Secretion of melatonin, a natural hormone whose receptors are present in the ciliary epithelium, displays diurnal variation in the aqueous humor (AH), potentially contributing to the regulation of intraocular pressure. This study aimed to determine the effects of melatonin on AH secretion in porcine ciliary epithelium. The addition of 100 µM melatonin to both sides of the epithelium significantly increased the short-circuit current (Isc) by ~40%. Stromal administration alone had no effect on the Isc, but aqueous application triggered a 40% increase in Isc, similar to that of bilateral application without additive effect. Pre-treatment with niflumic acid abolished melatonin-induced Isc stimulation. More importantly, melatonin stimulated the fluid secretion across the intact ciliary epithelium by ~80% and elicited a sustained increase (~50–60%) in gap junctional permeability between pigmented ciliary epithelial (PE) cells and non-pigmented ciliary epithelial (NPE) cells. The expression of MT3 receptor was found to be >10-fold higher than that of MT1 and MT2 in porcine ciliary epithelium. Aqueous pre-treatment with MT1/MT2 antagonist luzindole failed to inhibit the melatonin-induced Isc response, while MT3 antagonist prazosin pre-treatment abolished the Isc stimulation. We conclude that melatonin facilitates Cl− and fluid movement from PE to NPE cells, thereby stimulating AH secretion via NPE-cell MT3 receptors.
Treatment of many posterior-segment ocular indications would benefit from improved targeting of drug delivery to the back of the eye. Here, we propose the use of iontophoresis to direct delivery of negatively charged nanoparticles through the suprachoroidal space (SCS) toward the posterior pole of the eye. Injection of nanoparticles into the SCS of the rabbit eye ex vivo without iontophoresis led to a nanoparticle distribution mostly localized at the site of injection near the limbus and <15% of nanoparticles delivered to the most posterior region of SCS (>9 mm from the limbus). Iontophoresis using a novel microneedle-based device increased posterior targeting with >30% of nanoparticles in the most posterior region of SCS. Posterior targeting increased with increasing iontophoresis current and increasing application time up to 3 min, but further increasing to 5 min was not better, probably due to the observed collapse of the SCS within 5 min after injection ex vivo. Reversing the direction of iontophoretic flow inhibited posterior targeting, with just ~5% of nanoparticles reaching the most posterior region of SCS. In the rabbit eye in vivo, iontophoresis at 0.14 mA for 3 min after injection of a 100 μL suspension of nanoparticles resulted in ~30% of nanoparticles delivered to the most posterior region of the SCS, which was consistent with ex vivo findings. The procedure was well tolerated, with only mild, transient tissue effects at the site of injection. We conclude that iontophoresis in the SCS using a microneedle has promise as a method to target ocular drug delivery within the eye, especially toward the posterior pole.
Aerobic exercise is a common intervention for rehabilitation of motor, and more recently, cognitive function (Intlekofer and Cotman, 2013; Wood et al., 2012). While the underlying mechanisms are complex, BDNF may mediate much of the beneficial effects of exercise to these neurons (Ploughman et al., 2007; Griffin et al., 2011; Real et al., 2013). We studied the effects of aerobic exercise on retinal neurons undergoing degeneration. We exercised wild-type BALB/c mice on a treadmill (10 m/min for 1 h) for 5 d/week or placed control mice on static treadmills. After 2 weeks of exercise, mice were exposed to either toxic bright light (10,000 lux) for 4 h to induce photoreceptor degeneration or maintenance dim light (25 lux). Bright light caused 75% loss of both retinal function and photoreceptor numbers. However, exercised mice exposed to bright light had 2 times greater retinal function and photoreceptor nuclei than inactive mice exposed to bright light. In addition, exercise increased retinal BDNF protein levels by 20% compared with inactive mice. Systemic injections of a BDNF tropomyosin-receptor-kinase (TrkB) receptor antagonist reduced retinal function and photoreceptor nuclei counts in exercised mice to inactive levels, effectively blocking the protective effects seen with aerobic exercise. The data suggest that aerobic exercise is neuroprotective for retinal degeneration and that this effect is mediated by BDNF signaling.
