Albrecht v. Graefes Arch. klin. exp. Ophthal. 189, 159--164, (1974) �9 by Springer-Verlag 1974

A Comparison of Retinal Argon Laser Lesions in Man and in Cynomolgus Monkey* * *

I. H. L. Wallow, 0 . -E. Lund, and V.-P. Gabel

Augenklinik der Universitiit Miinehen (Direktor: Prof. Dr. 0.-E. Lund), 3{finehen

R. Birngruber and F. t I i l l enkamp

Gesellschaft fiir Strahlen- und Umweltforsehung m.b. tI., Neuherberg h. M/inchen

Received November 9, 1973

Summary. Suprathreshold argon laser lesions produced under comparable experimental conditions in man and in cynomolgus monkey showed the same degree of retinal destruction and approximately the same width of retinal damage. Four times the power necessary to produce an ophthalmoscopic threshold lesion was insufficient to inflict damage to the inner retinal half. In therapeutic argon laser photocoagulations, for instance in the treatment of diabetic microaneurysms, this power level should, therefore, be exceeded.

Zusammen/assung. ArgonlaserlEsionen oberhalb der ophthalmoskopischen Schwelle wurden unter vergleichbaren experimentellen Bedingungen in der Netz- haut eines menschlichen VoluntErs sowie eines Cynomolgusaffen produziert. Die L~sionen zeigten bei histopathologischer Untersuchung den gleichen Grad yon NetzhautzerstSrung und nngefiihr die gleiehe Schadensausdehnung. Selbst bei einer Laserleistung, die das Vierfache der zur Erzeugung einer ophthalmoskopischen Schwellenl~sion notwendigen Leistung betrug, blieb die iimere NetzhauthElfte un- versehrt. Wenn Argonlaserkoagulationen z.B. zur Behandlung yon diabetisehen Mikroaneurysmen eingesetzt werden, sollte dieses Leistungsniveau fiberschritten werden.

Introduction

Pr imate monkeys like the rhesus monkey (macaca mulatta) and the cynomolgus monkey (macaca iris) have become the most widely used exper imental animals in invest igat ions s tudying the effects of laser i r radiat ion to the ret ina. I t is generally accepted t ha t results found in these monkeys provide useful informat ion as to the s i tuat ion in man. Yet to our knowledge only Vassiliadis et al. (1969) have reported a comparison of re t inal laser lesions produced in both species under the same experi- menta l condit ions using similar laser powers and exposure times.

* Herrn Prof. Dr. H. K. Miiller zum 75. Geburtstag gewidmet. ** This investigation was supported in part by grant NT 188 of the Bundesmini- sterium fiir Forschung und Technologie (vormals Bundesministerium ffir Bildung and Wissenschaft, Bonn).

12 Albrecht v. Graefes Arch. klin. exp. Ophth~l., Bd. 189

160 I . H . L . Wallow et al.

R e c e n t l y we h a v e h a d t h e o p p o r t u n i t y t o p r o d u c e s u c h l e s ions in a

h u m a n v o l u n t e e r su f f e r i ng f r o m a m a l i g n a n t c h o r o i d a l m e l a n o m a , a n d

in e y n o m o l g u s m o n k e y . W h i l e Vass i l i ad i s et al. u s e d o p h t h a l m o s c o p i c

t h r e s h o l d l es ions for t h e i r c o m p a r i s o n , we p r o d u c e d s u p r a t h r e s h o l d les ions .

A h i s t o p a t h o l o g i c e v a l u a t i o n c o m p a r i n g t h e d e g r e e a n d e x t e n s i o n of

r e t i n a l d a m a g e in t h e s e l es ions wil l be p r e s e n t e d in t h i s r e p o r t .

Material and Methods

A 60 years' old emmetropie male Caucasian patient with greyblue irides and a fair fundus pigmentation was admitted to the Eye Hospital of the University of Munich for enueleation of his right eye which contained a malignant ehoroidal melanoma in the superior temporal quadrant, about one disc diameter above the macula. On subsequent histopathologic examinat ion the tumor proved to be a melanoma, spindle cell type B. The pa t ien t gave wri t ten consensus to have the central re t ina of his diseased eye exposed to some laser lesions prior to enucleation of this eye. The experimental arrangement included an argon laser (Spectra Physics, model 165) and is described in some detail below.

In to the ocular fundns of the non-anesthesized pat ient 4 argon laser lesions using a laser power of 60 mw (as determined a t the plane of the cornea) and an exposure t ime of 77 msec were produced nasally and beIow the macula to parts of the ret ina t ha t were not affected by the tumor. Ophthalmoscopically, the lesions appeared as areas of a round whitish retinal discoloration surrounded by one ring of grey-yellow retinal discoloration. 1 day after exposure the eye was enucteated, opened coronally at the pars plana and fixed in 4% buffered glutaraldehyde. Retinal areas containing laser lesions were dissected and embedded in Epon while the remaining globe was processed for routine histopathologic examination.

