Topically instilled caffeine selectively alters emmetropizing responses in infant rhesus monkeys.

Oral administration of the adenosine receptor (ADOR) antagonist, 7-methylxanthine (7-MX), reduces both form-deprivation and lens-induced myopia in mammalian animal models. We investigated whether topically instilled caffeine, another non-selective ADOR antagonist, retards vision-induced axial elongation in monkeys. Beginning at 24 days of age, a 1.4% caffeine solution was instilled in both eyes of 14 rhesus monkeys twice each day until the age of 135 days. Concurrent with the caffeine regimen, the monkeys were fitted with helmets that held either -3 D (-3D/pl caffeine, n = 8) or +3 D spectacle lenses (+3D/pl caffeine, n = 6) in front of their lens-treated eyes and zero-powered lenses in front of their fellow-control eyes. Refractive errors and ocular dimensions were measured at baseline and periodically throughout the lens-rearing period. Control data were obtained from 8 vehicle-treated animals also reared with monocular -3 D spectacles (-3D/pl vehicle). In addition, historical comparison data were available for otherwise untreated lens-reared controls (-3D/pl controls, n = 20; +3D/pl controls, n = 9) and 41 normal monkeys. The vehicle controls and the untreated lens-reared controls consistently developed compensating axial anisometropias (-3D/pl vehicle = -1.44 ± 1.04 D; -3D/pl controls = -1.85 ± 1.20 D; +3D/pl controls = +1.92 ± 0.56 D). The caffeine regime did not interfere with hyperopic compensation in response to +3 D of anisometropia (+1.93 ± 0.82 D), however, it reduced the likelihood that animals would compensate for -3 D of anisometropia (+0.58 ± 1.82 D). The caffeine regimen also promoted hyperopic shifts in both the lens-treated and fellow-control eyes; 26 of the 28 caffeine-treated eyes became more hyperopic than the median normal monkey (mean (±SD) relative hyperopia = +2.27 ± 1.65 D; range = +0.31 to +6.37 D). The effects of topical caffeine on refractive development, which were qualitatively similar to those produced by oral administration of 7-MX, indicate that ADOR antagonists have potential in treatment strategies for preventing and/or reducing myopia progression.

Adenosine receptor distribution in Rhesus monkey ocular tissue.

Adenosine receptor (ADOR) antagonists, such as 7-methylxanthine (7-MX), have been shown to slow myopia progression in humans and animal models. Adenosine receptors are found throughout the body, and regulate the release of neurotransmitters such as dopamine and glutamate. However, the role of adenosine in eye growth is unclear. Evidence suggests that 7-MX increases scleral collagen fibril diameter, hence preventing axial elongation. This study used immunohistochemistry (IHC) and reverse-transcription quantitative polymerase chain reaction (RT-qPCR) to examine the distribution of the four ADORs in the normal monkey eye to help elucidate potential mechanisms of action. Eyes were enucleated from six Rhesus monkeys. Anterior segments and eyecups were separated into components and flash-frozen for RNA extraction or fixed in 4% paraformaldehyde and processed for immunohistochemistry against ADORA1, ADORA2a, ADORA2b, and ADORA3. RNA was reverse-transcribed, and qPCR was performed using custom primers. Relative gene expression was calculated using the ΔΔCt method normalizing to liver expression, and statistical analysis was performed using Relative Expression Software Tool. ADORA1 immunostaining was highest in the iris sphincter muscle, trabecular meshwork, ciliary epithelium, and retinal nerve fiber layer. ADORA2a immunostaining was highest in the corneal epithelium, trabecular meshwork, ciliary epithelium, retinal nerve fiber layer, and scleral fibroblasts. ADORA2b immunostaining was highest in corneal basal epithelium, limbal stem cells, iris sphincter, ciliary muscle, ciliary epithelium, choroid, isolated retinal ganglion cells and scattered scleral fibroblasts. ADORA3 immunostaining was highest in the iris sphincter, ciliary muscle, ciliary epithelium, choroid, isolated retinal ganglion cells, and scleral fibroblasts. Compared to liver mRNA, ADORA1 mRNA was significantly higher in the brain, retina and choroid, and significantly lower in the iris/ciliary body. ADORA2a expression was higher in brain and retina, ADORA2b expression was higher in retina, and ADORA3 was higher in the choroid. In conclusion, immunohistochemistry and RT-qPCR indicated differential patterns of expression of the four adenosine receptors in the ocular tissues of the normal non-human primate. The presence of ADORs in scleral fibroblasts and the choroid may support mechanisms by which ADOR antagonists prevent myopia. The potential effects of ADOR inhibition on both anterior and posterior ocular structures warrant investigation.

The Adenosine Receptor Antagonist, 7-Methylxanthine, Alters Emmetropizing Responses in Infant Macaques.

