Horizontal Central Ridge of the Lamina Cribrosa and Regional Differences in Laminar Insertion in Healthy Subjects

Abstract

Purpose.: To assess the general morphology and position of the lamina cribrosa (LC) in healthy subjects using enhanced depth imaging–optical coherence tomography (EDI-OCT).

Methods.: Serial horizontal and vertical B-scans of the optic nerve head (interval between images, approximately 30 μm) were prospectively obtained using EDI-OCT for both eyes of each healthy subject. After delineation of the anterior laminar surface, mean and maximum LC depths were measured in 11 equally spaced horizontal B-scans, and the depth of the anterior LC insertion was measured at 32 points along its circumference (reference plane, Bruch's membrane edges) for one randomly selected eye of each subject. Three-dimensional (3D) images of the anterior laminar surface and the peripapillary sclera were reconstructed from serial horizontal EDI-OCT B-scans to assess the 3D morphology of the anterior laminar surface.

Results.: Among the 61 eyes (61 subjects) enrolled, 31 were excluded because of poor LC image quality, and 30 were included for analysis (mean age, 40 ± 18 [range, 21–78] years). Both mean and maximum LC depth profiles showed an elevation in the central area and a depression in the superior and inferior midperiphery of the LC. The anterior LC insertion was more posteriorly located in the superior and inferior than in the nasal and temporal regions. Three-dimensional LC images showed a bowtie-shaped horizontal central ridge of the LC.

Conclusions.: The LC has a central ridge ranging from the temporal to the nasal insertion areas and inserts more posteriorly in the superior and inferior than in the nasal and temporal regions. Further investigation is needed to elucidate the significance of these findings in the pathophysiology of glaucoma.

The lamina cribrosa (LC) is a series of sieve-like collagenous plates at the optic nerve head through which the retinal ganglion cell axons and the retinal blood vessels pass. ^{1 –3} It is often considered the primary site of axonal injury in glaucoma. ^{4 –10} Histologic studies using enucleated or postmortem eyes have demonstrated structural changes of the LC in glaucoma, ^{4 –14} but histologic processing of specimens may alter the architecture seen in vivo. A variety of imaging devices, including spectral domain–optical coherence tomography (SD-OCT), have recently been used to evaluate the LC in vivo. ^{15 –21} However, previous in vivo investigations ^{15 –21} reported an inability to visualize the anterior laminar surface beneath the neuroretinal rim, vascular structures, and scleral rim and were not able to demonstrate the morphology or position of the entire LC in detail. 

Conventional OCT has limited ability to image the deep structures of the posterior segment because of a depth-dependent decrease in sensitivity and a scattering of light by pigment and blood. ²² Enhanced depth imaging–OCT (EDI-OCT) was developed to improve the image quality of the deep posterior segment structures ²² and has been used to investigate the in vivo microanatomy of the outer retina, choroid, and sclera. ^{23 –25} Additionally, it has been demonstrated that EDI-OCT is able to better visualize the LC than conventional OCT. ^{26 –28}  

In the present study, we evaluated the general morphology and position of the LC in healthy subjects by measuring the depths of the LC and LC insertion at different locations using EDI-OCT. 

Methods

This is a cross-sectional analysis of data obtained from an ongoing, prospective, longitudinal study approved by the New York Eye and Ear Infirmary Institutional Review Board. Written, informed consent was obtained from all subjects, and the study adhered to the tenets of the Declaration of Helsinki. 

We included healthy subjects and performed a detailed medical history, slit-lamp biomicroscopy, Goldmann applanation tonometry, gonioscopy, and stereoscopic optic disc examination. For both eyes of each participant, an infrared optic disc photograph was taken, and peripapillary retinal nerve fiber layer thickness was measured along the 3.4-mm–diameter circle using SD-OCT (Spectralis; Heidelberg Engineering GmbH, Dossenheim, Germany). Serial horizontal and vertical B-scan images (interval between images, approximately 30 μm) of the optic nerve complex were obtained using the same OCT device with the EDI-OCT method. We excluded subjects who had previously undergone intraocular surgery; subjects who had systemic or ocular abnormalities known to affect the optic nerve structure or visual function; subjects with torted optic disc; and EDI-OCT images of poor quality because of media opacity or poor patient cooperation, causing diffusely unclear images or significant artifacts (e.g., mirror artifacts, out-of-range artifacts, z-alignment failure). The optic disc was classified as torted when the axis of longest optic disc diameter differed by >10° from the vertical axis of the optic disc based on optic disc photography. This is a stricter definition than that adopted in the Blue Mountains Eye Study. ²⁹  

