Faculty Bibliography

Fluorescence-mediated photoplethysmography (FM-PPG) is the first routine clinical methodology by which to quantifiably measure tissue blood perfusion in absolute terms (mL blood/sec ∗ mm² tissue). The FM-PPG methodology has been described in detail previously in this journal (MVR 114, 2017, 92-100), along with initial proof-of-concept measurements of blood perfusion in both ocular and forearm skin tissues. The motivation for the current study was to investigate whether FM-PPG can be used readily and routinely under realistic clinical conditions. The vehicle for doing this was to measure medial foot capillary blood flow, i.e., tissue perfusion, in 7 normal subjects, mean = 6.76 ± 2.29 E-005 mL/(sec ∙ mm²), and lesion-free areas of 8 type-2 diabetic patients with skin ulceration, mean = 4.67 + 3.15 E-005 mL/(sec ∙ mm²). Thus, perfusion in the diabetics was found to be moderately lower than that in the normal control subjects. Earlier skin perfusion measurements in medial forearms of 4 normal subjects, mean = 2.64 + 0.22 E-005 mL/(sec ∙ mm²), were lower than both the normal and diabetic foot perfusion measurements. Variability in the heartbeat-to-heartbeat blood perfusion pulses in the skin capillaries, defined as the ratio of the standard deviation among beat-to-beat pulses divided by the mean perfusion of those pulses, was determined for each subject. Average variability in foot skin was 21% in the diabetic population, versus 16% for normal subjects; and it was 18% in forearm skin. We conclude that absolute quantitative FM-PPG measurement of skin blood perfusion at the level of nutritive capillaries is feasible routinely under clinical conditions, allowing for quantitative measurement of skin tissue blood perfusion in absolute terms.

Fluorescence-mediated photoplethysmography (FM-PPG) was developed to facilitate determination of tissue microvascular (capillary) blood perfusion, requiring intra-venous injection of a small bolus (~0.33mL) of dye (25mg/mL ICG) and acquisition of high-speed angiogram images (>15images/s). The methodology is applicable to microvascular capillary beds of tissues that can be optically imaged, directly or laparoscopically. Proof of concept of the methodology and feasibility of its implementation are demonstrated in human forearm skin and in the choroid of a monkey eye, determined to have blood perfusion rates on the order of 1e-5mL/(s.mm2) and 10e-5mL/(s.mm2), respectively. Ability to obtain absolute quantified tissue perfusion data ultimately can provide the means by which to characterize blood flow information at the nutritive capillary-vessel level in an objective and universally understood manner, in much the same sense that body temperature or blood pressure are.

A clinical method for characterizing the state of micro-vasculature vasomotion is demonstrated, based on observing in capillaries the dynamics of autologously re-injected erythrocytes containing ICG dye. Since a manifestation of vasomotion is transient erythrocyte pausing, vasomotion state within a field of capillaries is characterized by an histogram plot of the number of paused erythrocytes as a function of pause duration during a fixed period of observation, then the ratio of long-pausing to short-pausing erythrocytes was calculated. The method was first applied to the posterior pole retinal vasculatures of anesthetized-monkey eyes, and normal vasomotion state during air-breathing was compared to the state induced by O2-breathing, known to cause mild arteriolar vasoconstriction in the mature eye. Subsequently, the effects of other antagonists to normal arteriolarvasotonia state (long-standing experimentally-induced ocular hypertension and branch-vein occlusion, as well as tissue edema) were similarly characterized and the results compared to those obtained during baseline air-breathing. The feasibility of applying the histogram characterization of vasomotion state to human eyes and skin was also preliminarily explored.

This technical note describes a modification to a fundus camera that permits simultaneous recording of pattern electroretinograms (pERGs) and pattern visual evoked potentials (pVEPs). The modification consists of placing an organic light-emitting diode (OLED) in the split-viewer pathway of a fundus camera, in a plane conjugate to the subject's pupil. In this way, a focused image of the OLED can be delivered to a precisely known location on the retina. The advantage of using an OLED is that it can achieve high luminance while maintaining high contrast, and with minimal degradation over time. This system is particularly useful for animal studies, especially when precise retinal positioning is required

