Angiocautery of the Aqueous Outflow Channels in the Rabbit Eye⋆

ANGIOCAUTERY OF THE AQUEOUS OUTFLOW CHANNELS IN THE RABBIT EYE* A TONOGRAPHIC STUDY J O S E P H R. GAZALA, M.D., W A L T E R J. GEERAETS, M.D., A N D D U P O N T GUERRY, I I I ,

M.D.

Richmond, Virginia

The discovery of the aqueous veins in the human eye by Ascher1-4 some 20 years ago marked a turning point in the understanding of the ocular aqueous hydrodynamics. Not only did this discovery help establish the validity of the then controversial theory of continuous aqueous circulation versus the stagnation theory but it also set the stage for a host of extremely valuable clinical observations and experimental studies. Goldmann,5"8 Ashton,9"11 Theobald,12"13 Sheppard4^15 and many others have contributed extensively to our present-day knowledge of the anatomic and biomicroscopic characteristics of these vessels in man and in laboratory animals. The results of surgical procedures designed to interrupt the aqueous outflow in laboratory animals have not been uniform. Weekers and Prijot 16-19 destroyed by diathermy all the visible vessels in the perilimbal area and detached the extraocular muscles from their insertions in the rabbit eyes. This resulted in a brief rise of the intraocular pressure, subsequently followed by hypotension. Hugert, 20 on the other hand, coagulated by diathermy the aqueous veins at the limbus of rabbit eyes and ligated the vascular trunks at the posterior pole of the eye and found that the intraocular pressure was elevated for periods ranging from two to 20 days. For obvious reasons the reports on the results of occlusion of the aqueous veins in man are sketchy and rather confusing.

Vail,21 for example, observed an increase in the intraocular pressure consequent to angiocautery by diathermy of the entire limbal area in a patient with severe corneal vascularization. Cyclodialysis became necessary on the eighth day to control the pressure. DeVries,22 on the contrary, argued that occlusive lesions of the aqueous outflow channels should not necessarily lead to glaucoma because of the presence of anastomotic channels which might allow a compensatory dilatation. With these and other contradictory views as a background, it was felt desirable to study the tonographic effects produced by occlusion of the aqueous veins in the rabbit eye. The first question that had to be answered concerned the feasibility of identifying for cauterization the aqueous veins in the anterior segment of the rabbit eye. Several rabbits, albino and chinchilla, were given intravenous pentobarbital sodium (Nembutal) for general anesthesia and studied carefully under the slitlamp with different magnifications and light intensities. The results of the instillation of adrenalin and dionin drops into the conjunctival sac, as well as those of compression on the globe, were thoroughly investigated. Ink was injected into the anterior chamber with a 25-gauge needle and the change in color of the episcleral outflow channels was observed. The results of this preliminary study may be summarized as follows : 1. The aqueous veins, although not easily seen under normal circumstances, are present in every normal eye. Because of a large amount of blood in these vessels, the

* From the Department of Ophthalmology, Medical College of Virginia. This study was supported by the Old Dominion Eye-Bank and Research, Inc., and Public Health Service research grant FR0001602 from the National Institutes of Health. 247

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J O S E P H R. GAZALA, W A L T E R J. G E E R A E T S A N D D u P O N T G U E R R Y , III

Fig. 1 (Gazala, Geeraets and Guerry). Lamination in an aqueous vein in the perilimbal area at about 6-o'clock position of a rabbit eye. The long black horizontal line is one of the clipped cilia which resisted saline irrigation. Note the confluence of the aqueous vein with a perilimbal venous trunk which ultimately joins the plexus of vessels around the inferior recrus muscle. This photograph was taken with a single-lens Exacta camera through the Zeiss slitlamp ( χ 2 5 . )