For EDI-OCT of the optic nerve complex, we used the method described in a previous report. ²² In brief, the OCT device was set to image a 15° × 10° rectangle for horizontal scans (and a 10° × 15° rectangle for vertical scans) centered on the optic disc. This rectangle was scanned with 97 sections, and each section had 20 OCT frames averaged. Because the distance between adjacent scans was ∼30 μm, 97 scans covered an area of ∼3 mm. The device was pushed close enough to the eye to create an inverted image with the inner portions of the retina shown facing downward. The OCT images shown in this article were inverted after they were exported from the OCT device. 

For one randomly selected eye of each subject, mean and maximum LC depths were measured in 11 horizontal B-scans that were equally spaced along the vertical diameter of the LC, and the depth of the anterior LC insertion was measured at 32 points along its circumference (Fig. 1). The line connecting Bruch's membrane edges was used as a reference plane for all the depth measurements (Fig. 1). A line was drawn from each of the two LC insertion points perpendicularly to the line connecting the two Bruch's membrane edges. The area surrounded by these two lines—the line connecting Bruch's membrane edges—and the anterior laminar surface was measured (Fig. 1B, area S). Then the mean LC depth was approximated by dividing area S by length D for each of the 11 horizontal EDI-OCT scans (Fig. 1B). The depth of the LC insertion was measured at both LC insertion points in each of the 11 horizontal B-scans (22 points) and the five vertical B-scans (10 points). The vertical scans were equally spaced between the LC insertion points in the most superior and inferior horizontal B-scans (Fig. 1C). All measurements were performed using ImageJ software (developed by Wayne Rasband, National Institutes of Health, Bethesda, MD; available at http://rsb.info.nih.gov/ij/index.html). The anterior laminar surface was manually delineated as if the LC had no pores, including those for retinal vessels. EDI-OCT images were appropriately magnified in a 24-inch monitor to delineate the anterior laminar surface more reliably and accurately. When the anterior laminar surface was unclear especially near the laminar insertion, we changed the contrast, brightness, or both of the OCT image portion of interest (Fig. 2). When the target microstructures were not clearly visualized, an adjacent horizontal EDI-OCT scan, approximately 30 μm from the original scan, was used for measurement. Eyes were excluded if they had an uncertain area of anterior laminar surface larger than the laminar pore or retinal vessel in any of the selected 11 horizontal and five vertical EDI-OCT scans. 

Figure 1.

 

(A) The maximum depth of the LC was measured (black arrow; perpendicular to the line connecting the two Bruch's membrane edges), (B) the mean depth of the LC was approximated (area S divided by length D) in EDI-OCT B-scans obtained (C) in 11 equally spaced horizontal scans, and the depth of the laminar insertion (A, white arrows) was measured at 32 points (C, black dots) along its circumference. (A, white and black dots) Bruch's membrane edges and laminar insertion points, respectively. Five vertical B-scans were equally spaced between the two laminar insertion points in the most superior and inferior horizontal B-scans.

Figure 2.

 

For the OCT image portion with an unclear anterior laminar surface, contrast, brightness, or both were changed for best visualization. (A) The anterior laminar surface is not clear in the inset. The anterior laminar surface near its insertion before (B) and after (C) the contrast and brightness of the OCT image were changed. (D, yellow dot and yellow line) LC insertion point and anterior laminar surface, respectively.

Three-dimensional (3D) images of the anterior laminar surface and the peripapillary sclera were reconstructed from all obtained serial horizontal cross-sectional EDI-OCT scans after manually outlining the anterior laminar and scleral surfaces and the anterior scleral canal wall using commercially available 3D reconstruction software (Amira, version 5.3.3.; Visage Imaging, Inc., San Diego, CA) to assess the 3D shape of the anterior laminar surface. The OCT scans, which had been automatically aligned by the built-in software of the OCT device, were exported and then uploaded to the 3D reconstruction software. Bruch's membrane edges were marked in each horizontal EDI-OCT image used for 3D reconstruction to assess the shape of the Bruch's membrane opening, which we used as a reference plane for the LC depth measurements. 

