Opposite effects of glucagon and insulin on compensation for spectacle lenses in chicks

Abstract

Purpose: Chick eyes compensate for the defocus imposed by positive or negative spectacle lenses. Glucagon may signal the sign of defocus. Do insulin (or IGF-1) and glucagon act oppositely in controlling eye growth, as they do in metabolic pathways and in control of retinal neurogenesis?

Methods: Chicks, wearing lenses or diffusers or neither over both eyes, were injected with glucagon, a glucagon antagonist, insulin, or IGF-1 in one eye (saline in the other eye). Alternatively, chicks without lenses received insulin plus glucagon in one eye, and either glucagon or insulin in the fellow eye. Ocular dimensions, refractive errors, and glycosaminoglycan synthesis were measured over 2 to 4 days.

Results: Glucagon attenuated the myopic response to negative lenses or diffusers by slowing ocular elongation and thickening the choroid; in contrast, with positive lenses, it increased ocular elongation to normal levels and reduced choroidal thickening, as did a glucagon antagonist. Insulin prevented the hyperopic response to positive lenses by speeding ocular elongation and thinning the choroid. In eyes without lenses, both insulin and IGF-1 speeded, and glucagon slowed, ocular elongation, but glucagon and insulin each increased the rate of thickening of the crystalline lens. When injected together, insulin blocked choroidal thickening by glucagon, at a dose that did not, by itself, thin the choroid.

Conclusions: Glucagon and insulin (or IGF-1) cause generally opposite modulations of eye growth, with glucagon mostly increasing choroidal thickness and insulin mostly increasing ocular elongation. These effects are mutually inhibitory and depend on the visual input.

FIGURE 1

The effects of glucagon on ocular length and the thickness and GAG synthesis of the choroid. Glucagon increased choroidal thickness (A, B, D, and J) and decreased the rate of ocular elongation (F, G, I, and J) in eyes wearing diffusers, -7 D lenses, or no devices. The opposite effect was found in eyes wearing positive lenses (C, H, and J). Glucagon also increased the rate of choroidal GAG synthesis (E) in eyes without lenses or diffusers. Panel J shows that, in eyes wearing positive lenses, glucagon (2 nmol) increased the rate of ocular elongation in nearly all birds and reduced choroidal thickening in half of them. Panel J includes 7 birds (five 1-week-old and two 2-week-old) from pilot experiments not shown in Table 1. Bar-charts show the mean changes and SEMs over the course of the experiments shown in the line-graphs, in which solid and interrupted lines show the time course of the experimental and fellow eyes, respectively. For choroidal GAG synthesis, the ratios of the scintillation counts in paired eyes (glucagon/PBS) of individual chicks (circles) are superimposed on the mean (bars) and SEM of each group. Analysis of Variance (ANOVA) with Bonferroni post hoc tests were used to test the statistical significance of interocular difference (X-N) among different days within a group with respect to day 0 (asterisks on line-graphs), and of relative changes (ΔX-ΔN) among the different doses (asterisks over horizontal lines in panels D and I). For ocular length and choroidal thickness, asterisks over bars indicate paired, 1-tailed Student t-tests comparing changes in experimental and fellow eyes, except for panels C and H, in which paired, 2-tailed Student t-tests were used. For GAG synthesis, one-sample t-tests were used to compare the ratio of the counts from paired eyes (glucagon/PBS, or X/N) with a value of 1. *p < 0.05, **p < 0.01, ***p < 0.001.

FIGURE 2

The effect of glucagon on lens thickness. A. Glucagon increased lens thickness in eyes wearing diffusers or negative lenses. B. Glucagon increased lens thickness in a dose-dependent fashion in eyes not wearing lenses. Asterisks over horizontal lines show comparisons of relative changes (ΔX-ΔN) that were significant by ANOVA with Bonferroni post hoc test. Asterisks over bars show significant difference in changes in experimental and fellow eyes from paired, 2-tailed Student t-tests.

FIGURE 3

The effects of a glucagon antagonist on changes in paired eyes (ΔX and ΔN) in ocular length (A) and choroidal thickness (B). The glucagon antagonist, at the lower dose, reduced the inhibition of ocular elongation caused by positive lenses and, at the higher dose, returned the rate of ocular elongation to normal or above (dotted line is normal elongation over 2 days without injections). The glucagon antagonist tended to reduced choroidal thickness only when the chicks had worn positive lenses for 2 days prior to the start of the injections (B, right-most bar). Statistics as in Fig. 1. Left-hand bars of each pair (change over first two days) are data pooled from eyes wearing +7 or +15 D lenses and injected with 26 pmol (groups 11 & 12) or from eyes wearing +10 or +15 D lenses and injected with 305 pmol (groups 14 & 15). The right-most bars are data pooled from eyes wearing +10 or +15 D lenses 2 days prior to the start of injections (groups 16 and 17).

FIGURE 4

The effects of insulin on changes in paired eyes in ocular length, scleral GAG synthesis, and choroidal thickness. (B-G) Ocular length (measured values) as a function of time (in days) are shown for each dose under the bar-chart showing the changes in paired eyes over the 3-day duration of the experiments (all y-axes are identical). Insulin increased the rate of ocular elongation in eyes wearing positive lenses (panels on left) and in eyes not wearing lenses (panels on right), as well as increasing scleral GAG synthesis in eyes wearing positive lenses (I). (H) Insulin also reduced choroidal thickness in eyes wearing positive lenses. Statistics as in Fig. 1. The dotted line shows the normal rate of ocular elongation (A) and choroidal thickness (H) over 3 days without injections. ‡: injections with a 35-gauge needle.

FIGURE 5

The effects of insulin on anterior chamber depth (A) and lens thickness (B). Insulin increased anterior chamber depth (A) and lens thickness (B) in a dose-dependent fashion. Statistics as in Fig. 2.

FIGURE 6

The effects of IGF-1 on changes in ocular length, choroidal thickness, and anterior chamber depth. IGF-1 at the highest dose increased the rate of ocular elongation (A), choroidal thickness (B), and anterior chamber depth (C) in eyes not wearing lenses. Statistics as in Figs. 1 and 2.

FIGURE 7

The interaction between glucagon- and insulin-dependent mechanisms on choroidal thickness (A, B), choroidal GAG synthesis (C), ocular length (D, E), and scleral GAG synthesis (F). Although insulin alone (at 0.01 U, 68 pmol) did not reduce choroidal thickness, it completely blocked the increase in both choroidal thickness (B: compare G vs. G+I) and choroidal GAG synthesis (C) caused by glucagon. Conversely, although glucagon by itself did not affect normal ocular elongation at either dose used, at the high dose it reduced the effect of insulin on the rate of ocular elongation by half (E: compare G+I vs. I). Glucagon also decreased the effect of insulin the rate of scleral GAG synthesis (F: compare G+I vs. I). Statistics as in Fig. 1. ‡: eyes injected with a 35-gauge needle.