Muscarinic receptor antagonist and an alpha-adrenergic agonist are required in combination to provide stable mydriasis following intravitreal injection in mice

Abstract

Tropicamide (muscarinic receptor antagonist) and phenylephrine (α-adrenergic receptor agonist) are commonly used to dilate the pupils by topical application. These two eye drops are often used, singly or in combination, to dilate the pupil and perform acute light-evoked physiological experiments (electroretinography, for example), before and after intravitreal injections of pharmacological agents, as an assay for their affect on retinal activity. This study wanted to determine whether treatment with one of these drugs, or with both, is most effective in maintaining mydriasis after intravitreal injections. Changes in pupillary dilation before and after intravitreal injection of balanced salt solution (0.5 µl) were recorded. Phenylephrine (α-adrenergic agonist) and tropicamide (muscarinic agonist) when combined, but not singly, produced full and stable pupillary dilation following intravitreal injections. Re-instillation of topical mydriatics after intravitreal injections was required for maximal pupillary dilation. A combination of a muscarinic receptor antagonist and an alpha-adrenergic agonist is required for stable mydriasis following intravitreal injection.

Figure 1. Experimental sequence

Three time-points of pupillary recording are shown in grey. Star: variable time interval between and the intravitreal injection (IVT) and recording of the pupillary area when the cylindrical bath from the eye was removed and the animal was injected with balanced salt solution.

Figure 2. Pupillary dilation after topical mydriatic instillation before and after intravitreal injections

Dashed circles indicate the pupillary circumference. The cylindrical bath was removed for intravitreal injections after A, D, G. White opacity in E is a cataract, which decreased over time after re-application of cylindrical bath. Black arrows indicate sparing of the sutures in the cataracts. White arrowhead: blood from the injection site. IVT: intravitreal.

Figure 3. Application of mydriatic followed by removal and reapplication of cylindrical bath does not alter the state of mydriasis caused by phenylephrine within one hour of mydriatic instillation

The two pairs A,B and C,D are from two different animals. A,C: pupil size 10 min after instillation of mydriatic in the cylindrical bath. The cylindrical bath was removed after A and C and reapplied after 15 min in B and 60 min in D. White opacity seen in B and D are cataracts. Black arrow shows the sparing of lens sutures in B, which is no longer visible at later times (D).

Figure 4. Average pupillary dilation after topical mydriatic instillation and following intravitreal injections

A. Pupillary area (PA) was divided by area enclosed by the limbus (LA) to yield the normalized pupillary area. B. Averaged normalized pupillary area measured for the three groups (phenylephrine treatment, Tropicamide treatment and phenylephrine + tropicamide treatment) at three time points (x-axis). The pupillary area recorded at 10 min after application of mydriatic in the bath showed no statistically significant difference (One-way ANOVA). Star: statistically significant difference (Bonferroni-Holm post-hoc test; adj. alpha < 0.05); n=4 for each group.