Purpose

To assess the validity of and compare applanation and rebound tonometry readings of intraocular pressure in alert normal chicks from ages 3 to 45 days.

Methods

Intraocular pressures (IOPs) were measured weekly in awake White Leghorn chicks, from ages 3-45 days (n = 22-30 per age group), with both applanation Tono-Pen and rebound TonoLab tonometers. Three repeated measurements on individual eyes were used to derive variance data for both instruments at each age. Calibration curves were also derived for each instrument and each age, weekly from ages 10-45 days (n = 3-4 per age group), from in situ manometry data collected over IOP settings of 0 to 100 mmHg in 5 mmHg steps in cannulated eyes.

Results

The TonoLab showed less within measurement variability, but more variability with age, than the Tono-Pen. The coefficient of variation ranged from 3.8-8.3% for the TonoLab, compared to 11.0-19.7% for the Tono-Pen across all ages. For the youngest, 3 day-old chicks, mean IOPs recorded with the Tono-Pen and TonoLab were not significantly different (17.0 ± 5.6 and 15.2 ± 3.7 mmHg, respectively, P = .27). However, with increasing age, IOP readings significantly increased for the TonoLab (P < .001), whereas Tono-Pen readings did not. Compared to manometry settings, the Tono-Pen tended to underestimate IOPs while the TonoLab overestimated IOPs over the range 20-60 mmHg, saturating thereafter; there were also age-dependent differences for the TonoLab.

Conclusions

Both the Tono-Pen and TonoLab gave IOP readings that differed from manometry settings in normal young chicks over some or all of the ages tested. These results reinforce the importance of calibrating clinical tonometers in animal studies involving IOP as a key variable.

The guinea pig is becoming an increasingly popular model for studying human myopia, which carries an increased risk of glaucoma. As a step towards understanding this association, this study sought to characterize the normal, developmental intraocular pressure (IOP) profiles, as well as the anatomy of the optic nerve head (ONH) and adjacent sclera of young guinea pigs. IOP was tracked in pigmented guinea pigs up to 3 months of age. One guinea pig was imaged in vivo with OCT and one with a fundus camera. The eyes of pigmented and albino guinea pigs (ages 2 months) were enucleated and sections from the posterior segment, including the ONH and surrounding sclera, processed for histological analyses - either hematoxylin and eosin (H&E) staining of paraffin embedded, sectioned tissue (n = 1), or cryostat sectioned tissue, processed for immunohistochemistry (n = 3), using primary antibodies against collagen types I-V, elastin, fibronectin and glial fibrillary acidic protein (GFAP). Transmission and scanning electron microscopy (TEM, SEM) studies of ONHs were also undertaken (n = 2 & 5 respectively). Mean IOPs ranged from 17.33 to 22.7 mmHg, increasing slightly across the age range studied, and the IOPs of individual animals also exhibited diurnal variations, peaking in the early morning (mean of 25.8, mmHg, ∼9 am), and decreasing across the day. H&E-stained sections showed retinal ganglion cell axons organized into fascicles in the prelaminar and laminar region of the ONHs, with immunostained sections revealing collagen types I, III, IV and V, as well as elastin, GFAP and fibronectin in the ONHs. SEM revealed a well-defined lamina cribrosa (LC), with radially-oriented collagen beams. TEM revealed collagen fibrils surrounding non-myelinated nerve fiber bundles in the LC region, with myelination and decreased collagen posterior to the LC. The adjacent sclera comprised mainly crimped collagen fibers in a crisscross arrangement. Both the sclera and LC were qualitatively similar in structure in pigmented and albino guinea pigs. The well-organized, collagen-based LC of the guinea pig ONH is similar to that described for tree shrews and more similar to the human LC than that of other rodents that lack collagen. Based on these latter structural similarities the guinea pig would seem a promising model for investigating the relationship between myopia and glaucoma.

The chick is widely used in studies of eye growth regulation and myopia. The aim of this study was to explore the utility of pattern (p)ERG as a tool to assess retinal function in such studies. Effects of optical defocus and diffusing blur, manipulations used to alter eye growth experimentally, were evaluated. PERGs were recorded from White-Leghorn chickens, using a checkerboard pattern, including 8 spatial frequencies (0.05-2.2c/d SF), 13 contrast levels (1-100%), and 8 temporal reversal frequencies (0.5-20Hz). The acute effects of defocus and diffusing blur were examined. Flash- and pERGs were also recorded from chicks that underwent monocular optic nerve section (ONS), to explore the contribution of retinal ganglion cells (RGCs). Measurements were made up to 6weeks post-ONS, complemented with SD-OCT imaging. In normal chicks, the response to 1Hz, 100% contrast stimuli showed positive- and negative-going waveforms at 43ms (P1) and 75ms (N95), respectively, with 0.06-0.1c/d SF eliciting the largest P1 amplitudes of 21.9±2.5μV. Contrast levels above 5% yielded measurable P1 responses. Responses were transient and monophasic for 0.5-5Hzreversal rates, with higher temporal frequencies yielding steady state responses. Defocus and diffusing blur decreased pERG amplitude across all SFs. pERG responses remained normal after ONS, despite the loss of RGCs. In conclusion, chicks show robust pERG responses, which are attenuated by defocus and diffusing blur. The pERG response is not affected by ONS, suggesting that RGCs do not contribute to the chick pERG.