Results

Among the 61 healthy subjects who met the entry criteria, 31 were excluded because of poor image quality of the anterior laminar surface. Nineteen (63%) of 30 included subjects were women. Mean ± SD age was 40 ± 18 years (range, 21–78 years). All eyes had normal-appearing open iridocorneal angles, nontorted optic discs, intraocular pressure between 10 and 18 mm Hg, and no apparent ocular or systemic abnormality that could have affected the optic nerve structure or visual function. According to the manufacturer-provided normative database, retinal nerve fiber layer thickness was within normal limits in all eyes. Among the 330 (11 scans × 30 eyes) horizontal and 150 (5 scans × 30 eyes) vertical EDI-OCT scans analyzed, 39 (12%) and 27 (18%) were adjacent scans next to (∼30 μm apart from) the initially selected scans with unclear anterior laminar surfaces, respectively. Among 960 (32 points × 30 eyes) laminar insertion points analyzed, 114 (12%) points were decided after adjusting the image brightness, contrast, or both. 

Both mean and maximum LC depth profiles (superior to inferior horizontal scans) assumed the shape of a W, demonstrating that the LC is elevated in the central area and has a depression in the superior and inferior midperiphery (P ≤ 0.001 for scans 1 vs. 3, 3 vs. 7, 7 vs. 9, and 9 vs. 11 by generalized estimating equation regression model; Fig. 3A). The mean depth of LC insertion was 265 ± 68 μm (range, 166–451 μm). The depth profile of the LC insertion along its circumference (temporal-superior-nasal-inferior-temporal points) also assumed the shape of a W, revealing that the LC insertion is more posteriorly located in the superior and inferior than the nasal and temporal quadrants (P < 0.001 for LC insertion point 1 vs. 9, 9 vs. 17, 17 vs. 25, and 25 vs. 1 by generalized estimating equation regression model; Fig. 3B). The anterior laminar surface in the vertical EDI-OCT images assumed the shape of a W because of an elevation in the central area and a depression in the superior and inferior midperiphery (Figs. 4B, 4H). However, central elevation was absent or unremarkable in the horizontal EDI-OCT images (Figs. 4E, 4K). In addition, the depth of LC insertion was greater in the superior and inferior than in the nasal and temporal regions in all eyes (Figs. 4B, 4E, 4H, 4K). 

Figure 3.

 

(A) Mean and maximum depth profiles of the anterior laminar surface in 11 horizontal scans (scan 1 → 11 = superior → inferior). (B) Depth profile of the laminar insertion at 32 points along the laminar circumference (point 1 → 32 = temporal → superior → nasal → inferior → temporal). Depth with a blue asterisk was significantly greater than depth with a red asterisk in the same profile (P ≤ 0.001, A; P < 0.001, B; by generalized estimating equation regression model). Negative value reflects the position below the reference plane (Bruch's membrane edges). Error bars represent SD. Measurement points of the LC insertion depth are shown equally spaced even though they were not measured as such.

Figure 4.

 

(A, D, G, J) Infrared optic disc photographs and (B, H) vertical and (E, K) horizontal EDI-OCT scans of two normal right eyes. (C, F, I, L) The same images as in (B, E, H, K) without the labels (A–F, subject 4; G–L, subject 11). The anterior laminar surface assumes an extended W shape in vertical B-scans, but not in horizontal B-scans, because of the presence of the horizontal central ridge of the LC. Note that the depths of laminar insertion are greater in the superior and inferior than in the nasal and temporal regions of the LC. (M) Mean and maximum depth profiles of the anterior laminar surface and (N) depth profiles of the laminar insertion of subjects 4 and 11. Measurement points of the LC insertion depth are shown equally spaced even though they were not measured as such. (A, D, G, J, green arrows) Location and direction of the vertical and horizontal OCT scans. (B, E, H, K, green lines, yellow dots, yellow lines, red lines, yellow dashed lines, and numbers) Intersection points of the vertical and horizontal OCT scans, LC insertion points, anterior laminar surface, anterior scleral surface and scleral canal, line connecting Bruch's membrane edges, and depth of laminar insertion in micrometers, respectively.

These W-shaped depth profiles of the LC and the LC insertion implied that the LC has a central ridge ranging from the temporal to the nasal insertion areas. This horizontal central ridge of the LC was also observed in 3D images, revealing its bowtie shape and its variability in size and location among eyes (Fig. 5A–D). The central retinal vessel trunk passed through the horizontal laminar ridge in all cases. Therefore, the location of the horizontal laminar ridge corresponded to that of the central retinal vessel trunk in the LC. The LC tissue surrounding the central retinal vessel trunk constituted a humplike structure in the center of the horizontal laminar ridge (knot of the bowtie). When viewed orthogonally from the superior, inferior, nasal, and temporal sides, the landmarks of the Bruch's membrane edges were well aligned in one plane or were located more posteriorly in the superior and inferior areas than in the nasal and temporal areas, with no significant outliers in all 30 eyes (Figs. 5E, 5F). 

