Effects of Hydrostatic Pressure on Electrical Retinal Activity in a Multielectrode Array-Based ex vivo Glaucoma Acute Model.

Glaucoma is a heterogeneous eye disease causing atrophy of the optic nerve head (ONH). The optic nerve is formed by the axons of the retinal ganglion cells (RGCs) that transmit visual input to the brain. The progressive RGC loss during glaucoma leads to irreversible vision loss. An elevated intraocular pressure (IOP) is described as main risk factor in glaucoma. In this study, a multielectrode array (MEA)-based ex vivo glaucoma acute model was established and the effects of hydrostatic pressure (10, 30, 60, and 90 mmHg) on the functionality and survival of adult male and female wild-type mouse (C57BL/6) retinae were investigated. Spontaneous activity, response rate to electrical and light stimulation, and bursting behavior of RGCs was analyzed prior, during, and after pressure stress. No pressure related effects on spontaneous firing and on the response rate of the RGCs were observed. Even a high pressure level (90 mmHg for 2 h) did not disturb the RGC functionality. However, the cells' bursting behavior significantly changed under 90 mmHg. The number of spikes in bursts doubled during pressure application and stayed on a high level after pressure stress. Addition of the amino sulfonic acid taurine (1 mM) showed a counteracting effect. OFF ganglion cells did not reveal an increase in bursts under pressure stress. Live/dead staining after pressure application showed no significant changes in RGC survival. The findings of our ex vivo model suggest that RGCs are tolerant toward high, short-time pressure stress.

New epiretinal implant with integrated sensor chips for optical capturing shows a good biocompatibility profile in vitro and in vivo.

BACKGROUND: Retinal degenerative diseases, e.g., retinitis pigmentosa, cause a severe decline of the visual function up to blindness. Treatment still remains difficult; however, implantation of retinal prostheses can help restoring vision. In this study, the biocompatibility and surgical feasibility of a newly developed epiretinal stimulator (OPTO-EPIRET) was investigated. The previously developed implant was extended by an integrated circuit-based optical capturing, which will enable the immediate conversion of the visual field into stimulation patterns to stimulate retinal ganglion cells. RESULTS: The biocompatibility of the OPTO-EPIRET was investigated in vitro using the two different cell lines L-929 and R28. Direct and indirect contact were analyzed in terms of cell proliferation, cell viability, and gene expression. The surgical feasibility was initially tested by implanting the OPTO-EPIRET in cadaveric rabbit eyes. Afterwards, inactive devices were implanted in six rabbits for feasibility and biocompatibility testings in vivo. In follow-up controls (1-12 weeks post-surgery), the eyes were examined using fundoscopy and optical coherence tomography. After finalization, histological examination was performed to analyze the retinal structure. Regarding the in vitro biocompatibility, no significant influence on cell viability was detected (L929: < 1.3% dead cells; R-28: < 0.8% dead cells). The surgery, which comprised phacoemulsification, vitrectomy, and implantation of the OPTO-EPIRET through a 9-10 mm corneal incision, was successfully established. The implant was fixated with a retinal tack. Vitreal hemorrhage or retinal tearing occurred as main adverse effects. Transitional corneal edema caused difficulties in post-surgical imaging. CONCLUSIONS: The OPTO-EPIRET stimulator showed a good biocompatibility profile in vitro. Furthermore, the implantation surgery was shown to be feasible. However, further design optimization steps are necessary to avoid intra- and postoperative complications. Overall, the OPTO-EPIRET will allow for a wide visual field and good visual acuity due to a high density of electrodes in the central retina.

A multielectrode array-based hypoxia model for the analysis of electrical activity in murine retinae.

Several eye diseases, for example, retinal artery occlusion, diabetic retinopathy, and glaucoma, are associated with retinal hypoxia. The lack of oxygen in the retina, especially in retinal ganglion cells (RGCs), causes cell damage up to cell degeneration and leads to blindness. Using multielectrode array recordings, an ex vivo hypoxia acute model was established to analyze the electrical activity of murine wild-type retinae under hypoxic stress conditions. Hypoxia was induced by exchanging the perfusion with oxygen-saturated medium by nitrogen-saturated medium. Hypoxic periods of 0 min (control) up to 60 min were tested on the retinae of adult female C57BL/6J mice. The electrical RGC activity vanished during hypoxia, but conditionally returned after the reestablishment of conventional test conditions. With increasing duration of hypoxia, the returning RGC activity decreased. After a hypoxic period of 30 min and a subsequent recovery time of 30 min, 59.43 ± 11.35% of the initially active channels showed a restored RGC activity. The survival rate of retinal cells after hypoxic stress was analyzed by a live/dead staining assay using two-photon laser scanning microscopy. For detailed information about molecular changes caused by hypoxia, a microarray gene expression analysis was performed. Furthermore, the effect of 2-aminoethanesulfonic acid (taurine, 1 mM) on retinae under hypoxic stress was tested. Treatment with taurine resulted in an increase in the RGC response rate after hypoxia and also increased the survival rate of retinal cells under hypoxic stress, confirming its potential as promising candidate for neuroprotective therapies of eye diseases.

A new microfluidic device design for a defined positioning of neurons in vitro.

A new triangle-shaped microfluidic channel system for defined cell trapping is presented. Different variants of the same basic geometry were produced to reveal the best fitting parameter combinations regarding efficiency and sensitivity. Variants with differences in the trap gap width and the inter-trap distance were analyzed in detail by Computational Fluid Dynamics simulations and in experiments with artificial beads of different sizes (30, 60, 80 µm). Simulation analysis of flow dynamics and pressure profiles revealed strongly reduced pressure conditions and balanced flow rates inside the microfluidic channels compared to commonly used systems with meandering channels. Quantitative experiments with beads showed very good trapping results in all channel types with slight variations due to geometrical differences. Highest efficiency in terms of fast trap filling and low particle loss was shown with channel types having a larger trap gap width (20 µm) and/or a larger inter-trap distance (400 µm). Here, experimental success was achieved in almost 85% to 100% of all cases. Particle loss appeared significantly more often with large beads than with small beads. A significantly reduced trapping efficiency of about 50% was determined by using narrow trap gaps and a small inter-trap distance in combination with large 80 µm beads. The combination of the same parameters with small and medium beads led to an only slight decrease in trapping efficiency (80%). All channel types were tested qualitatively with invertebrate neurons from the pond snail Lymnaea stagnalis. The systems were appropriate to trap those sensitive neurons and to keep their viability in the trapping area at the same time.