Purpose

Spectral domain optical coherence tomography (OCT) was used to examine the influence of refractive error (RE) on foveal retinal and choroidal thicknesses and scleral canal width (SCW). The performance of the Cirrus and Bioptigen spectral domain OCT instruments was compared in the same eyes.

Methods

Both eyes of 40 healthy human subjects, aged 22 to 38 years, were dilated and imaged, with the Cirrus OCT, using 6-mm five-line rasters collapsed into one line, one centered on the fovea and one bisecting the optic nerve head. Seventy-two of the same eyes were imaged with the Bioptigen OCT, using 6- by 6-mm scans, one centered on the fovea and one on the optic nerve head. Subfoveal retinal and choroidal thicknesses and SCW were measured. Axial lengths (ALs) and REs were obtained using an IOLMaster and a Grand Seiko autorefractor, respectively.

Results

Only right eyes were included in analyses. Spherical equivalent REs ranged from -12.18 to +8.12 diopters (mean [±SD], -3.44 [±4.06] diopters), and ALs ranged from 20.56 to 29.17 mm (mean [±SD], 24.86 [±1.91] mm). Myopia was associated with relatively thin choroids at the fovea (p < 0.05) but normal retinal thickness. Scleral canal width was significantly correlated with AL as measured with the Bioptigen OCT (p < 0.05). Retinal and choroidal thicknesses recorded with the Bioptigen OCT tended to be smaller than values obtained with the Cirrus OCT (mean difference, 5.63 and 24.76 μm, respectively), whereas the converse was true for the SCW (mean difference, 25.45 μm).

Conclusions

The finding that high myopes tend to have a thinner subfoveal choroid is consistent with previous studies. That high myopia was linked to enlarged scleral canals may help to explain the increased risk of glaucoma in myopia. Observed differences obtained with the Cirrus and Bioptigen instruments urge caution in comparing results collected with different instruments.

Purpose

The guinea pig is being used increasingly as a model of human myopia. As accommodation may influence the effects of manipulations used in experimental myopia models, understanding the accommodative ability of guinea pigs is important. Here, nonselective muscarinic agonists were used as pharmacological tools to study guinea pig accommodation.

Methods

Measurements were made on 15 pigmented guinea pigs. For in vivo testing, animals were anesthetized and, following baseline measurements, 2% pilocarpine was applied topically. Measurements included A-scan ultrasonography, optical coherence tomography (OCT) imaging, corneal topography, and refraction. In vitro lens scanning experiments were performed using anterior segment preparations, with measurements before and during exposure to carbachol. Anterior segment structures were examined histologically and immunohistochemistry was done to characterize the muscarinic receptor subtypes present.

Results

In vivo, pilocarpine induced a myopic shift in refractive error coupled to a small, but consistent decrease in anterior chamber depth (ACD), a smaller and more variable increase in lens thickness, and a decrease in pupil size. Lens thickness increases were short-lived (10 minutes), while ACD and pupil size decreased over 20 minutes. Corneal curvature was not significantly affected. Carbachol tested on anterior segment preparations in vitro was without effect on lens back vertex distance, but did stimulate pupil constriction. Immunohistochemistry indicated the presence of muscarinic receptor subtypes 1 to 5 in the iris and ciliary body.

Conclusions

The observed pilocarpine-induced changes in ACD, lens thickness, and refraction are consistent with active accommodation in the guinea pig, through cholinergic muscarinic stimulation.

Protein disulfide isomerase (PDI) family members regulate protein folding and calcium homeostasis in the endoplasmic reticulum (ER). The PDI family member anterior gradient (AGR) 3 is expressed in the airway, but the localization, regulation, and function of AGR3 are poorly understood. Here we report that AGR3, unlike its closest homolog AGR2, is restricted to ciliated cells in the airway epithelium and is not induced by ER stress. Mice lacking AGR3 are viable and develop ciliated cells with normal-appearing cilia. However, ciliary beat frequency was lower in airways from AGR3-deficient mice compared with control mice (20% lower in the absence of stimulation and 35% lower after ATP stimulation). AGR3 deficiency had no detectable effects on ciliary beat frequency (CBF) when airways were perfused with a calcium-free solution, suggesting that AGR3 is required for calcium-mediated regulation of ciliary function. Decreased CBF was associated with impaired mucociliary clearance in AGR3-deficient airways. We conclude that AGR3 is a specialized member of the PDI family that plays an unexpected role in the regulation of CBF and mucociliary clearance in the airway.

The choroid is the richly vascular layer of the eye located between the sclera and Bruch's membrane. Early studies in animals, as well as more recent studies in humans, have demonstrated that the choroid is a dynamic, multifunctional structure, with its thickness directly and indirectly subject to modulation by a variety of physiologic and visual stimuli. In this review, the anatomy and function of the choroid are summarized and links between the choroid, eye growth regulation, and myopia, as demonstrated in animal models, discussed. Methods for quantifying choroidal thickness in the human eye and associated challenges are described, the literature examining choroidal changes in response to various visual stimuli and refractive error-related differences are summarized, and the potential implications of the latter for myopia are considered. This review also allowed for the reexamination of the hypothesis that short-term changes in choroidal thickness induced by pharmacologic, optical, or environmental stimuli are predictive of future long-term changes in axial elongation, and the speculation that short-term choroidal thickening can be used as a biomarker of treatment efficacy for myopia control therapies, with the general conclusion that current evidence is not sufficient.