aqueous content is masked and can be brought into view only after certain procedures. In the rabbit eye the aqueous veins have, to a large extent, the same characteristics as the blood veins constituting the perilimbal venous plexus. They usually take a very irregular course, concentric with the limbus, dividing several times and joining several blood veins in the 3 to 4-mm zone around the limbus. They ultimately converge toward the upper and lower rectus muscles and disappear into the deeper parts of the orbit (fig. 1). 2. Procedures that help bring into view the aqueous veins either by showing marked lamination or blanching of the vessels (glass-rod phenomenon) include the instillation of vasoconstrictor drops into the conjunctival sac and application of pressure on the globe. The former brings about a substantial reduction in the erythrocyte contents of the vessels of the anterior segment of the globe. The latter causes temporary but marked increase in the aqueous outflow. The end-result in both procedures is a substantial increase in the amount of aqueous over that of the blood in the vessels of the anterior segment of the globe. Thus the aqueous content in these vessels becomes

easily recognizable. Injection of a contrast fluid (Shaefer commercial ink) into the anterior chamber allows drainage of the contrast fluid along with the aqueous, thus making it possible to identify even very small aqueous outflow channels. The results of ink injection into the anterior chamber are in accordance with Sheppard's14"15 work which revealed numerous (up to 90) outflow channels in the rabbit eye (fig. 2). 3. In albino rabbits the aqueous veins are more difficult to see because of poor contrast. The sciera of these rabbits appears pinkish in color due to the absence of pigment in the choroidal layer, while in the chinchilla rabbits, the sciera is bluish white, giving a good contrast to the faded red aqueous veins to be identified and traced. Careful examination showed that the albino rabbits have as many aqueous vessels as the chinchilla. 4. Application of the Hildreth cautery to the proximal end of the aqueous vessel or to what appears to be the emissary opening of the aqueous vein resulted in cessation of aqueous flow and elimination of lamination. The vessel then assumed a dark reddish color because of the undiluted nature of its blood contents. These results made it clear that the rabbit

ANGIOCAUTERY OF THE AQUEOUS CHANNELS

249

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Fig. 2 (Gazala, Geeraets and Guerry). The limbal and perilimbal vessels seen after the injection of ink into the anterior chamber. (Upper left) Photograph represents the 6-o'clock position of the rabbit's limbal and perilimbal area. (Upper right) The area between the 8-o'clock and 9-o'clock positions. (Lower left) The 7- and 8-o'clock positions. (Lower right) The 12-o'clock position.

eye lends itself nicely to a procedure designed to occlude by thermocautery the visible aqueous outflow channels followed by a tonographic study of sequential pressure changes. Furthermore, the study just described demonstrated that aqueous vessels could be identified without instillation of drops or application of pressure on the globe. PROCEDURES

This study consisted of two experiments: 1. In the first experiment, 15 chinchilla rabbits were given Nembutal intravenously through the marginal vein of the ear and tonography was carried out on both eyes. The next morning, again under general anesthesia, the right eye was studied carefully under the Zeiss slitlamp and the aqueous vessels were identified and cauterized at the

limbal end of their course in the episclera. Cauterization was carried out through the conjunctiva and each point was only large and deep enough to occlude the vessel, care being taken that the sciera and the adjacent blood veins were spared (fig. 3 ) . Twenty to 28 such applications, as a rule, were enough to occlude all the visible aqueous veins. T h e process of cauterization took about five minutes. Immediately after cauterization tonographic recording was done on the control, left, eye first. This was to allow sufficient time for the intraocular pressure to build up in the experimental right eye following the occlusion of the outflow channels. Thus, tonography on the right eye, was done about 10 to 15 minutes after cauterization. During tonography on the left eye, every effort was made to prevent pressure on the globe of

250

JOSEPH R. GAZALA, WALTER J. GEERAETS AND DuPONT GUERRY, III

the right eye. Tonography was repeated five and 24 hours later. At the beginning of this study, another recording was made 48 hours postcauterization ; however, this was found unnecessary since the tonographic characteristics by that time were generally normal. 2. The second experiment, done on 15 rabbits, followed a similar procedure except

that cautery applications were placed on the episclera instead of the aqueous veins. The blood and aqueous veins in the perilimbal plexus were avoided (fig. 3). This control experiment was thought necessary in order to eliminate the possibility that the changes in the tonographic findings of the right eye in the first experiment might have been due

Fig. 3 (Gazala, Geeraets and Guerry). (Upper half) The Hildreth cautery applied to occlude the perilimbal vessels, in the first experiment, at the 12-o'clock position. The arrows point to the cautery marks. (Lower half) The Hildreth cautery marks placed away from the perilimbal vessels, in the second experiment, at the upper posterior sector of the globe.