Purpose: Previous studies suggest that the adenosine receptor antagonist, 7-methylxanthine (7-MX), retards myopia progression. Our aim was to determine whether 7-MX alters the compensating refractive changes produced by defocus in rhesus monkeys. Methods: Starting at age 3 weeks, monkeys were reared with -3 diopter (D; n = 10; 7-MX -3D/pl) or +3D (n = 6; 7-MX +3D/pl) spectacles over their treated eyes and zero-powered lenses over their fellow eyes. In addition, they were given 100 mg/kg of 7-MX orally twice daily throughout the lens-rearing period (age 147 ± 4 days). Comparison data were obtained from lens-reared controls (-3D/pl, n = 17; +3D/pl, n = 9) and normal monkeys (n = 37) maintained on a standard diet. Refractive status, corneal power, and axial dimensions were assessed biweekly. Results: The -3D/pl and +3D/pl lens-reared controls developed compensating myopic (-2.10 ± 1.07 D) and hyperopic anisometropias (+1.86 ± 0.54 D), respectively. While the 7-MX +3D/pl monkeys developed hyperopic anisometropias (+1.79 ± 1.11 D) that were similar to those observed in +3D/pl controls, the 7-MX -3D/pl animals did not consistently exhibit compensating myopia in their treated eyes and were on average isometropic (+0.35 ± 1.96 D). The median refractive errors for both eyes of the 7-MX -3D/pl (+5.47 D and +4.38 D) and 7-MX +3D/pl (+5.28 and +3.84 D) monkeys were significantly more hyperopic than that for normal monkeys (+2.47 D). These 7-MX-induced hyperopic ametropias were associated with shorter vitreous chambers and thicker choroids. Conclusions: In primates, 7-MX reduced the axial myopia produced by hyperopic defocus, augmented hyperopic shifts in response to myopic defocus, and induced hyperopia in control eyes. The results suggest that 7-MX has therapeutic potential in efforts to slow myopia progression.

Protective effects of high ambient lighting on the development of form-deprivation myopia in rhesus monkeys.

PURPOSE: Time spent outdoors reduces the likelihood that children will develop myopia, possibly because light levels are much higher outdoors than indoors. To test this hypothesis, the effects of high ambient lighting on vision-induced myopia in monkeys were determined. METHODS: Monocular form deprivation was imposed on eight infant rhesus monkeys. Throughout the rearing period (23 ± 2 to 132 ± 8 days), auxiliary lighting increased the cage-level illuminance from normal lighting levels (15-630 lux) to ∼25,000 lux for 6 hours during the middle of the daily 12-hour light cycle. Refractive development and axial dimensions were assessed by retinoscopy and ultrasonography, respectively. Comparison data were obtained in previous studies from 18 monocularly form-deprived and 32 normal monkeys reared under ordinary laboratory lighting. RESULTS: Form deprivation produced axial myopia in 16 of 18 normal-light-reared monkeys. In contrast, only 2 of the 8 high-light-reared monkeys developed myopic anisometropias, and in 6 of these monkeys, the form-deprived eyes were more hyperopic than their fellow eyes. The treated eyes of the high-light-reared monkeys were more hyperopic than the form-deprived eyes of the normal-light-reared monkeys. In addition, both eyes of the high-light-reared monkeys were more hyperopic than those of normal monkeys. CONCLUSIONS: High ambient lighting retards the development of form-deprivation myopia in monkeys. These results are in agreement with the hypothesis that the protective effects of outdoor activities against myopia in children are due to exposure to the higher light levels encountered outdoors. It is possible that therapeutic protection against myopia can be achieved by manipulating indoor lighting levels.

Effects of foveal ablation on emmetropization and form-deprivation myopia.

PURPOSE: Because of the prominence of central vision in primates, it has generally been assumed that signals from the fovea dominate refractive development. To test this assumption, the authors determined whether an intact fovea was essential for either normal emmetropization or the vision-induced myopic errors produced by form deprivation. METHODS: In 13 rhesus monkeys at 3 weeks of age, the fovea and most of the perifovea in one eye were ablated by laser photocoagulation. Five of these animals were subsequently allowed unrestricted vision. For the other eight monkeys with foveal ablations, a diffuser lens was secured in front of the treated eyes to produce form deprivation. Refractive development was assessed along the pupillary axis by retinoscopy, keratometry, and A-scan ultrasonography. Control data were obtained from 21 normal monkeys and three infants reared with plano lenses in front of both eyes. RESULTS: Foveal ablations had no apparent effect on emmetropization. Refractive errors for both eyes of the treated infants allowed unrestricted vision were within the control range throughout the observation period, and there were no systematic interocular differences in refractive error or axial length. In addition, foveal ablation did not prevent form deprivation myopia; six of the eight infants that experienced monocular form deprivation developed myopic axial anisometropias outside the control range. CONCLUSIONS: Visual signals from the fovea are not essential for normal refractive development or the vision-induced alterations in ocular growth produced by form deprivation. Conversely, the peripheral retina, in isolation, can regulate emmetropizing responses and produce anomalous refractive errors in response to abnormal visual experience. These results indicate that peripheral vision should be considered when assessing the effects of visual experience on refractive development.

