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148 4. 5. 6. 7. 8. 9. 10. 11. Investigative Reports glucose deprivation on function of isolated mammalian retina, J. Neurophysiol. 26: 617, 1963. Smith, C. C , and Baird, C. D.: Survival time of retinal cells when deprived of their blood supply by increased intraocular pressure, Am. J. Ophthalmol. 35: 133, 1952. Reinecke, R. D., Kuvvabara, T., Cogan, D. C , et al.: Retinal vascular patterns. Part V. Experimental ischemia of the cat eye, Arch. Ophthalmol. 07: 470, 1962. Popp, C : Die retinafunktion nach intraocularer ischemie. v. Craefes Arch. Ophthalmol. 156: 395, 1955. Lamb, A.: Ocular changes occurring during cardiac surgery under profound hypothemiia and occlusion, Br. |. Ophthalmol. 45: 490, 1961. Cole, S., and Corday, E.: 4-Minute limit for cardiac resuscitation, |.A.M.A. 161: 1454, 1956. Ames, A.: Discussion of paper by Briefly, J. B., et al. The threshhold and neuropalhology ol cerebral "anoxic-ischemic" cell change. Arch. Neural. 29: 373, 1973. Drance, S. M. Some factors involved in the production of low-tension glaucoma, in: The optic nerve, Cant, [. S., editor. London, 1972, Henry Kimpton Publisher, pp. 339366. Drance, S. M.: Personal communication, April, 1974. Physical properties of experimental vitreous membranes. I. Tensile strength. TETSUTO NUMATA, IAN J. CONSTABLE, AND DANIEL E. WHITNEY. The tensile strength of vitreous membranes, induced by silk sutures imbedded in the vitreous body and removed one to two weeks later, was investigated in rabbits. Three types of membranes were distinguished by their appearance under an operating microscope: dense, opaque, cylindrical membranes, (Type 1), broken with 2 to 16 Cm. of force and elongated from 127 to 200 per cent before breaking; thin, cylindrical membranes, (Type 2), broken with 200 mg. to 2 Cm. of force and elongated from 51 to 152 per cent before breaking; and thin film-like membranes imbedded in formed vitreous gel, (Type 3), broken with 200 mg. to 5.5 Cm. of force and elongated from 53 to 1S6 per cent before breaking. Vitreous membranes formed two to eight weeks after surgery resisted forces greater than those required to detach rabbit retina. The results of the experiment are rclcvatit in the design of vitrcctomy instruments. A satisfactory surgical instrument for vitreous membrane removal by the closed transcleral route Ophthalmology February 1975 must possess properties such as small size (maximum 2.0 mm. diameter tube), a separate infusion chamber to maintain pressure, and a suction line to remove cut membranes without pulling on the easily detachable retina. These requirements have been quite well recognized by others.'••s However, knowledge of the mechanical properties of the materials being cut would be very useful for the systematic design of specific details such as shape, material, and finish of the cutting edge, and speed and stroke of the cutting mechanism. The present experiment was carried out to investigate the tensile strength of vitreous membranes produced in rabbit eyes. Materials and. methods. Production of vitreous membranes. Ten rabbits were used for this experiment weighing between 1.7 to 4.2 kilograms. Animals were anesthetized with intravenous pentobarbital sodium (25 mg. per kilogram) and pupils were dilated prior to surgery with phenylephrine 10 per cent and scopolamine 0.3 per cent. The eyes were proptosed and held by forceps applied to the superior and inferior recti. Three to six sutures (4-0 silk) were passed through conjunctiva and sclera across the vitreous cavity at the level of the equator by means of a straight sewing needle. After one or two weeks, sutures were removed and, thereafter, mcmbiane formation was observed by indirect ophthalmoscopy and by slit lamp biomicroscopy. Membrane preparation for testing. Two rabbits were killed every week by an overdose of anesthetic and the eyes were enucleated and put in normal saline at 25" C. Each eye was sutured into a molded plastic hemisphere, then opened by an incision around the limbus. The anterior segment was lifted with forceps and the lens cut away from the vitreous face. The membranes were then dissected out from the eye under an operating microscope and transferred onto a flat, black plexiglass plate. Then, membranes were cut into test specimen sizes. The individual form clamps were made by cutting soft polyurethane form into 1.6 by 1.6 by 0.4 mm. blocks and glueing them onto strips of aluminum foil (Fig. 1). Membrane test specimens were transferred from the black plexiglass plate onto form clamps submerged in normal saline at 25U C , and the ends of each membrane were then pressed between two such forms (Fig. 1). Form clamps prevented membranes from slipping during pulling and from being pinched directly by the plexiglass clamps. Pinching might create a local weakness in membranes where they would break with very small forces. Each membrane specimen secured at both ends by form clamps was carefully transferred into a plexiglass testing bath filled with normal saline at 25° C , and secured tightly into two plexiglass clamps as shown in Fig. 1. The specimen was then pulled until broken by a trans- Downloaded From: http://iovs.arvojournals.org/pdfaccess.ashx?url=/data/journals/iovs/933292/ on 06/11/2017 Volume 14 Number 2 Reports 149 Polyurethone foam Upper plexiglass clamp Membrane Aluminum-form clamps Lower plexiglass clamps Fig. 1. Apparatus for testing tensile strength, The transducer (above) moves up at constant speed, stretching the vitreous membranes which are held between two plexiglass clamps (lower left). The form clamps (lower right) prevent uneven force from being applied to the vitreous membrane. ducer mounted on a microscope stand, which was driven up and down by a motor at V2 rev. per hour (0.3 mm. per minute extension) and Vi rev. per minute (9 mm. per minute extension). The transducer was calibrated using milligram weights and its outputs were recorded on a Sanborn 350 recording machine. Results. Ohaervatumx mudv on the membranes formed. The number of membranes obtained from each eye varied between nine and twenty-two. Membranes formed were classifiable into three categories on the basis of appearance under the operating microscope (Fig. 2). Dense, opaque, white membranes approximately 1 mm. in diameter or less were found bridging two nearby suture perforation sites (Type 1). Incomplete and thin films of vitreous gel surrounded these membranes. This type of membrane tended to be thicker and wider near the perforation sites tapering down in width toward its middle. Fine, cylindrical membranes, approximately 0.05 mm. in diameter, wider near the perforation sites, tapering down (Type 2). Diameters, however, varied and were never constant throughout the length of the membranes. Therefore, small sections of each membrane specimen were selected for testing so that they were as regular as possible. Considerable amounts of formed gel surrounded this type Fig. 2. Schematic diagram of three vitreous membrane types observed under an operating microscope. Type 1 bridged adjacent perforation sites, Type 2 extended along the suture tracks, and Type 3 was embedded in formed vitreous gel. of membrane, but most of it was broken down by the preparation for testing. These membranes were often transparent and hard to observe under the operating microscope. Thin, film-like mem- Downloaded From: http://iovs.arvojournals.org/pdfaccess.ashx?url=/data/journals/iovs/933292/ on 06/11/2017 150 Reports Investigative Ophthalmology February 1975 Fig, 3. Serial photographs of a Type 1 membrane being pulled and broken. branes imbedded in formed vitreous gel were found at random locations in the vitreous cavity (Type 3). This material was difficult to handle and trim into the small size necessary for testing. These membranes were often quite transparent, and could only be seen by means of oblique illumination. Variable amounts of vitreous gel surrounded each type of membrane and could not be easily separated from them, but was continuous only in Type 2 and Type 3 membranes. Although, Type 1 and Type 2 membranes were found radiating out from suture tracks, Type 3 membranes bore no direct relationship to them, being present at random sites within formed vitreous gel. All membranes tended to increase in density up to approximately four weeks, then became thinner and more transparent. None of the membranes had caused actual retinal detachment by pulling on the retina at local attachment points within the period of observation. Tensile strength and membrane extension. Membranes first elongated, then finally broke at a random site (Fig. 3). The tensile strength and the extent of elongation of membranes before breaking are depicted in Fig. 4 as a function of age. Two different extension speeds, 0.3 mm. per minute and 9.0 mm. per minute, did not reveal any strength differences. Type 1 membranes showed a breaking strength of 2 to 16 Cm. and a breaking elongation of 127 per cent to 200 per cent. Type 2 membranes showed a breaking strength of 200 mg. to 2 Cm. and breaking elongation of 51 per cent to 152 per cent. Type 3 membranes showed a breaking strength of 200 mg. to 5.5 Cm. and a breaking elongation of 53 per cent to 186 per cent. All membranes tested were stronger than the force required to detach the retina.!l< in No strength differences were observed between two-week-old and eightweek-old membranes. A clear association between membrane age, strength, and elongation could not be made. Discussion, The method of producing experimental vitreous membranes by means of sutures was found to be reliable. This method has several Downloaded From: http://iovs.arvojournals.org/pdfaccess.ashx?url=/data/journals/iovs/933292/ on 06/11/2017 Reports 151 Volume N Number 2 10,000 1000 100 I 10 200 150 t I Uj 50 SI S2 S3 S4 S5 S6 Fig. 4. Tension data (above). Si to S« represent six rabbits tested. Forces measured are plotted on a long scale in milligrams. S, and S.