ANGIOCAUTERY OF T H E AQUEOUS CHANNELS

to manipulation of the eye, changes in the scierai rigidity or to dilatation or constriction of the episcleral blood vessels from any type of cauterization and/or irritation of the anterior segment of the globe. All tonographic recordings were done with a V. Mueller tonography apparatus with a built-in electronic tonometer and ink recorder. General anesthesia was given 108 times and a total of 216 tonograms of the right and left eye were obtained. Each tonogram was individually analyzed using Friedenwald (1955) tables. The rate of aqueous flow was calculated according to the classical formula of F = C (Po — 10) ( F being the rate of aqueous flow, C the coefficient of facility of outflow, Po the initial intraocular pressure, 10 the pressure in the episcleral blood veins). The figures for each eye included the initial pressure, the coefficient of facility of aqueous outflow, the rate of aqueous flow and the Po/C ratio (fig. 4 ) . ANALYSIS OF RESULTS

In the right eyes of the first experiment, the most prominent changes were observed in the tonograms obtained immediately following the angiocautery as the intraocular pressure was increased in 14 eyes out of a total of 15. The rise ranged from 5.0 to 22 mm Hg. One eye, however, showed a drop in tension of 9.0 mm Hg. The facility was reduced in 14 eyes and the reduction ranged from 0.02 to 0.37 (mm/min/mm H g ) . In one, it rose by 0.03. In several of these eyes the tonogram was almost a straight line with the C value reaching almost the zero

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251

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level (fig. 5). The Po/C in all 15 eyes was raised above the arbitrary figure of 100 which is found by Becker23 to permit recognition of 70% of proved cases of glaucoma. The lowest figure obtained was 103 ; the highest, 660. The aqueous flow value showed an increase in five eyes, a decrease in nine, and no change in one. It should be recognized that there is only an inadequate analogy between glaucoma cases in man and these experimental conditions. Application of Becker's data should, therefore, be mere speculation. The findings in the control left eyes of the first experiment showed no specific trend in the change of the P,C, Po/C or F values. The pressure, for example, slightly increased in seven and decreased in eight. The same general trend was observed in the C,Po/C and F figures. In the second experiment the right eyes showed a minor increase of tension in the immediate postcauterization reading in 11 eyes, the range of rise was between 1.0 and 6.0 mm Hg only. In three it was lower and the decline ranged from 2.0 to 4.0 mm Hg.

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JOSEPH R. GAZALA, WALTER J. GEERAETS AND ΠυΡΟΝΤ CHERRY, III

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Fig. 6 (Gazala, Geeraets and Guerry). The P o (intraocular pressure) graph. The part of the curves to the left of o represents the mean values of P o obtained from tonographic tracings made the day before cauterization. O marks the time of cauterization. * marks the time of tonography done soon after cauterization. For simplification the graph was interrupted between the fifth and 24th hours after cauterization.