Diffuse correlation spectroscopy (DCS) is an optical modality used to measure an index of blood flow in biological tissue. This blood flow index depends on both the red blood cell flow rate and density (i.e., hematocrit), although the functional form of hematocrit dependence is not well delineated. Herein, we develop and validate a novel tissue-simulating phantom containing hundreds of microchannels to investigate the influence of hematocrit on blood flow index. For a fixed flow rate, we demonstrate a significant inverse relationship between hematocrit and blood flow index that must be accounted for to accurately estimate blood flow under anemic conditions.
by
Edelle Field-Fote;
Daniel Pinto;
Allen W Heinemann;
Shuo-Hsiu Chang;
Susan Charlifue;
Catherine L Furbish;
Arun Jayaraman;
Candace Tefertiller;
Heather B Taylor;
Dustin D French
Background: Few, if any estimates of cost-effectiveness for locomotor training strategies following spinal cord injury (SCI) are available. The purpose of this study was to estimate the cost-effectiveness of locomotor training strategies following spinal cord injury (overground robotic locomotor training versus conventional locomotor training) by injury status (complete versus incomplete) using a practice-based cohort. Methods: A probabilistic cost-effectiveness analysis was conducted using a prospective, practice-based cohort from four participating Spinal Cord Injury Model System sites. Conventional locomotor training strategies (conventional training) were compared to overground robotic locomotor training (overground robotic training). Conventional locomotor training included treadmill-based training with body weight support, overground training, and stationary robotic systems. The outcome measures included the calculation of quality adjusted life years (QALYs) using the EQ-5D and therapy costs. We estimate cost-effectiveness using the incremental cost utility ratio and present results on the cost-effectiveness plane and on cost-effectiveness acceptability curves. Results: Participants in the prospective, practice-based cohort with complete EQ-5D data (n = 99) qualified for the analysis. Both conventional training and overground robotic training experienced an improvement in QALYs. Only people with incomplete SCI improved with conventional locomotor training, 0.045 (SD 0.28), and only people with complete SCI improved with overground robotic training, 0.097 (SD 0.20). Costs were lower for conventional training, $1758 (SD $1697) versus overground robotic training $3952 (SD $3989), and lower for those with incomplete versus complete injury. Conventional overground training was more effective and cost less than robotic therapy for people with incomplete SCI. Overground robotic training was more effective and cost more than conventional training for people with complete SCI. The incremental cost utility ratio for overground robotic training for people with complete spinal cord injury was $12,353/QALY. Conclusions: The most cost-effective locomotor training strategy for people with SCI differed based on injury completeness. Conventional training was more cost-effective than overground robotic training for people with incomplete SCI. Overground robotic training was more cost-effective than conventional training for people with complete SCI. The effect estimates may be subject to limitations associated with small sample sizes and practice-based evidence methodology. These estimates provide a baseline for future research.
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Joshua A Chu-Tan;
Adrian V Cioanca;
Yvette Wooff;
Max Kirkby;
Marissa Ellis;
Pranay Gulati;
Tim Karl;
Jeffrey Boatright;
Katie Bales;
John Nickerson;
Riccardo Natoli
Background: Exercise has been shown to promote a healthier and longer life and linked to a reduced risk of developing neurodegenerative diseases including retinal degenerations. However, the molecular pathways underpinning exercise-induced cellular protection are not well understood. In this work we aim to profile the molecular changes underlying exercise-induced retinal protection and investigate how exercise-induced inflammatory pathway modulation may slow the progression of retinal degenerations. Methods: Female C57Bl/6J mice at 6 weeks old were given free access to open voluntary running wheels for a period of 28 days and then subjected to 5 days of photo-oxidative damage (PD)-induced retinal degeneration. Following, retinal function (electroretinography; ERG), morphology (optical coherence tomography; OCT) and measures of cell death (TUNEL) and inflammation (IBA1) were analysed and compared to sedentary controls. To decipher global gene expression changes as a result of voluntary exercise, RNA sequencing and pathway and modular gene co-expression analyses were performed on retinal lysates of exercised and sedentary mice that were subjected to PD, as well as healthy dim-reared controls. Results: Following 5 days of PD, exercised mice had significantly preserved retinal function, integrity and reduced levels of retinal cell death and inflammation, compared to sedentary controls. In response to voluntary exercise, inflammatory and extracellular matrix integrity pathways were significantly modulated, with the gene expression profile of exercised mice more closely trending towards that of a healthy dim-reared retina. Conclusion: We suggest that voluntary exercise may mediate retinal protection by influencing key pathways involved in regulating retinal health and shifting the transcriptomic profile to a healthy phenotype.