Figure 5.

 

(A, B) 3D images of the anterior laminar surface and peripapillary sclera of two normal right eyes (subjects 4 and 11, respectively). Note the bowtie-shaped horizontal central ridge of the LC and the humplike structure in the center of the ridge (knot of the bowtie). (C, D) The same figures as A and B without the labels. (E, F) Landmarks for Bruch's membrane edges in all horizontal OCT scans viewed from the vitreous, nasal, and inferior sides. I, inferior; N, nasal; S, superior; T, temporal; A, anterior; P, posterior, respectively)

Discussion

In the present study, the W-shaped depth profiles of the LC and LC insertion as well as the 2D EDI-OCT images of the LC and 3D reconstructions of the anterior laminar surface suggested the presence of a horizontal central ridge spanning the LC. Figures in previous reports showed some clues to the presence of horizontal central ridge (see Fig. 6), ^5,13 and the shape of anterior laminar surface was described as a saddle. ⁵ However, its presence or clinical and pathophysiological significance was not specifically mentioned or described, possibly because of its subtle structure. Previous histologic studies demonstrated a lower density of connective tissue and glial cell processes and a greater diameter of laminar pores in the superior and inferior compared with the nasal and temporal regions of the LC. This has been suggested as the reason for the inherently greater susceptibility to glaucomatous damage of the optic nerve fibers passing through the superior and inferior poles of the optic nerve head. ^5,6 Another histologic study revealed that the successive LC sheets are compressed early in glaucoma and that the LC undergoes backward bowing at a later stage, which involves the superior and inferior poles more than the middle region of the optic nerve head. ⁸ These previous findings suggest that the horizontal central ridge of the LC may have connective tissue and glial cell processes of greater density and laminar pores of smaller diameter and, therefore, may be associated with regional difference in the susceptibility of the optic nerve head to glaucomatous damage; this requires further investigation. 

Figure 6.

 

Human optic nerve head (right eye) after osmotic and detergent digestion of cellular tissue, oriented with superior part of the optic nerve head above and viewed from vitreous side by light microscopy (50×). A horizontal central ridge is visible, consistent with the EDI-OCT findings shown in Figure 5. Reprinted with permission from Quigley HA, Addicks EM. Regional differences in the structure of the lamina cribrosa and their relation to glaucomatous optic nerve damage. Arch Ophthalmol. 1981;99:137–143.

The connective tissue around the central retinal vessel trunk appeared to be important to the integrity of the horizontal central ridge of the LC, forming a humplike structure in its center. In 1947, Wilczek ¹ called the shallow depression consisting of the scleral canal and the anterior laminar surface the “nest of the optic disc ” and described the bottom of this nest as “uneven and slightly knob-like.” In the fifth month of fetal development, connective tissue trabeculae grow into the optic nerve head and strengthen primitive neuroglial septa to form the LC. ^30,31 Although not substantiated, it has been described that this occurs from all directions and from all available sources of connective tissue such as the sclera, pia mater, and connective tissue that accompanies the central retinal vessels. ¹ Therefore, it can be postulated that the presence of a humplike structure in the center of the horizontal laminar ridge may be attributed to the outgrowing trabeculae from the connective tissue around the central retinal vessel trunk in addition to those from the sclera, pia mater, or both during fetal development. 

In the present study, we demonstrated the regional difference in the position of LC insertion, which was more posteriorly located in the superior and inferior than in the nasal and temporal regions. Wilczek ¹ suggested that the LC may insert into the pia mater in some eyes. This was confirmed by Jonas et al. ³² and Sigal et al., ¹⁴ who demonstrated a substantial amount of LC inserting into the pia mater. Regional differences in the position of the LC insertion may affect the pial insertion of the LC, in relation to the regional difference in the scleral thickness and the LC thickness. In our EDI-OCT images, however, we could not determine whether the LC was inserting into the sclera or the pia mater. The effect of a more posteriorly positioned LC insertion in the superior and inferior regions on the pialization of the LC insertion and its significance in the pathogenesis of glaucoma must be investigated further. 