In one eye there was no change. The C value dropped in five and increased in 10 eyes. T h e P o / C and F showed no particular pattern change. In the left eyes of the second experiment, the figures followed no particular trend, thus exhibiting the same pattern as that for the left eyes of the first experiment. Since the results, as just seen, were not uniform at a first approximation, the data obtained from all the tonograms were evaluated by computer analysis. Comparison of the right eyes of the first and second experiments revealed statistically significant differences in the values obtained from the tonograms recorded immediately following the cautery. T h e rise of P o in the first experiment over that of the second was significant at the 0.01 level. ( T h e probability of this difference being due to chance is one out of a hundred.) T h e mean value of P o is 30.66 with standard deviation of 6.779 in the first experiment as compared with the 23.13 and 5.43 respectively in the second experiment. The rise of the P o / C of the first experiment over that of the second was significant at the 0.001 level. ( T h e probability of this

difference being due to chance is one out of a thousand.) The mean value was 300.5 with standard deviation of 12.62 in the first experiment as compared with 73.47 and 32.6 respectively in the second experiment. The decline of the C value of the first experiment below that of the second was significant at the 0.001 level with the mean value of 0.1521 and standard deviation of 0.09 in the first experiment compared with 0.3413 and 0.089, respectively, in the second experiment. T h e values of the rate of flow showed no significant difference between the first and second experiments. T h e values obtained at five and 24 hours after the cautery also revealed no statistical significance. T h e graphic analysis proved interesting. The P o graph (fig. 6) shows a marked rise in tension of the animals' right eyes in the first experiment immediately after cautery with a mean value of about 10 mm H g . On the fifth hour the mean value of tension shows a sharp decline to a level almost equal to that of the precautery value, and below that on the 24th hour. T h e right eyes in the second experiment showed an elevation of a lesser magnitude immediately after the cautery. Five hours later the tension went down below the initial value and, after 24 hours, it rose again to a level almost equal to that of the precautery value. T h e changes in tension of the left eyes in both experiments were within the range of average experimental variations. T h e increase of tension in the right eyes of the first experiment immediately after cauterization is easily explained on the basis of embarrassment by cautery blockage of the aqueous outflow. T h e slight increase in the tension of the right eyes of the second experiment was due presumably to occlusion of some of the outflow channels which were inevitably involved during the process of cauterization or more likely to a reflex vasospasm of the episcleral vessels. I n the graph representing the coefficient of outflow ( C ) (fig. 7 ) , the right eyes of the

ANGIOCAUTERY OF THE AQUEOUS CHANNELS animals in the first experiment showed a sharp decline immediately after the cautery from 0.35 to 0.15. At the fifth and 24th hours it reached a level just below that of the initial value. However, the right eyes of the second experiment revealed some increase immediately after the cautery which reached a higher level five hours and 24 hours later. Variation in the left eyes are insignificant. The changes observed in the C value again do not seem to be too difficult to explain. T h e decline in the C value of the right eyes of the first experiment is due to the blockage of the outflow channels. T h e increase at the fifth and 24th hours is presumably due to a compensatory dilatation and distension of the vessels spared by the cautery. This assumption was based on a clinical observation made by studying the eyes on the second day under the slitlamp at which time quite a number of distended vessels in the areas between the cautery marks were noticed. T h e explanation for the increase in the C value in the right eyes of the second experiment lies probably in the dilatation and distention of the vessels in the perilimbal area produced by the inflammatory process as a result of the cautery in the episclera of the anterior segment of the globe.

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Fig. 8 (Gazala, Geeraets and Guerry). Po/C ratio graph. See the legend of Figure 6 for further explanation. I n the graph representing the P o / C value of the right eyes (fig. 8 ) , of the animals in the first experiment, there is a sharp immediate rise after cauterization u p to a level of about 280. Five hours later it falls to the initial level of less than 100. T h e right eyes of the second experiment and the left eyes of both experiments show no significant change. T h e rise in P o / C ratio in the right eye of the first experiment is an expression of the combined effect of increase in the tension and decrease in the C values. T h e F graph (fig. 9) shows a higher rate of aqueous flow in the right eyes over that of the left eyes in both experiments in all three recordings following the angiocautery. T h e finding of higher aqueous flow in the right eyes is difficult to explain. Theoretically, it might have been caused by a reflex stimulation of aqueous production in the ciliary body induced by the cautery in the episclera. This seems to be the more likely explanation and justifies further experimental studies.