Although effective drugs that lower intraocular pressure (IOP) in the management of glaucoma exist, their efficacy is limited by poor patient adherence to the prescribed eye drop regimen. To replace the need for eye drops, in this study we tested the hypothesis that IOP can be reduced for one month after a single targeted injection using a microneedle for administration of a glaucoma medication (i.e., brimonidine) formulated for sustained release in the supraciliary space of the eye adjacent to the drug's site of action at the ciliary body. To test this hypothesis, brimonidine-loaded microspheres were formulated using poly(lactic acid) (PLA) to release brimonidine at a constant rate for 35 days and microneedles were designed to penetrate through the sclera, without penetrating into the choroid/retina, in order to target injection into the supraciliary space. A single administration of these microspheres using a hollow microneedle was performed in the eye of New Zealand White rabbits and was found to reduce IOP initially by 6 mm Hg and then by progressively smaller amounts for more than one month. All administrations were well tolerated without significant adverse events, although histological examination showed a foreign-body reaction to the microspheres. This study demonstrates, for the first time, that the highly-targeted delivery of brimonidine-loaded microspheres into the supraciliary space using a microneedle is able to reduce IOP for one month as an alternative to daily eye drops.
We introduce Airy-beam tomographic microscopy (ATM) for high-resolution, volumetric, inertia-free imaging of biological specimens. The work exploits the highly adjustable Airy trajectories in the 3D space, transforming the conventional telecentric wide-field imaging scheme that requires sample or focal-plane scanning to acquire 3D information. The results present a consistent near-diffraction-limited 3D resolution across a tenfold extended imaging depth compared to widefield microscopy. We anticipate the strategy to not only offer a promising paradigm for 3D optical microscopy, but also be translated to other non-optical waveforms.
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Rachael S Allen;
Adam M Hanif;
Marissa A Gogniat;
Brian C Prall;
Raza Haider;
Moe H Aung;
Megan C Prunty;
Lukas M Mees;
Monica M Coulter;
Cara T Motz;
Jeffrey Boatright;
Machelle Pardue
Diabetic retinopathy is a leading cause of vision loss. Treatment options for early retinopathy are sparse. Exercise protects dying photoreceptors in models of retinal degeneration, thereby preserving vision. We tested the protective effects of exercise on retinal and cognitive deficits in a type 1 diabetes model and determined whether the TrkB pathway mediates this effect. Hyperglycaemia was induced in Long Evans rats via streptozotocin injection (STZ; 100 mg/kg). Following confirmed hyperglycaemia, both control and diabetic rats underwent treadmill exercise for 30 min, 5 days/week at 0 m/min (inactive groups) or 15 m/min (active groups) for 8 weeks. A TrkB receptor antagonist (ANA-12), or vehicle, was injected 2.5 h before exercise training. We measured spatial frequency and contrast sensitivity using optokinetic tracking biweekly post-STZ; retinal function using electroretinography at 4 and 8 weeks; and cognitive function and exploratory behaviour using Y-maze at 8 weeks. Retinal neurotrophin-4 was measured using ELISA. Compared with non-diabetic controls, diabetic rats showed significantly reduced spatial frequency and contrast sensitivity, delayed electroretinogram oscillatory potential and flicker implicit times and reduced cognitive function and exploratory behaviour. Exercise interventions significantly delayed the appearance of all deficits, except for exploratory behaviour. Treatment with ANA-12 significantly reduced this protection, suggesting a TrkB-mediated mechanism. Despite this, no changes in retinal neurotrohin-4 were observed with diabetes or exercise. Exercise protected against early visual and cognitive dysfunction in diabetic rats, suggesting that exercise interventions started after hyperglycaemia diagnosis may be a beneficial treatment. The translational potential is high, given that exercise treatment is non-invasive, patient controlled and inexpensive.
The diurnal peak of phagocytosis by the retinal pigment epithelium (RPE) of photoreceptor outer segments (POS) is under circadian control and believed that this process involves interactions from the retina and RPE. Previous studies have demonstrated that a functional circadian clock exists within multiple retinal cell types and RPE. Thereby, the aim of this study was to determine whether the clock in the retina or RPE controls the diurnal phagocytic peak and whether disruption of the circadian clock in the RPE would affect cellular function and the viability during aging. To that, we generated and validated an RPE tissue-specific KO of the essential clock gene, Bmal1, and then determined the daily rhythm in phagocytic activity by the RPE in mice lacking a functional circadian clock in the retina or RPE.
Then, using electroretinography, spectral domain-optical coherence tomography, and optomotor response of visual function we determined the effect of Bmal1 removal in young (6 months) and old (18 months) mice. RPE morphology and lipofuscin accumulation was determined in young and old mice. Our data shows that the clock in the RPE, rather than the retina clock, controls the diurnal phagocytic peak. Surprisingly, absence of a functional RPE clock and phagocytic peak does not result in any detectable age-related degenerative phenotype in the retina or RPE. Thus, our results demonstrate that the circadian clock in the RPE controls the daily peak of phagocytic activity. However, the absence of the clock in the RPE does not result in deterioration of photoreceptors or the RPE during aging.