (Sigma: M3262) was injected. The contralateral eye served as an un-injected control. After 24 hours, eyes were processed for FACs and subsequent scRNA-seq.