BackgroundUncontrolled microglial activation contributes to the pathogenesis of various neurodegenerative diseases. Previous studies have shown that proinflammatory microglia are powered by glycolysis, which relays on high levels of glucose uptake. This study aimed to understand how glucose uptake is facilitated in active microglia and whether microglial activation can be controlled by restricting glucose uptake.MethodsPrimary murine brain microglia, BV2 cells and the newly established microglial cell line B6M7 were treated with LPS (100 ng/ml) + IFNγ (100 ng/ml) or IL-4 (20 ng/ml) for 24 h. The expression of glucose transporters (GLUTs) was examined by PCR and Western blot. Glucose uptake by microglia was inhibited using the GLUT1-specific inhibitor STF31. The metabolic profiles were tested using the Glycolysis Stress Test and Mito Stress Test Kits using the Seahorse XFe96 Analyser. Inflammatory gene expression was examined by real-time RT-PCR and protein secretion by cytokine beads array. The effect of STF31 on microglial activation and neurodegeneraion was further tested in a mouse model of light-induced retinal degeneration.ResultsThe mRNA and protein of GLUT1, 3, 4, 5, 6, 8, 9, 10, 12, and 13 were detected in microglia. The expression level of GLUT1 was the highest among all GLUTs detected. LPS + IFNγ treatment further increased GLUT1 expression. STF31 dose-dependently reduced glucose uptake and suppressed Extracellular Acidification Rate (ECAR) in naïve, M(LPS + IFNγ) and M(IL-4) microglia. The treatment also prevented the upregulation of inflammatory cytokines including TNFα, IL-1β, IL-6, and CCL2 in M(LPS + IFNγ) microglia. Interestingly, the Oxygen Consumption Rates (OCR) was increased in M(LPS + IFNγ) microglia but reduced in M(IL-4) microglia by STF31 treatment. Intraperitoneal injection of STF31 reduced light-induced microglial activation and retinal degeneration.ConclusionGlucose uptake in microglia is facilitated predominately by GLUT1, particularly under inflammatory conditions. Targeting GLUT1 could be an effective approach to control neuroinflammation.Electronic supplementary materialThe online version of this article (10.1186/s13024-019-0305-9) contains supplementary material, which is available to authorized users.

High-resolution visual prostheses require small, densely packed pixels, but limited penetration depth of the electric field formed by a planar electrode array constrains such miniaturization. We present a novel honeycomb configuration of an electrode array with vertically separated active and return electrodes designed to leverage migration of retinal cells into voids in the subretinal space. Insulating walls surrounding each pixel decouple the field penetration depth from the pixel width by aligning the electric field vertically, enabling a decrease of the pixel size down to cellular dimensions. We demonstrate that inner retinal cells migrate into the 25 μm deep honeycomb wells as narrow as 18 μm, resulting in more than half of these cells residing within the electrode cavities. Immune response to honeycombs is comparable to that with planar arrays. Modeled stimulation threshold current density with honeycombs does not increase substantially with reduced pixel size, unlike quadratic increase with planar arrays. This 3-D electrode configuration may enable functional restoration of central vision with acuity better than 20/100 for millions of patients suffering from age-related macular degeneration.

