Dr. Lujan: I wanted to talk about spectral-domain OCT and some interesting observations that we’ve made clinically. The retina appealed to me and to most retinal specialists partially because of its layered anatomy that is present from early drawings from (unintelligible) to more recent histological images that really display the retina in exquisite detail. And, certainly that’s what initially appealed to us with OCT, this ability to see the retina and to see its different layers and to learn about it

Now, obviously OCT has evolved over the last several years from its original commercial incantations as Stratus OCT into spectral-domain OCT systems such as Cirrus and more recent software innovations to improve images, reduce noise and generate very beautiful cross-sectional images using frame averaging and other software manipulations

But, the basics of OCT remain the same from its initial inception to now, and that there’s a source of light that is entering through the pupil and is reflecting off of the different layers of the retina. And, as that light reflects off of different surfaces, the amount of light returning will be dependent upon the optical properties of the retinal tissue, and the amount of time it takes between light returning from one part of the retina to the other part of the retina allows for the reconstruction of the cross-sectional images that we see

This difference is too fast to measure directly, so it’s interferometry and the brilliance of OCT and how to relate that difference into an image that we’re able to see, but there really is a difference in time

When you’re looking at the Cirrus acquisition screen, you can see that there’s multiple different visualizations. I draw your attention to the one in the upper left. It’s showing you the pupil position. This shows us exactly where light is entering the pupil during the scan. In real time, you’re getting a cross-sectional V scan [sp] showing us what you’re seeing entering through that pupil position

As operators of OCT devices have learned, that appearance of that cross-section will vary depending on where through the pupil the scan is entered, and if you go through one position, the V scan will appear tilted. If you go through another position, it will appear to tilt in a different direction, and it’s one of the things that the operators are doing, is being aware of this and trying to acquire an image usually in a flat position

It’s important to understand why optically this occurs, and that is related to the time it takes for light to enter through the pupil and bounce back. So the appearance of a flat V scan, of a cross-sectional scan occurs when the path length from one side of what’s scanned in the retina to the other side of what’s scanned is equal and you generate a--what’s termed a flat cross-sectional image, a relatively flat image where one side looks to be about the same as the other

However, when you’re entering through the side of the pupil, this doesn’t appear flat anymore. It actually appears tilted on the screen, as I showed you. The reason for that is that these path lengths are now different. It takes a shorter amount of time for light to travel down path A all the way on one side than it does to path B

Now, obviously the back of the eye, the retina, hasn’t changed, but the software is interpreting the amount of time it takes for light to come back as greater distance and is rendering a tilted cross-sectional image

So, that--but that’s--it’s easy to understand why that’s occurring just from this optical--or just from an understanding of how the software and OCT technology works

What’s interesting, though, is that there’s more than just this apparent tilt in these images that changes, and if you go through the center of the eye--you see in this image it appears quite flat. When you go in through the side, again you get this tilted image. This is the same eye, just taken a few seconds later, a few seconds between scans

Then, if go to the other side, you get this tilt going the other way, and what you’ll notice is that a certain part of the retina very obviously looks different depending on these two images. So, you see on the top image, just focusing on the left side of what you see, a very bright, hyper-reflective band, whereas just moving the position of the entry, that goes away and actually becomes dark

The right side of the image applies the exact same phenomenon. This alternates between dark and light

So, this is interesting. Why is this occurring optically? Well, if we look at this exact part of the retina where this difference occurs, alternating between dark and light now in this registered image--this is the same retina. Nothing has changed. If we try to understand why that is, we need to look to the histology

So, in this beautiful image published by Curseo [sp] in 2011, we can see what is present at that layer where there’s variable, and that structure is Henle’s fiber layer

Henle’s fiber layer is the axons of the photoreceptor nuclei that are located lower--where it denotes the optical nuclear layer, the ONL. It’s the axons going from those photoreceptor nuclei up to the next level of cells in the retina. And, as the foveal pit is developing from early in life, the distance between where the photoreceptor nuclei are and where the cells of the inner nuclear layer are increases. There’s a spread of those cells as the foveal pit develops, and the result is that you have these obliquely oriented Henle’s fibers that are radiating out from the center of the retina

If we see them in close detail from electron microscopy, you can see that immediately out of the outer nuclear layer of these photoreceptor nuclei there are these obliquely oriented, tilting axons that are coming out

So, optically, why is this the case? What’s occurring? Well, when we go in from the side of the eye, we’re actually hitting those obliquely oriented fibers flat on, and they’re acting like mirrors and generating a very bright signal, whereas on the opposite side of the eye, light is bouncing off those fibers, but it’s not coming out of the pupil. It’s basically bouncing into the side of the eye and isn’t seen and appears dark, so the entrance path through the pupil actually affects how this layer is visualized

But, this layer may be the first layer that’s obvious, but in our publication in IOVS where I demonstrated this phenomenon, we noticed that this is true in clinical patients as well. So, this is something you’ll see, and now that, you know, you’re aware of this, it becomes very obvious that this occurs all the time