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Fig. 7 (Gazala, Geeraets and Guerry). The C (coefficient of facility of outflow) graph. See the legend of Figure 6 for further explanation.

DISCUSSION

T h e tonographic measurements expressed numerically as values for the P , C, P o / C and F were derived for this study from Friedenwald's 1955 modified tables. This was necessary since, to the best of our knowl-

254

J O S E P H R. GAZALA, W A L T E R J. G E E R A E T S A N D D u P O N T G U E R R Y , I I I

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edge, there are, as yet, no tables available for the rabbit eye based on calculations from studies on this widely used experimental animal. Kornblueth and Linnér24 after an extensive tonographic study on the rabbit eye, reached the following conclusion: ". . . The ocular rigidity of rabbit eyes was fairly close to that of human eyes. The tonographic tracings extending to a steady-state showed that there was a good agreement between initial P 0 and final P t . P t values as found by Friedenwald are not significantly different in rabbit and human eyes. It seemed, therefore, justifiable to make calculations from the tonographic tracings in rabbits using the tables compiled for human eyes." Furthermore, the current study was not so much concerned with the absolute numerical measurements of these data as with the comparative measurements before and after the cautery in the same eye. It can be assumed, therefore, that the described figures correctly reflect the actual changes in the eyes of rabbits in spite of utilizing data originally designed for human eye studies. The broad conclusions that may be drawn from this study are as follows : A transitory condition, analogous to open-

angle glaucoma, can be obtained in the rabbit eye by cautery of the aqueous channels on the surface of the sciera. This condition is characterized by an increase of the intraocular pressure and Po/C and a decrease of the facility of outflow. This finding is of significance since it demonstrates that the facility of outflow is not exclusively dependent on the state of the trabecular meshwork as has been suggested by a number of investigators ; rather it shows that there are other ocular structures that, at least experimentally, influence the facility of outflow. Moreover, this experiment suggests that in the rabbit eye there is a compensatory mechanism that can be brought into action when the intraocular pressure is elevated by occlusion of some aqueous outflow channels. The compensatory mechanism is presumably related to the intrascleral vascular system which has the capability of dilating, thus allowing for an extra load of the aqueous outflow. SUMMARY

1. A preliminary study was conducted to observe the aqueous vessels and to determine their changes as a result of pressure on the globe and of instillation of vasoconstrictor drops into the conjunctival sac. 2. Tonography was carried out on rabbits' eyes before, immediately following, five and 24 hours after the angiocautery of the aqueous vessels. 3. Analysis of the tonograms showed a statistically significant rise in the Po and Po/C and a decline in the C values in the recordings obtained immediately following the angiocautery. 6001 Lakeside Avenue (23228). ACKNOWLEDGMENT

We wish to acknowledge the help of Mrs. Rhoda Maddox of the Biophysics and Biometry Department of the Medical College of Virginia for the processing and interpretation of the statistical data and to the Visual Education Department for technical assistance in preparing the illustrations.

