Comparison of invasive and non-invasive tonometry in the mouse

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Abstract

Assessment of the accuracy of non-invasive rebound tonometry, and comparison with invasive cannulation tonometry. An in vivo calibration technique was devised to improve the accuracy of the rebound tonometer. IOP was then measured in SW mice using both rebound and cannulation tonometry. The ability of the rebound tonometer to accurately measure small IOP reductions after instillation of a topical prostaglandin was also determined. With the rebound method, mid-afternoon IOP in two groups of similar aged SW mice was 15.9±3.9 mmHg (mean±s.d., n=25) compared to 16.3±1.2 mmHg (n=32) using the cannulation technique. This difference was not statistically significant (p=0.6). For serial measurements using both techniques in the same eyes of a third group of SW mice (n=14), mean IOP was 15.0±3.9 mmHg for rebound tonometry but only 13.4±2.3 mmHg for subsequent cannulation tonometry. This effect was subsequently shown to be a consequence of the rebound tonometry, as multiple rebound measurements induced a statistically significant reduction in IOP. The average IOP reduction observed 2 hr after a single application of topical latanoprost (200 ng) was 2.8±1.3 mmHg (p<0.001) and 2.4±4.7 mmHg (p=0.03) with cannulation and rebound tonometers, respectively. These differences were not significantly different (p=0.8). In vivo calibration of the rebound tonometer increased measurement accuracy and provided IOP values within the physiological range that agreed closely with the IOP measured by cannulation tonometry. However, IOP measurement with the rebound tonometer had larger variability compared with the cannulation method. Repeat IOP measurements with the rebound tonometer led to a reduction in IOP. The rebound tonometer was sufficiently sensitive to detect a 2–3 mmHg reduction in IOP following application of topical latanoprost. Despite these limitations, the rebound tonometer has a significant advantage over cannulation tonometry in that it permits longitudinal IOP measurement in conscious mice.

Introduction

Mouse and human eyes share a number of anatomic and physiological similarities including well-established uveoscleral and trabecular meshwork outflow pathways, comparable intraocular pressures (IOP), and similar IOP responses to topical glaucoma medications (Aihara et al., 2003, Crowston et al., 2004a). These features along with the availability of genetically modified mice are particularly attractive for glaucoma research. Technical difficulties associated with the measurement of IOP in mouse eyes has, however, limited the use of these animals in certain experimental paradigms.

Cannulation of the anterior chamber permits measurement of true manometric IOP in the mouse (John et al., 1997). Although highly accurate, cannulation can be technically difficult. Further, it necessitates perforation of the cornea and the administration of general anesthesia; both of these may affect IOP. In addition, several days are required for the corneal perforation to heal before a subsequent IOP measurement can be reliably obtained. These disadvantages have led to the development of non-invasive techniques for IOP measurement in the mouse.

Non-invasive techniques include the induction-impact (rebound) tonometer (Danias et al., 2003), a modified Goldmann applanation tonometer (Cohan and Bohr, 2001b), a Schiotz-like indentation tonometer (Gross et al., 2003) and the tonopen (Reitsamer et al., 2004). Previous studies using the rebound tonometer have reported IOPs that were lower than the IOP range reported for the same strain of mice using cannulation tonometry (Danias et al., 2003). This may be a direct consequence of the calibration technique, which was performed on enucleated eyes. We hypothesized that an in vivo calibration technique performed in anesthetized mice would improve the accuracy of the rebound tonometer and provide IOP values within the physiological range. In addition, this study sought to determine the accuracy and variability of the rebound tonometer in the measurement of mouse IOP and to compare these with IOP measurements obtained with the cannulation tonometer.

Section snippets

Animal husbandry

All experiments were performed in compliance with the ARVO Statement for the Use of Animals in Ophthalmic and Vision Research. NIH Swiss White (SW) mice were obtained from Harlan Sprague–Dawley (San Diego, CA). The environment was kept at 21 °C with a 12-hr light (6:00–18:00) and 12-hr dark cycle. All mice were fed ad libitum. Animal age ranged from 8 to 24 weeks.

Anesthesia

Mice were anesthetized by intraperitoneal injection of a mixture of ketamine (100 mg kg−1, Ketaset, Fort Dodge Animal Health, Fort

Tonometer calibration in vivo

The correlation between the pressure transducer used for IOP measurements and a digital manometer revealed a close linear correlation (y=0.995x+0.313, r2=0.99, Fig. 1a). In vivo calibration of the rebound tonometer in anesthetized SW mice produced calibration curves with a quadratic best fit with the equation: y=−0.0013x2+0.10x+0.035 (r2=0.99, Fig. 1b). Four calibrations were performed on four separate SW mice. Each calibration produced the same shaped calibration curve, which were also similar

Discussion

The induction-impact (rebound) tonometer provides a straightforward, technically undemanding and non-invasive method of measuring mouse IOP. These data indicate that the rebound tonometer provides accurate and reproducible IOP measurements in the mouse eye. Use of an in vivo calibration technique provided slope conversion values of mean IOP that were within the physiological range and agreed closely with cannulation IOP measurements. The calibration curve suggests that the rebound tonometer may

Acknowledgements

This work was supported by grants R03 EY13732, EY 01867 and RPB.

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Supported in part by the National Eye Institute EY05990 (RNW).

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