BackgroundReactive microglia are commonly seen in retinal degenerative diseases, and neurotoxic microglia responses can contribute to photoreceptor cell death. We and others have previously shown that translocator protein (18 kDa) (TSPO) is highly induced in retinal degenerations and that the selective TSPO ligand XBD173 (AC-5216, emapunil) exerts strong anti-inflammatory effects on microglia in vitro and ex vivo. Here, we investigated whether targeting TSPO with XBD173 has immuno-modulatory and neuroprotective functions in two mouse models of acute retinal degeneration using bright white light exposure.MethodsBALB/cJ and Cx3cr1GFP/+ mice received intraperitoneal injections of 10 mg/kg XBD173 or vehicle for five consecutive days, starting 1 day prior to exposure to either 15,000 lux white light for 1 h or 50,000 lux focal light for 10 min, respectively. The effects of XBD173 treatment on microglia and Müller cell reactivity were analyzed by immuno-stainings of retinal sections and flat mounts, fluorescence-activated cell sorting (FACS) analysis, and mRNA expression of microglia markers using quantitative real-time PCR (qRT-PCR). Optical coherence tomography (OCT), terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) stainings, and morphometric analyses were used to quantify the extent of retinal degeneration and photoreceptor apoptosis.ResultsFour days after the mice were challenged with bright white light, a large number of amoeboid-shaped alerted microglia appeared in the degenerating outer retina, which was nearly completely prevented by treatment with XBD173. This treatment also down-regulated the expression of TSPO protein in microglia but did not change the TSPO levels in the retinal pigment epithelium (RPE). RT-PCR analysis showed that the microglia/macrophage markers Cd68 and activated microglia/macrophage whey acidic protein (Amwap) as well as the pro-inflammatory genes Ccl2 and Il6 were reduced after XBD173 treatment. Light-induced degeneration of the outer retina was nearly fully blocked by XBD173 treatment. We further confirmed these findings in an independent mouse model of focal light damage. Retinas of animals receiving XBD173 therapy displayed significantly more ramified non-reactive microglia and more viable arrestin-positive cone photoreceptors than vehicle controls.ConclusionsTargeting TSPO with XBD173 effectively counter-regulates microgliosis and ameliorates light-induced retinal damage, highlighting a new pharmacological concept for the treatment of retinal degenerations.

Objective. To restore central vision in patients with atrophic age-related macular degeneration, we replace the lost photoreceptors with photovoltaic pixels, which convert light into current and stimulate the secondary retinal neurons. Clinical trials demonstrated prosthetic acuity closely matching the sampling limit of the 100 μm pixels, and hence smaller pixels are required for improving visual acuity. However, with smaller flat bipolar pixels, the electric field penetration depth and the photodiode responsivity significantly decrease, making the device inefficient. Smaller pixels may be enabled by (a) increasing the diode responsivity using vertical p–n junctions and (b) directing the electric field in tissue vertically. Here, we demonstrate such novel photodiodes and test the retinal stimulation in a vertical electric field. Approach. Arrays of silicon photodiodes of 55, 40, 30, and 20 μm in width, with vertical p–n junctions, were fabricated. The electric field in the retina was directed vertically using a common return electrode at the edge of the device. Optical and electronic performance of the diodes was characterized in-vitro, and retinal stimulation threshold measured by recording the visually evoked potentials in rats with retinal degeneration. Main results. The photodiodes exhibited sufficiently low dark current (<10 pA) and responsivity at 880 nm wavelength as high as 0.51 A W−1, with 85% internal quantum efficiency, independent of pixel size. Field mapping in saline demonstrated uniformity of the pixel performance in the array. The full-field stimulation threshold was as low as 0.057 ± 0.029 mW mm−2 with 10 ms pulses, independent of pixel size. Significance. Photodiodes with vertical p–n junctions demonstrated excellent charge collection efficiency independent of pixel size, down to 20 μm. Vertically oriented electric field provides a stimulation threshold that is independent of pixel size. These results are the first steps in validation of scaling down the photovoltaic pixels for subretinal stimulation.

Localized stimulation of the inner retinal neurons for high-acuity prosthetic vision requires small pixels and minimal crosstalk from the neighboring electrodes. Local return electrodes within each pixel limit the crosstalk, but they over-constrain the electric field, thus precluding the efficient stimulation with subretinal pixels smaller than 55 μm. Here we demonstrate a high-resolution prosthetic vision based on a novel design of a photovoltaic array, where field confinement is achieved dynamically, leveraging the adjustable conductivity of the diodes under forward bias to turn the designated pixels into transient returns. We validated the computational modeling of the field confinement in such an optically-controlled circuit by in-vitro and in-vivo measurements. Most importantly, using this strategy, we demonstrated that the grating acuity with 40 μm pixels matches the pixel pitch, while with 20 μm pixels, it reaches the 28 μm limit of the natural visual resolution in rats. This method enables customized field shaping based on individual retinal thickness and distance from the implant, paving the way to higher acuity of prosthetic vision in atrophic macular degeneration.