We did not investigate the regional differences in LC thickness because the posterior sclera surface, pia mater, and posterior laminar surface near the laminar insertion were invisible. Results of previous histologic studies on the regional thickness of the LC are inconsistent. ^13,33,34 Dichtl et al. ³³ reported that the LC was significantly thicker in the peripheral parts of the disc than in its central region, but the LC was thinner in the area close to the optic disc border than in the central area in a study by Jonas et al. ³⁴ In another report, LC thickness varied according to the location of the measurement (center, midperiphery, and periphery). ¹³ Inconsistency among the previous results may be attributed to the complexity and high interindividual variability of the LC structure and thickness. Further in vivo investigation of regional differences in laminar thickness is warranted using more sophisticated imaging devices. 

Approximately half the examined subjects were excluded from analysis because the quality of the EDI-OCT images was not good enough for reliable measurement of the depths of the LC or its insertion, possibly because of thicker prelaminar neural tissue, thicker scleral rim, or more significant vascular shadowing above the LC in the course of OCT beams in the excluded eyes. Therefore, it is possible that our results may be representative of a subgroup of healthy subjects with good visibility of the LC in EDI-OCT images. Additionally, as described in Methods, we delineated the anterior laminar surface as if the LC had no pores. It should be kept in mind that this study assessed the general morphology and position of the LC in healthy subjects, not focal undulation or spikiness of the LC in detail. We did not quantify the variability of our delineations of the anterior laminar surface. However, we think the variability was low enough not to have influenced the main outcomes of our study. Although we had 3D information, we used 2D EDI-OCT images to measure the depths of the LC and the LC insertion. This is because the measurements from the original 2D images might have been more accurate than those from the 3D images derived from 2D images. The Bruch's membrane edges, which were used as a reference point for LC depth measurements in this study, were aligned in one plane or were more posteriorly located in the superior and inferior areas than in the nasal and temporal areas in some eyes. This finding increases the significance of our results (horizontal central ridge of the LC and more posterior laminar insertion in the superior and inferior than in the nasal and temporal areas). To measure LC depth, we used the same principle used in previous reports (shortest distance from each delineated point on the anterior laminar surface to the line connecting the two Bruch's membrane edges). ^11,20 LC depth was measured at discrete points in these reports. ^11,20 To represent the LC depths from an infinite number of points on the anterior laminar surface, the mean LC depth was calculated by dividing the area S by the length D (Fig. 1B) in our study. However, our LC depth data do not represent the LC depth of all the points on the entire anterior laminar surface because we measured them in discrete horizontal scans (11 scans). The measurement points of the LC insertion depth were not equally spaced along the laminar circumference, as shown in Figure 1C. However, they were plotted in Figure 3B and Figure 4N as if they had been equally spaced because the relative distances between adjacent points varied among eyes. For example, in eyes with horizontally oval LC, this distance is greater in the superior and inferior areas than in the nasal and temporal areas, and vice versa. In addition, this distance is greater between laminar insertion points farther from the horizontal and vertical midlines. Because the OCT scans were directly exported from the OCT device and then uploaded to the 3D reconstruction software, our 3D results (Fig. 5) depended on the ability of SD-OCT image alignment/registration software. In addition, our 3D images (Fig. 5) contain interpolated information between each scan (distance between scans, approximately 30 μm). Lastly, our results depended solely on the EDI-OCT findings, which may be different from those of histologic examination or other imaging modalities. 

In conclusion, the LC depth profile and 3D LC images obtained using in vivo EDI-OCT demonstrated that there is a horizontal central ridge in the LC and that the LC insertion was more posteriorly located in the superior and inferior than in the nasal and temporal regions. These regional differences in anterior laminar surface concur with previously reported regional differences in susceptibility to glaucomatous damage. Further work is needed to determine whether this association is causal. 

Footnotes

 Presented at the annual meeting of the Association for Research in Vision and Ophthalmology, Fort Lauderdale, Florida, May 2011.

Footnotes

 Supported by a grant from The Gertrude Josephine Bennett Family Foundation (New York Glaucoma Research Institute).

Footnotes

 Disclosure: S.C. Park, None; S. Kiumehr, None; C.C. Teng, None; C. Tello, None; J.M. Liebmann, Carl Zeiss Meditec (F), Heidelberg Engineering (F), Optovue, Inc. (C), Topcon Medical Systems (C); R. Ritch, None