Biology subjects

Scalabrino, M. L.

Publications and source records attributed to Scalabrino, M. L..

5 recordsLinked to original sources

bioRxiv2023

Late gene therapy limits the restoration of retinal function in a mouse model of retinitis pigmentosa

Retinitis pigmentosa is an inherited photoreceptor degeneration that begins with rod loss followed by cone loss and eventual blindness. Gene therapies are being developed, but it is unknown how retinal function depends on the time of intervention. To uncover this dependence, we utilized a mouse model of retinitis pigmentosa capable of artificial genetic rescue. This model enables a benchmark of best-case gene therapy by removing the variables that complicate the ability to answer this vital question. Complete genetic rescue was performed at 25%, 50%, and 70% rod loss (early, mid and late, respectively). Early and mid treatment restored retinal function to near wild-type levels, specifically the sensitivity and signal fidelity of retinal ganglion cells (RGCs), the output neurons of the retina. However, some anatomical defects persisted. Late treatment retinas exhibited continued, albeit slowed, loss of sensitivity and signal fidelity among RGCs, as well as persistent gliosis. We conclude that gene replacement therapies delivered after 50% rod loss are unlikely to restore visual function to normal. This is critical information for administering gene therapies to rescue vision.

neuroscience↗

bioRxiv2022

Thousandfold Cell-Specific Pharmacology of Neurotransmission

Cell-specific pharmaceutical technologies promise mechanistic insight into clinical drugs[-]those that treat, and often define, human disease. In particular, DART (drug acutely restricted by tethering) achieves genetically programmable control of drug concentration over cellular dimensions. The method is compatible with clinical pharmaceuticals and amenable to studies in behaving animals. Here, we describe DART.2, comprising three advances. First, we improve the efficiency of chemical capture, enabling cell-specific accumulation of drug to [~]3,000-times the ambient concentration in 15 min. Second, we develop tracer reagents, providing a behavior-independent measure of cellular target engagement in each animal. Third, we extend the method to positive allosteric modulators and outline design principles for this clinically significant class. We showcase the platform with four pharmaceuticals[-]two that weaken excitatory (AMPAR) or inhibitory (GABAAR) chemical neurotransmission, and two that strengthen these forms of synaptic communication. Across four labs, we tested reagents in the mouse cerebellum, basal ganglia, visual cortex, and retina. Collectively, we demonstrate robust, bidirectional editing of chemical neurotransmission. We provide for distribution of validated reagents, community design principles, and synthetic building blocks for application to diverse pharmaceuticals.

neuroscience↗

bioRxiv2022

Large scale interrogation of retinal cell functions by 1-photon light-sheet microscopy

Visual processing in the retina depends on the collective activity of large ensembles of neurons organized in different layers. Current techniques for measuring activity of layer-specific neural ensembles rely on expensive pulsed infrared lasers to drive 2-photon activation of calcium-dependent fluorescent reporters. Here, we present a 1-photon light-sheet imaging system that can measure the activity in hundreds of ex vivo retinal neurons over a large field of view while simultaneously presenting visual stimuli. This allowed for a reliable functional classification of different retinal ganglion cell types. We also demonstrate that the system has sufficient resolution to image calcium entry at individual synaptic release sites across the axon terminals of dozens of simultaneously imaged bipolar cells. The simple design, a large field of view, and fast image acquisition, make this a powerful system for high-throughput and high-resolution measurements of retinal processing at a fraction of the cost of alternative approaches.

neuroscience↗

bioRxiv2022

Cones and Cone Pathways Remain Functional in Advanced Retinal Degeneration

Most defects causing retinal degeneration in retinitis pigmentosa (RP) are rod-specific mutations, but the subsequent degeneration of cones, which produces loss of daylight vision and high-acuity perception, is the most debilitating feature of the disease. To understand better why cones degenerate and how cone vision might be restored, we have made the first single-cell recordings of light responses from degenerating cones and retinal interneurons after most rods have died and cones have lost their outer-segment disk membranes and synaptic pedicles. We show that degenerating cones have functional cyclic-nucleotide-gated channels and can continue to give light responses, apparently produced by opsin localized either to small areas of organized membrane near the ciliary axoneme or distributed throughout the inner segment. Light responses of second-order horizontal and bipolar cells are smaller and less sensitive but surprisingly similar to those of normal retina. Furthermore, retinal output as reflected in responses of ganglion cells is largely intact at cone-mediated light levels. Together, these findings show that cones and their retinal pathways can remain functional even as degeneration is progressing, an encouraging result for future research aimed at enhancing the light sensitivity of residual cones to restore vision in patients with genetically inherited retinal degeneration.

neuroscience↗

bioRxiv2022

Robust cone-mediated signaling persists late into rod photoreceptor degeneration

Rod photoreceptor degeneration causes deterioration in the morphology and physiology of cone photoreceptors along with changes in retinal circuits. These changes could diminish visual signaling at cone-mediated light levels, thereby limiting the efficacy of treatments such as gene therapy for rescuing normal, cone-mediated vision. However, the impact of progressive rod death on cone-mediated signaling remains unclear. A mouse model of rod degeneration was used to investigate the fidelity of retinal ganglion cell (RGC) signaling throughout disease progression. Despite clear deterioration of cone morphology with rod death, cone-mediated signaling among RGCs remained surprisingly robust: spatiotemporal receptive fields changed little and the mutual information between stimuli and spiking responses was relatively constant. This relative stability held until nearly all rods had died and cones had completely lost well-formed outer segments. Interestingly, RGC information rates were higher and more stable for natural movies than checkerboard noise as degeneration progressed. The main change in RGC responses with photoreceptor degeneration was a decrease in response gain. These results suggest that gene therapies for rod degenerative diseases are likely to successfully prolong cone-mediated vision even if there are changes to cone morphology and density.

neuroscience↗