From the number of lesions which we have produced previously in eynomolgus monkeys (Wallow, 1973), 2 argon laser lesions were chosen for comparison. These had been created in one eye below the macula using a laser power of 70 mw (deter- mined at the level of the cornea) and an exposure t ime 77 reset. Ophthalmoscopically, the lesions appeared similar to those in human, i. e. they presented as areas of a round whitish retinal discoloration surrounded by one ring of grey-yellow retinal discolora- tion. 2 days after exposure the eye was enueleated, fixed and dissected as described above.

The lesions of both , man and monkey, were then cut serially a t 1.5 i~m thick sections. The sections were stained with toluidine blue and examined by light microscopy unt i l the center of each lesion was established. The dimensions of the lesions in the central section and the retinal thickness of the irradiated retinal re- gions were measured using a calibrated graticule.

The abil i ty of our ins t rumentat ion in producing consistent experimental laser lesions of the re t ina has been established previously in systematic investigations of laser threshold lesions in eynomolgus monkeys, and chinchilla rabbi ts (Bi,~agi~ber et al., 1972; Wallow, 1973). An argon laser beam (A~5145 ~_) was guided into a modified Zeiss-slitlamp through a series of optical and mechanical elements. I t left the sli t lamp collinearly with the i l luminat ing light beam and was focused into the plane of observation of the microscope. I r radia t ion and observation were done through a Goldmann-type corneal contact lens. The exposure t ime was determined by an electromagnetic shutter. The laser power was measured by a photodiode a t the entrance of the beam into our modified slit lamp, calibrated to read the power incident upon the contact lens. The details of this ins t rumenta t ion have been pro-

Retinal Laser Lesions in Man and Monkey 161

sented by one of us (Birngruber et al., 1972) and are similar to the arrangement described by L'Esperanee and Kelly (1969). The retinal image size in cynomolgus monkeys using this instrumentation was estimated to be 16 t~m in diameter (not taking into account intraocular scatter). In man, a retinal image size of 18 ~zm in diameter was calculated. The total error of the system in calculating the laser power density incident upon the retina was ~-40 %.

Results

The ret inal thickness in the 4 i r radiated fundus regions in m a n was 290, 310, 280, 240 ~xm. I n the 2 re t inal areas of cynomolgus monkey it was 275 and 230 ~zm.

tI istopathologically, in both species all lesions showed marked de- s t ruct ion of the ehoriocapillaris and dest ruct ion of She outer half of the re t ina up to the external plexiform layer. Brueh's membrane remained in tac t (Fig. 1A, B). I n the ehoriocapillaris, there were occlusion of numerous blood vessels, ex t ravasa t ion of f ibrin and in t ravasa l as well as extravasal accumula t ion of large irregular-shaped mononuelear cells.

I n the retina, the cells of the ret inal p igment epi thel ium were vaeu- olated in the periphery of the lesions. Centrally, the p igment epithelial cells were condensed. Their nuclei were no t recognized. The outer seg- ments of the photoreeeptor cells were par t ia l ly fragmented. I n other areas, they had a stretched appearance with dist inct outlines and an almost normal parallel orientat ion. The inner segments were shrunken and hyperehromatie . There was pycnosis and karyolysis of the nuclei of photoreceptor cells wi thin the external nuclear layer. Numerous axons of photoreeeptor cells extending to the middle l imit ing membrane were hyperchromatie . No damage to cells in ternal to the middle l imit ing membrane was noted.

Table 1. Extension of damage and physical parameters of retinal argon laser lesions in a human eye and in a cynomolgus monkey eye

Species Lesion Age Extension of damage Laser Exposure No. (days) in ~m at level of power duration

(row) (reset) RPE a ENL b

Man 1 1 190 140 60 77 2 1 170 150 60 77 3 1 160 120 60 77 4 1 150 100 60 77

Cynomolgus 5 2 200 160 70 77 6 2 180 140 70 77

a RPE = retinal pigment epithelium; b ENL = external nuclear layer.

1'2"

Fig. 1. Light mierographs showing argon laser lesion of the perimaeular ret ina of man (A) and of eynomolgus monkey (B). The lesion in A is 1 day old and was produced with 60 mw at 77 msee. The lesion in B is 2 days old and was produced with 70 mw at 77 msee. A and B Within the lesions, the retinal pigment epithelium (RPE) and the photoreeeptor cells including outer segments (os) inner segments (is), nuclei in the external nuclear layer (enl), and axons in the "external plexiform layer" (epl) are partially vaeuolated, partially condensed and hyperehromatie. No damage to the inner nuclear layer (in[) is present. B M Brueh's membrane; chc ehorioeapillaris. [n A. damaged photoreceptor cell axons which leave the maeula, are cut obliquely.

