Region of lamina (measurements in p) Age (years) Posterior Mid posterior Mid- anterior Anterior 9 5.33 4.98 4.98 4.51 25 7.03 6.41 3.59 3.23 26 8.15 4.59 5.46 5.29 40 7.16 5.42 3.74 3.62 45 7.06 5.08 4.75 4.57 56 7.73 5.59 5.29 4.03 60 6.12 7.31 4.57 3.97 72 9.69 7.81 7.31 6.34 76 7.5 7.2 6.69 6.41 83 7.33 7.7 6 6.31 84 6.72 5.01 4.07 4.27 90 8.72 8.18 6.74 6.09
Average thickness in microns of cribrosal plates in each of the areas as defined in figure 6.2. The cribrosal plates are thickest at the posterior lamina cribrosa region, and increase in thickness with age (linear regression, r^ = 0.42, p<0.05).
6.1.4 Discussion
The purpose of this study was to investigate the longitudinal architecture of the lamina cribrosa to see if it offered any clues as to the limitations of our in vivo
imaging technique.
Previous measurements of the thickness of the lamina area in fixed tissue have shown it to be of the order of 0.237mm (Quigley, Hohman et al. 1983).
Experimental studies by Yan et al have shown it to range in thickness from 0.1- 0.140mm under different pressure loads (Yan, Coloma et al. 1994). Our results using fresh, unfixed specimens indicate that, in conditions that more closely resemble the in vivo situation, the region is somewhat thicker. In the earlier study of lamina thickness (Quigley, Hohman et al. 1983), the tissue was investigated using scanning electron microscopy techniques. This involves the digestion and dehydration of tissue during its preparation. It has been shown that removal of axons from lamina cribrosa samples can induce shrinkage and distortion of the extracellular matrix of the region, and that up to 30% shrinkage in tissue occurs during preparation procedures for scanning electron microscopic techniques (Birch, Brotchie et al. 1997). Our findings of thickness approximately 50% greater (fig 6.4) would be expected due to the use of fresh material. The lamina area can be divided into two distinct parts, the prominent posterior lamina area, and the less densely packed anterior area (Radius and Gonzales 1981; Bron, Tripathi et al. 1997). It is possible that under ex vivo experimental conditions, as in those in Yan et al’s study, the anterior area becomes less prominent, leading to an
underestimation of the total lamina thickness.
W e also found that the thickness of the lamina cribrosa increased with age, which agrees with most other studies looking at the change in collagen content. Albon et al reported a finding of an increase in the total collagen content of the lamina cribrosa to change from 20% in the young to over 50% in the elderly (Albon, Karwatowski et al. 1995). Morrison et al (Morrison, Jerdan et al. 1989) showed that collagen accumulates after birth and increases throughout life. Hernandez et al found similar changes in the macromolecular content of the optic nerve head with age (Hernandez, Luo et al. 1989), finding that in the ageing
lamina cribrosa there was an increase in the fibrillar forms of collagen in the core of the cribriform plates. Ogden et al (Ogden, Duggan et al. 1988) also found an increased area of connective tissue within the area with progressive age.
It has been observed that the appearance of the lamina cribrosa differs across its thickness, and that the trabeculae, or beams of the lamina cribrosa are less dense in the ‘choroidal’ (or anterior) lamina than the ‘scleral’ (or posterior) lamina (Radius and Gonzales 1981). W e also observed a change in the degree of compactness of the cribriform plates across the lamina cribrosa, with a more regular arrangement of plates in the posterior regions compared to that found in the anterior region. Figure 6.6 shows that the thickness of the individual cribrosal plates increase with age, and that this rate of increase does not differ regionally i.e. increase is uniform in the posterior and anterior regions. No previous
measures of cribrosal plate thickness are available to compare our findings, and this is the first time that measurements of the longitudinal lamina cribrosa area have been performed on hydrated tissue.
This study shows that the thickness of the lamina cribrosa in conditions that closely mimic in vivo conditions is greater than previously estimated in
dehydrated tissue. It is essential to know the three-dimensional structure of the lamina cribrosa area in order to help us with our understanding of the mechanism of glaucomatous damage. Bellezza et al have published preliminary work
modelling the optic nerve head as a biomechanical structure, using finite element modelling to estimate the stress caused by intraocular pressure(Bellezza, Hart et al. 2000). It may be important to not only know the diameter of the scleral
opening, but the longitudinal thickness of the lamina cribrosa in further modelling techniques. In addition, in these conditions we have found that cribrosal plates are thicker in the posterior portion of the lamina cribrosa. This is of importance in
in vivo studies of the lamina cribrosa area, and may also be of importance in development of models to study the effects of mechanical forces on the area to help with our understanding of the mode of glaucomatous damage.
For the purposes of this project, knowing that the longitudinal lamina thickness is of the order of 450 microns upwards helps with our understanding of the quality of images obtained. The FWHM of the Zeiss cSLO system is of the order of 360 microns (section 2.1.2). This means that the ‘confocal’ slices of the lamina area actually encompass the whole of the lamina cribrosa, possibly including some retrobulbar material as well. The images will, therefore, only be of the surface pore structure, and will be unable to go any deeper without optical modifications to the cSLO. These modifications have been used previously to image the retinal cone mosaic (Fitzke, Woon et al. 1991; Wade 1998). A micro-scanning adapter (MSA) can be attached to the cSLO to reduce the scanning angle of the laser and increase its magnification. This can reduce the FWHM to approximately 30 microns (through a 7mm pupil) (Wade 1998).
Attempts were made in this project to use such an adaptor to image the optic disc in a human subject. However, it was virtually impossible to gain any image data. It was found that using the MSA, only a small proportion of the optic disc would be imaged at any one time (2 degrees visual angle at the retina). For imaging the retinal cone mosaic, the subject was able to fixate directly on the laser imaging light source, (as that was the area that was being imaged). However, for optic disc imaging, off-axis fixation is required. The influence of microsaccadic eye movements is magnified when using the MSA, so only a fleeting glimpse of the optic disc was achieved at any one time. Also, in the images that were gained, it was impossible to establish which area of the disc was actually being imaged. Although a vast improvement in the confocal width of the cSLO is possible with the MSA, it is not feasible to use for imaging the optic disc.
As mentioned in section 1.5.2.4, OCT is becoming an increasingly popular mode of imaging due to the substantial improvement in resolution (both lateral and longitudinal) of the instrumentation compared with the cSLO. Commercially available instrumentation is able to give ‘B scan’ like images of the retina (figure 6.7). However, although research is being conducted into producing en face
increase in resolution of the instrument does not appear in the en face images due to difficulties in imaging the tilted/ spherical retina (Rogers J.A., Podoleanu A. Gh. et al. 2001). Adequate lamina cribrosa images have not been obtained yet using the current en face set up (figure 6.8). The results shown in this project show far superior in vivo lamina cribrosa images using the cSLO.