So, in this case, a central serous retinopathy, you see this variability in that layer. That occurs above this pigment epithelial attachment, where the orientation of those fibers has changed relative to the entrance beam, relative to the light coming into the eye

Similarly in drusen, and above these drusen, you have a similar phenomenon, where there’s both lighter bands and darker bands depending on the exact orientation of those fibers with respect to the light that’s coming in

Schematically, this is what’s occurring in this case of central serous retinopathy. Those normally obliquely oriented fibers, as sub-retinal fluid occurs underneath the retina, it’s pushing them to the side and light is reflecting off of them in areas where it’s brighter at such an angle to act like a mirror and to reflect very brightly what is being seen, to reflect very brightly

In areas where it’s dark, the exact same phenomenon as when you’re coming in from one side of the eye occurs, and it actually appears dark simply because of the relative orientation of those fibers to the entry position

So, this is important to be aware of and to note. These are not pathological changes. Early, there was a suspicion that overlying these drusen, that this actually represented inflammatory cells that were coming down to these drusen. While that’s an interesting hypothesis, it’s more likely that this is simply an optical phenomenon of the relative change of these fibers

Importantly, it’s not just Henle’s fiber layer that changes based on the orientation of light coming into the eye. If we look at these registered images taken from different positions, we can see a large variability in the outer retina as well

This asterisk shows you the position of light reflecting off of the inner limiting membrane at the top, and you can see how it shifts, showing that I’ve actually come in from the side, but then I’ve just manipulated these images to show them together. But, you can see that the cells, excuse me, in the outer retina actually change as well, that cells--you can see the brightness shift there as I toggle back and forth between these images

This is important to be aware of and makes us want to consider what the source of these reflections that we see are, what the sources are. So, this layer, the external limiting membrane, actually has a physical source in the dense bodies that are connecting photoreceptors and muller cells. The external limiting membrane does not change based on the directional position because it is truly a reflective tissue. It actually physically has substrate that is reflecting light, and it does not depend on the direction for which light is coming at it

Versus the inner segment/outer segment junction. This junction, going from mitochondria--as you see here, the light is passing through, is reflecting either off of those mitochondria or, more likely, off the outer segment discs that are below it. This is extremely sensitive to changes in position. So, if light is not coming off of it directly perpendicular and reflecting back, you’re not going to visualize this layer as you do when it’s flat

Similarly, the outer segment RPE junction, where the tips of the outer segments are interacting with the RPE, has directional dependence as well, and this layer varies depending on the position of entry through the pupil

So, why is this important? Because if you have a patient with central serous retinopathy who comes in with sub-retinal fluid and a PED, you--if you are trying to say is this patient going to see again, you know, you know that--first of all, you know that they can because they’re 20/20. You know that their photoreceptors are intact

But, when you look to those bands that we’re learning now on spectral-domain OCT are telling us what the state of the photoreceptors are, they don’t appear crisp and distinct. The inner segment/outer segment junction that you see nicely on the left side of the image as it goes into the area of fluid, its orientation relative to the entrance beam changes, and this affects how it’s visualized

Here, this PED being elevated has caused this fluid and is accounting for that change from this inner segment/outer segment junction into the area of disruption

But, we know inherently that these photoreceptors aren’t loss because of the vision and that when the patient comes back later, they’re 20/20 and that line, that returns to normal. So, there’s been no, you know, biological disruption. This is an optical phenomenon, and it’s something we all need to be aware of

Similarly, in a patient with (unintelligible) disease who’s seeing 20/40, we see an OCT image taken head on and we may wonder what’s the status of this patient’s photoreceptors. When we follow that inner segment/outer segment junction, it appears to fade away or to be very low in intensity. However, if we come in from different angles--we can tell what angle I’ve come in at based on the shadows from the retinal vessels, which will always tell you how the scan was acquired. You can see the large difference in the reflectivity of this layer, of the inner segment/outer segment junction

So, if you’re trying to prognosticate if your patient’s going to regain photoreceptors or know what the status of their photoreceptors are, the interpretation of this layer will help you, and it’s something to be aware of that it’ll appear different depending on how you actually acquire the image

So, in summary, with SD OCT, this is not just like a photograph where taking a photo by any photographer is going to result in an image that should be interpreted by the physician. We have to be aware of how the images are being taken actually affects what is being seen and that specific tissues do have retinal directional reflectivity, meaning that the direction in which their imaged affects how they’re visualized

So, this is particularly true in Henle’s fiber layer and in the photoreceptor layers deep to that, and it behooves all of us to be training our imagers, the people actually acquiring these OCT images, to use Cirrus to actually use the camera, the photograph that’s present of the pupil, and the V scan to be aware of imaging horizontally, to be imaging the same way each time or to be aware of the effect that changes in position have with regard to what’s actually being visualized

Thank you very much