ANGIOCAUTERY OF THE AQUEOUS CHANNELS

255

REFERENCES

1. Ascher, K. : Zur Mikroskopie des lebenden Auges. Klin. Mbl. Augenh., 72:628 (May-June) 1964. 2. : Aqueous veins. Am. J. Ophth., 25:31 (Jan.) 1942. 3. : Physiologie importance of the visible elimination of intraocular fluid. Am. J. Ophth., 25 : 1174 (Oct.) 1942. 4. : Local pharmacologie effects on aqueous veins. Am. J. Ophth., 25:1301 (Nov.) 1942. 5. Goldmann, H . : Abfluss des Kammerwassers beim Menschen. Ophthalmologica, 111:146 (Feb.Mar.) 1946. 6. : Weitere Mitteilung über den Abfluss des Kammerwassers beim Menschen. Ophthalmologica, 112:344 (Dec.) 1946. 7. : Studien über den Abflussdruck des Kammerwassers beim Menschen. Ophthalmologica, 114: 81 (Aug.) 1947. 8. : Écoulement de l'humeur aqueuse chez l'homme. Ann. Ocul. ( P a r i s ) , 180:114, 1947. 9. Ashton, N. : Neoprene casts of Schlemm's canal, showing the origin of aqueous veins. X V I Internat. Cong. Ophth., London, July, 1950, Clinicopathological Exhibitions, p. 17. 10. : Anatomical study of Schlemm's canal and aqueous veins by means of neoprene casts: Part I. Schlemm's canal. Brit. J. Ophth., 35 :291 (May) 1951. 11. : Anatomical study of Schlemm's canal and aqueous veins by means of neoprene casts: Part II. Aqueous veins. Brit. J. Ophth., 36:265 ( M a y ) 1952. 12. Theobald, G. D. : Schlemm's canal: Its anastomoses and anatomic relations. Trans. Am. Ophth. Soc, 32:574, 1934. 13. : Further studies on the canal of Schlemm: Its anastomoses and anatomic relations. Am. J. Ophth., 39:65 (Apr. Pt. I I ) 1955. 14. Sheppard, L. B. : Anatomy and histology of the normal rabbit eye with special reference to the ciliary zone. Arch. Ophth., 66:896-904 (Dec.) 1961. 15. : The anatomy and histology of the normal rabbit eye with special reference to the ciliary zone. Arch. Ophth., 67:87-100 (Jan.) 1962. 16. Weekers, R., and Prijot, E. : Recherches expérimentales sur les fonctions des veines aqueuses. Ophthalmologica, 119:321, 1950. 17. : Recherches expérimentales sur le vasomotricité des veines aqueuses. Bull. Soc. belge Ophthal., 96:535 (Nov.) 1950. 18. : Vasoconstriction des veines aqueuses et ophthalmotonus. Ophthalmologica, 121:264 (May) 1951. 19. : Étude du mode d'action de la diathermie rétrociliare par l'épreuve de compression au tonometre électronique. Bull. Soc. belge Ophtal., 99:424 (Nov.) 1951. 20. Huggert, A. : Obstruction of the outflow of aqueous humor produced experimentally. Acta ophth. (Kbh.), 35:1 (Jan.) 1957. 21. Vail, D., and Ascher, K. : Corneal vascularization problems. Am. J. Ophth., 26:1025 (Oct.) 1943. 22. Vries, S. de: Aqueous veins at the anterior surface of the eyeball. Ophthalmologica, 115:361, 1948. 23. Becker, B. : Tonography in the diagnosis of single (open-angle) glaucoma. Am. Acad. Ophth., 65 : 156 (Mar.-Apr.) 1961. 24. Kornbleuth, W., and Linnér, E. : Experimental tonography in rabbits. Arch. Ophth., 54:717, 1955.

T H E U S E O F CORNEA TO REPLACE TARSUS* HENDERSON C. ALMEIDA, M.D. Belo Horizonte, Minas, Brasil O p h t h a l m i c s u r g e o n s , in g e n e r a l , a g r e e t h a t it is m u c h m o r e difficult t o r e p a i r t i s s u e loss of t h e u p p e r eyelid t h a n t h a t of t h e l o w e r eyelid. S m a l l c o l o b o m a s c a n b e r e paired b y different techniques1"3 b u t larger

d e f e c t s p r e s e n t a r e a l p r o b l e m . I t is t h e p u r p o s e of t h i s p a p e r t o a n a l y z e t h e u s e of c o r n e a t o r e p l a c e t a r s u s a n d s u b s e q u e n t l y to r e p a i r l a r g e r d e f e c t s of t h e u p p e r eyelid.

* From the Department of Ophthalmology, Northwestern University Medical School, and the Veterans Administration Research Hospital, Chicago, Illinois.

T w e l v e albino rabbits and four dogs w e r e u s e d . T h e e x p e r i m e n t c o n s i s t e d of t w o

METHODS

parts: (1) dogs' corneas were transplanted