Recovery from form-deprivation myopia in rhesus monkeys.

PURPOSE: Although many aspects of vision-dependent eye growth are qualitatively similar in many species, the failure to observe recovery from form-deprivation myopia (FDM) in higher primates represents a significant potential departure. The purpose of this investigation was to re-examine the ability of rhesus monkeys (Macaca mulatta) to recover from FDM. METHODS: Monocular form deprivation was produced either with diffuser spectacle lenses (n = 30) or by surgical eyelid closure (n = 14). The diffuser-rearing strategies were initiated at 24 +/- 3 days of age and continued for an average of 115 +/- 20 days. Surgical eyelid closure was initiated between 33 and 761 days of age and maintained for14 to 689 days. After the period of form deprivation, the animals were allowed unrestricted vision. The ability of the animals to recover from treatment-induced refractive errors was assessed periodically by retinoscopy, keratometry, and A-scan ultrasonography. Control data were obtained from 35 normal monkeys. RESULTS: At the onset of unrestricted vision, the deprived eyes of 18 of the diffuser-reared monkeys and 12 of the lid-sutured monkeys were at least 1.0 D less hyperopic or more myopic than their fellow eyes. The mean (diffuser = -4.06 D, lid-suture = -4.50 D) and range (diffuser = -1.0 to -10.19 D, lid-suture = -1.0 to -10.25 D) of myopic anisometropia were comparable in both treatment groups. All 18 of these diffuser-reared monkeys demonstrated recovery, with 12 animals exhibiting complete recovery. The rate of recovery, which was mediated primarily by alterations in vitreous chamber growth rate, declined with age. None of the lid-sutured monkeys exhibited clear evidence of recovery. Instead, 8 of the 12 lid-sutured monkeys exhibited progression of myopia. CONCLUSIONS: Like many other species, young monkeys are capable of recovering from FDM. However, the potential for recovery appears to depend on when unrestricted vision is restored, the severity of the deprivation-induced axial elongation, and possibly the method used to produce FDM.

Glutathione content is altered in Müller cells of monkey eyes with experimental glaucoma.

Extracellular levels of glutamate are thought to be increased in glaucoma and thus contribute to retinal damage. An increase in glutamate concentration or duration in the extracellular retinal space is expected to impact glutathione content in Müller cells since glutamate is the rate-limiting constituent in glutathione synthesis. We have investigated whether glutathione content is changed in retinal Müller cells of monkeys with experimental glaucoma using immunocytochemistry and image analysis. Müller cells in glaucomatous retinas showed significantly greater immunoreactivity (27-57%) for glutathione than those in fellow control retinas, increasing with the duration of elevated intraocular pressure (IOP). This pattern of labeling was prominent in all four monkeys studied. The presence of more glutathione in Müller cells of glaucomatous retinas is consistent with an increase in extracellular glutamate and an increase in transport and metabolism of glutamate.

Effects of brief periods of unrestricted vision on the development of form-deprivation myopia in monkeys.

PURPOSE: To characterize the temporal integration properties of the mechanisms responsible for form-deprivation myopia (FDM), the effects of brief daily periods of unrestricted vision on the degree of FDM were investigated in infant monkeys. METHODS: Starting at approximately 3 weeks of age, unilateral form deprivation was produced in 24 infant rhesus monkeys by securing a diffuser spectacle lens in front of one eye and a clear, zero-powered lens in front of the fellow eye. During the treatment period (17 +/- 2 weeks), six infants wore the diffuser lenses continuously. In the other experimental infants, the diffuser lenses were removed each day and replaced with clear, zero-powered lenses for 1 (n = 7), 2 (n = 7), or 4 hours (n = 4). Refractive development was assessed by retinoscopy and A-scan ultrasonography. Control data were obtained from 11 normal infants and 3 infants reared with zero-powered lenses over both eyes. RESULTS: The degree of FDM varied significantly with the duration of unrestricted vision. Continuous form deprivation produced -5.2 +/- 3.6 D of relative axial myopia. However, 1 hour of unrestricted vision was sufficient to reduce the degree of axial FDM by more than 50% (-1.7 +/- 3.2 D). The infants that were allowed 4 hours of unrestricted vision exhibited only -0.4 +/- 0.5 D of FDM. CONCLUSIONS: As observed in chickens and tree shrews, relatively long periods of form deprivation can be counterbalanced by quite short periods of unrestricted vision. These results indicate that the processes or signals that promote axial elongation in monkeys are comparatively weak or easily overridden by factors that slow ocular growth.