Guinea pig ciliary muscle development

Abstract

Purpose: The purpose of this study was to develop a method for quantifying guinea pig ciliary muscle volume (CMV) and to determine its relationship to age and ocular biometric measurements.

Methods: Six albino guinea pigs' eyes were collected at each of five ages (n = 30 eyes). Retinoscopy and photography were used to document refractive error, eye size, and eye shape. Serial sections through the excised eyes were made and then labeled with an α-smooth muscle actin antibody. The ciliary muscle was then visualized with an Olympus BX51 microscope, reconstructed with Stereo Investigator (MBF Bioscience), and analyzed using Neurolucida Explorer (MBF Bioscience). Full (using all sections) and partial (using a subset of sections) reconstruction methods were used to determine CMV.

Results: There was no significant difference between the full and partial volume determination methods (p = 0.86). The mean (±SD) CMV of the 1-, 10-, 20-, 30-, and 90-day-old eyes was 0.40 (±0.16) mm, 0.48 (±0.13) mm, 0.67 (±0.15) mm, 0.86 (±0.35) mm, and 1.09 (±0.63) mm, respectively. Ciliary muscle volume was significantly correlated with log age (p = 0.001), ocular length (p = 0.003), limbal circumference (p = 0.01), and equatorial diameter (p = 0.003). It was not correlated with refractive error (p = 0.73) or eye shape (p = 0.60). Multivariate regression determined that biometric variables were not significantly associated with CMV after adjustment for age.

Conclusions: Three-dimensional reconstruction was an effective means of determining CMV. These data provide evidence that ciliary muscle growth occurs with age in tandem with eye size in normal albino guinea pigs. Additional work is needed to determine the relationship between CMV and abnormal ocular growth.

Figure 1

Examples of best-fit circles for determining structural areas. Note that white lines are outlining examples of the coronal (A), transverse (B), and limbal (C) areas.

Figure 2

An example tracing of stained ciliary muscle. Note that the ciliary muscle is outlined in red (10x objective).

Figure 3

Determination of optimal number of ciliary muscle sections. The mean area of ciliary muscle sections are plotted with an increasing number of sections at each point. Note that the mean area is approximately stable with ten or more sections and that each side of an error bar represents one standard deviation from the mean.

Figure 4

An example of stacked ciliary muscle sections prior to computer generated three-dimensional reconstruction. Note how tracings like seen in Figure 2 are taken from throughout the entire eye, stacked, and aligned to allow for three-dimensional reconstruction of the ciliary muscle.

Figure 5

Reconstructed images of ciliary muscle volume at 1, 10, 20, 30, and 90 days of age. Note that each arrow and associated value represents the mean inner ciliary muscle ring diameter at the listed time-point. Also note that superior and inferior ring thinning can be seen in some images; this artifact is due to the software during the reconstruction process.

Figure 6

Ciliary muscle volume at different ages. The presented data are best represented as a log growth pattern (R² = 0.81). Each side of an error bar represents one standard deviation from the mean.

Figure 7

An example picture of 1-day-old guinea pig ciliary muscles. Note that the ciliary muscle spans its natural attachment points at 1 day of age (4x objective).

Figure 8

Length of ciliary muscle sections over time. The presented data are best represented as a log growth pattern (R² = 0.81). Each side of an error bar represents one standard deviation from the mean.

Figure 9

An Estimate of Mean Ciliary Muscle Thickness Over Time. The presented data are best represented as a log growth pattern (R² = 0.75). Each side of an error bar represents one standard deviation from the mean.