(1.5 tzm Epon section, toluidine blue, A and B • 420)

Retinal Laser Lesions in Man and Monkey 163

In both species, the extension of the retinal damage was wider at the level of the retinal pigment epithelium as compared to the level of the external nuclear layer. The widths as measured in the central section through each lesion are given in Table 1.

Comment

Retinal argon laser lesions above the ophthalmoscopic threshold (subsequently also referred to as suprathreshold lesions) were produced in man and in cynomolgus monkey (macaca iris), the closest relative of rhesus monkey (macaca mulatta), to the perimacular fundus under identical experimental conditions using a similar laser power (60 and 70 row, respectively, determined at the plane of the cornea) and the same exposure time (77 msec). The retinal image size in man was calculated to be 18 ~z in diameter, in cynomolgus monkey ist was estimated to be t6 ~z in diameter. Histopathologically, the lesions showed the same con- figuration of damage, i.e. they all involved the retinal pigment epithelium (RPE) and the photoreceptor cells including their nuclei within the external nuclear layer (ENL) and their axons. The width of retinal de- struction as measured at the RPE-level and at the ENL-level was not significantly different between man and cynomolgus monkey. These data though very limited as yet are taken as an indication tha t a difference in fundus sensitivity between the two species, if there is any, is rather small at least for minimal spot sizes relatively low laser powers, and exposure durations within the range of the blinking reflex. This result is compatible with observations by Vassiliadis et al. (1969) made in two human volun- teers and in rhesus monkey under somewhat different experimental conditions. More data based on human volunteers with an intact ocular'

fundus at the posterior pole are required to arrive at more definite con-

clusions.

When compared to ophthalmoscopic threshold lesions which we have produced in cynomolgus monkeys for other purposes (Birngruber et al., 1972), the total power for the suprathreshold lesions described in this repoIt was approximately four times higher. The ophthalmoscopic threshold lesions typically showed a destruction of the outer retinal half, extending over an area of approximately 60-80 ~zm in diameter at the RPE-level and of 30-40 ~zm in diameter at the ENL-level (Wallow et al., 1973, in press). Interestingly, the present suprathreshold lesions exhibited the same pat tern of damage and differed merely in width of retinal damage (approximately 190 ~zm at the RPE-level and 150 ~zm at the ENL-leve]) ; the inner retinal half internal to the middle limiting mem- brane still remained uninvolved.

I t is tempting to apply the ratio of 4 to the relationship between ophthalmoscopic threshold lesions and suprathreshold lesions in man.

164 I . H . L . Wallow et al.

I n man, the argon laser is now f requent ly employed in the t h e r a p y of re t ina l diseases, for ins tance in order to des t roy leaking microaneurysms in background d iabet ic r e t inopa thy . These aneurysms are a lmost a lways loca ted wi th in the inner re t ina l half wi th only few aneurysms bulging o u t w a r d into the ou te r p lexi form layer. Threshold lesions jus t visible ophtha lmoscopica l ly , as well as lesions p roduced wi th a laser power four t imes above t h a t for such threshold lesions would then no t suffice to des t roy microaneurysms.

Re~erenees

Birngruber, R., Wallow, I. H. L., Gabel, V.-P., Hillenkamp, F., Schmidt, R. : Zur Bestimmung der Seh~digungsschwellenwerte der Netzhaut durch Laserstrahlen. Gespr~chskreis: Laseranwendung in der Ophthalmologie anl~Blich der 72. Ta- gung der Deutschen Ophthalmologischen GesellschMt in Hamburg, 23.9. 1972

L'Esperance, F. A., Kelly, G. R. : The threshold of the retina to damage by argon laser radiation. Arch. Ophthal. 81, 583-588 (1969)

Vassiliadis, A., Pmsan, R. C., Zweng, H. C.: Research on ocular laser threshold. Stanford Research Institute, )~enlo Park, Calif.; Final Report, August 1969

Wallow, I. H. L. : Netzhautreaktionen nach Intensivlichtbestrahlung. Experimen- telle, morphologische und klinische Untersuchungen fiber pathologische und therapeutische Wirkungen yon Laserlicht und yon weii~em Licht. Habilitations- schrift (medical thesis), Med. Fakult~t d. Ludwig-Maximilians-Universit~t, Mfinchen, 1973

Wallow, I. H.L. , Gabel, V.-P., Birngruber, R., Hillenkamp, F. : Klinische und histologische Untersuchungen nach Argonlasereinwirkung auf die ~Tetzhaut.-- Die histopathologische Auswertung yon Laserl~sionen als Mittel zur Ab- schi~tzung einer funktionellen Laserschi~digungsschwelle. Ber. dtsch, ophthal. Ges. 73 (1974), in press

Dr. Ingolf H. L. Wallow Univ.-Augenklinik D-8000 Mfinchen 2 Mathildenstral3e 8 Federal Republic of Germany