PURPOSE: To find evidence of retinal vasomotion and to examine the relationship between erythrocyte dynamics and previously observed high-frequency pulsatile blood flow through the choriocapillaris. METHODS: An osmotic shock technique was used to encapsulate indocyanine green (ICG) dye in erythrocyte ghost cells at a concentration that produced maximum cell fluorescence. By obviating the plasma staining that results from aqueous ICG's high affinity for plasma proteins, high contrast was maintained between reinjected ICG-loaded erythrocytes and their plasma background. High-speed, high-magnification ICG angiograms showing individual cell movement were recorded from the intact eyes of four monkeys and three rabbits for periods up to 30 seconds. RESULTS: In monkey retinal perifoveal capillaries, numerous erythrocytes were seen to pause for as long as 20 seconds and then resume transit. Similar pausing behavior was observed in the subfoveal choriocapillaris. Individual erythrocytes also were seen to pause in the rabbits' choriocapillaries below the medullary rays, where visualization of the cells was uninhibited by overlying retinal vasculature or dense pigment. CONCLUSIONS: Reinjected ICG-loaded erythrocytes permit routine visualization of retinal capillary and choriocapillaris hemodynamics of the intact eye. It is speculated that erythrocyte-pausing in both microcirculations facilitates metabolic exchange across capillary walls. In retinal capillaries, pausing is presumed to result from vasomotion-which has been postulated as necessary for the inhibition of retinal edema-and in choriocapillaries, to result from the shifting distributions of local perfusion pressures within the network of capillary vessel segments that comprise each lobular area of the choriocapillaris vascular plexus

Laser photocoagulation of the feeder vessels of age-related macula degeneration-related choroidal neovascularization (CNV) membranes is a compelling treatment modality, one important reason being that the treatment site is removed from the fovea in cases of sub- or juxtafoveal CNV. To enhance the energy absorption in a target feeder vessel, an indocyanine green dye bolus is injected intravenously, and the 805 nm wavelength diode laser beam is applied when the dye bolus transits the feeder vessel; this tends to reduce concomitant damage to adjacent tissue. A 3D theoretical simulation, using the Pennes bioheat equation, was performed to study the temperature distribution in the choroidal feeder vessel and its vicinity during laser photocoagulation. The results indicate that temperature elevation in the target feeder vessel increases by 20% in dye-enhanced photocoagulation, compared to just photocoagulation alone. The dye bolus not only increases the laser energy absorption in the feeder vessel but also shifts the epicenter of maximum temperature away from the sensitive sensory retina and retinal pigment epithelial layers and toward the feeder vessel. Two dominant factors in temperature elevation of the feeder vessel are location of the feeder vessel and blood flow velocity through it. Feeder vessel temperature elevation becomes smaller as distance between it and the choriocapillaris layer increases. The cooling effect of blood flow through the feeder vessel can reduce the temperature elevation by up to 21% of the maximum that could be produced. Calculations were also performed to examine the effect of the size of the laser spot. To achieve the same temperature elevation in the feeder vessel when the laser spot diameter is doubled, the laser power level has to be increased by only 60%. In addition, our results have suggested that more studies are needed to measure the constants in the Arrhenius integral for assessing thermal damage in various tissues

PURPOSE: To review and update techniques of posterior segment ophthalmic fundus imaging. DESIGN: Literature review and collective experience of the authors. METHODS: This perspective includes sections on digital imaging, fundus autofluorescence, ultrasonography, angiography, and optical coherence tomography (OCT) -ophthalmoscopy written by leading clinicians and researchers in these areas. RESULTS: Digital angiography has become the new standard in the ophthalmic community based upon improved technology which has enhanced resolution, processing time, and ease of image duplication, manipulation, and transmission. A relatively new imaging technique, fundus autofluorescence, highlights lipofuscin deposits and improves our understanding of the metabolic status of the retinal pigment epithelium. Diagnostic ultrasonography continues to be a major adjunct to ocular evaluation where advances now allow for exceptional versatility and portability. High speed angiographic techniques provide detailed visualization of choroidal perfusion which improves our understanding of both normal and pathologic vascular phenomenon. Advances in high-resolution OCT currently under development promise an even more detailed fundus representation. The integration of the scanning laser ophthalmoscope and OCT has produced a dynamic new instrument, the OCT ophthalmoscope, which simultaneously images the fundus in numerous ways with point to point correlation. CONCLUSIONS: Ophthalmic imaging technology has revolutionized fundus examination. Currently available systems have contributed significantly to our understanding of the pathophysiology and treatment of various retinal diseases. Future advances promise near histologic resolution of retinal structures as well as real-time image manipulation and instantaneous transmission world-wide

PURPOSE. Anterior ischemic optic neuropathy (AION) is caused by sudden loss of vascular supply to retinal ganglion cell (RGC) axons in the anterior portion of the optic nerve and is a major cause of optic nerve dysfunction. There has been no easily obtainable animal model of this disorder. The current study was conducted to design a novel model of rodent AION (rAION), to enable more detailed study of this disease. METHODS. A novel rodent photoembolic stroke model was developed that is directly analogous to human AION. Using histologic, electrophysiological, molecular- and cell biological methods, the early changes associated with isolated RGC axonal ischemia were characterized. RESULTS. Functional (electrophysiological) changes occurred in RGCs within 1 day after rAION, with a loss of visual evoked potential (VEP) amplitude that persisted in the long term. The retinal gene expression pattern rapidly changed after rAION induction, with an early (<1 day) initial induction of c-Fos mRNA, and loss of RGC-specific gene expression. RGC-specific protein expression declined 2 days after detectable mRNA level changes, and immunostaining suggested that multiple retinal layers react to isolated RGC axonal ischemia. CONCLUSIONS. rAION rapidly results in electrophysiological and histologic changes similar to clinical AION, with reactive responses in primary and supporting neuronal cell layers. The rAION model can enable a detailed analysis of the individual retinal and optic nerve changes that occur after optic nerve stroke, which may be useful in determining possible therapeutic interventions for this disorder

PURPOSE: To optimize the method of treating choroidal neovascularization (CNV) associated with age-related macular degeneration (AMD). DESIGN: Experimental study and interventional case series. METHODS: The parameters associated with locating and then photocoagulating CNV feeder vessels were identified and optimized using published data and data derived from modeling the choroidal vasculature. Based on these optimized parameters, a prototype diagnostic/treatment system was designed that captures high-speed indocyanine green (ICG) angiogram images and facilitates analysis of the images by enhancing visualization of dye movement through CNV feeder vessels (FVs). The system also permits precise aiming and delivery of 810- nm wavelength photocoagulation laser energy to target FVs on a real-time ICG angiogram image of the choroidal vasculature. Target FVs are tracked by a joy-stick controlled laser aiming beam until an intravenously-injected high-concentration ICG dye bolus is observed to enter the target vessel, at which time the laser is fired. Proof of principle of the combined diagnosis/treatment system design for performing dye-enhanced photocoagulation (DEP) in the clinical setting and determination of the minimum DEP laser energy needed to close CNV FVs was made in 11 AMD patients requiring treatment of CNV, but for whom other treatment was not appropriate. RESULTS: Using ICG-DEP, CNV feeder vessels were closed with single pulse laser energy, delivering as little as 0.6 to 1.8 J of energy to the fundus, producing no visible change in the fundus. Successful FV closure was usually indicated immediately by presence of incarcerated ICG dye in the vessel adjacent to the burn site. The prototype system proved relatively easy to operate. After acquiring and interpreting diagnostic angiograms and repositioning a patient in front of the device, feeder vessel DEP and treatment evaluation required 15 to 20 minutes. CONCLUSIONS: Indocyanine green dye-enhanced photocoagulation of CNV feeder vessels, facilitated by use of a device that permits real-time visualization of the choroidal circulation while aiming the treatment laser beam, appears to minimize the amount of energy applied to the fundus and the volume of fundus tissue affected by treatment, compared with other treatment modalities. The combination diagnosis/treatment device should be useful in optimizing FV treatment and in refining and evaluating the efficacy of DEP in future clinical trials

PURPOSE: To report clinical observations consistent with conclusions from a previous theoretical investigation indicating that photocoagulation of choroidal neovascularization (CNV) efferent vessels can be, in some instances, an effective treatment.DESIGN: Interventional case series.METHODS: In five eyes with age-related macular degeneration (five patients with mean age +/- SD of 65 +/- 11 years, range 55-79 years) requiring treatment of CNV. In each case, the appropriate treatment was location and photocoagulation of the CNV efferent vessels, since the afferent vessels were not identifiable or were located beneath the fovea. Targeted vessels were determined to be draining vessels by analysis of pretreatment high-speed indocyanine green (ICG) angiograms, and successful vessel closure by photocoagulation was demonstrated by posttreatment ICG angiograms. The eyes subsequently were followed from 2 to 12 months.RESULTS: After photocoagulation of efferent vessels, CNV-related retinal edema was significantly reduced or resolved within 1 to 4 days. Visual acuity became stabilized in three eyes and improved in two eyes. In a few days, metamorphopsia disappeared in four of the eyes and was stable for a period longer than the duration of the associated efferent vessel closure. Initial efferent vessel closure by photocoagulation persisted on average for 7 to 15 days, after which additional treatment was required. It is significant that in no case did hemorrhage result from the photocoagulation treatment.CONCLUSIONS: These observations are consistent with the earlier theoretical study prediction that photocoagulation of efferent CNV vessels can be effective in reducing CNV-associated edema. That no hemorrhage was induced by photocoagulation is consistent with the theoretical concept that there appears to be no direct hydrostatic connection between the CNV and its afferent vessels. Rather, that connection appears to be a functional one made through the choriocapillaris, which may dissipate excess CNV hydrostatic pressure produced by occlusion of a draining vessel. However, this finding is not intended to be a recommendation to attempt CNV efferent vessel photocoagulation