> were twoweek-old membranes pulled at 0.3 mm. per minute speed. S:, and Si were four-week-old membranes pulled at 9.0 mm. per minute speed. Sr, represents two-week-old membranes pulled at 9.0 mm. per minute speed. S,: represents eightweek-old membranes pulled at 9.0 mm. per minute speed. Elongation data (below). Elongation data of each membrane specimen are plotted in the same order as the tension data. advantages over the commonly used method of injecting blood. The vitreous body remains clear, allowing detailed clinical examination during formation of membranes and during experimental surgery. A reproducible variety of membranes are forme:I and, most importantly, formed vitreous gel is not radically destroyed as occurs after blood injection in the rabbit. This last point is particularly relevant when assessing the efficiency of vitrcctomv instruments. Type I membranes were usually much stronger than either Type 2 or Type 3: Type I membranes represent ingrowth of scar tissue at perforation sites. Their greater strength was probably due principally to larger amounts of these materials rather than to actual structural differences. The fine strands bridging suture tracks (Type 2) were of similar strength to lacelike membranes embedded in formed gel (Type 3). It was difficult to be certain to what degree their tensile strength was dependent on the newly formed membranous tissue and to what degree it was dependent on the formed gel. Attempts to mea- sure the tensile strength of normal formed vitreous gel were unsatisfactory for several reasons. Normal vitreous gel could not easily be cut into small regular sizes suitable for testing, repeated handling tended to break up the gel, and the clear gel was invisible when immersed in saline. However, handling indicated that the formed gel alone was not strong enough to contribute much of the strength of Type 2 and Type 3 membranes. It was inevitable that a wide range of tensile strength and extension would be obtained since individual membranes varied in diameter along the segments being tested, and since only gross trimming of surrounding formed vitreous gel and thicker Type I membranes was done. No attempt was made to measure cross-sectional areas and express results as force per unit area, because it was felt that the range of breaking strength was more relevant for instrument designs than these idealized values. Implications of the results for design of vitrcctomy instruments. The present experiments showed that even the finest vitreous membranes visible only with the aid of the operating microscope were stronger than the forces necessary to detach the rabbit retina in vitro and in vivo and membianes can invariably be broken if stretched a maximum of 200 per cent. These facts suggest cutting mechanism designs in which membranes are first clamped before cutting. With such a design, the dependence on perfectly fitting shearing surfaces would become less critical. Systematic determination of the clamping surface and clamping force necessary would require data on adhesive force and frictional force characteristics of vitreous membranes. Experiments to define these properties for rabbit vitreous membranes will be reported later. The authors thank Charles L. Schepens, M.D., for his support of this project, and Mr. Oleg I'omerantzell and Dr. F. Delori for their advice and assistance. From the Department of Retina Research, Eye Research Institute of Retina Foundation, Boston and the Department of Mechanical Engineering, Massachusetts Institute of Technology, Cambridge, Mass. Supported by the |ohn A. Hartford Foundation, Massachusetts Lion's Eye Research Inc., and by Grant EY 01030-01 from the National Eye Institute of Health. Submitted for publication Aug. 27, 1974. Reprint requests: Editorial Services Unit, Eye Research Institute of Retina Foundation, 20 Staniford St., Boston, Mass. 02114. Key words: tensile strength, vitreous membranes, vitreous gel, vitrectomy. REFERENCES 1. Couvillion, C. C, Freeman, H. M., and Schepens, C. L.: V. Modification of the Downloaded From: http://iovs.arvojournals.org/pdfaccess.ashx?url=/data/journals/iovs/933292/ on 06/11/2017 152 2. 3. 'I. 5. o'. 7. S. 9. 10. Reports vitreous scissors, Arch. Ophthamol. 83: 722, 1970. Machemer, R., Parel, J. M., and Buertner, II.: A new concept (or vitreous surgery. I. Instrumentation, Am. |. Ophthalmol. 73: 1, 1972. Peyman, C. A., Daily, M. |., and Ericson, E. S.: Experimental vitrectomy: new technical aspects, Am. |. Ophthalmol. 75: 773, 1973. Brighbill, F. S , Kaufman, H. E., and Levenson, J. E.: A vitreous suction cutter for aphakic keratoplastv, Am. |. Ophthalmol. 7(>: 331, 1973. Kreiger, A. E., Straatsma, B. R., Griffin, J. R., ct al.: A vitrectomy instrument in stereotaxie intraocular surgery, Am. ). Ophthalmol. 76: 527, 1973. O'Malley, C, and Heintz, R. M.: Vitrectomy via the pars plana—a new instrument system, Trans. Pacific Coast. Oto-Ophthalmol. Soc. 53: 121, 1972. Duvas, N. C : The cataract roto-extractor, Trans. Am. Acad. Ophthalmol. Otolaryngol. 7: 792, 1973. Tolentino, F. I., Bunko, A., Sehepens, C. L., et al.: Vitreous surgery. XII. A new surgical system for vitrectomy. Part I. Instrumentation, Arch. Ophthalmol. (in press) 1974. deGuillebon, H., and Zauberman, H.: Experimental retinal detachment. Biophysical aspect of retinal peeling and stretching, Arch. Ophthalmol. 87: 545, 1972. Zauberman, H., and deCuillebon, II.: Retinal traction in vivo and postmortem, Arch. Ophthalmol. 87: 549, 1972. Qunntitation of pilocarpine flux enhancement across isolated rabbit cornea by hydrogel polymer lenses. DAVID L. KHOHN AND JULIANNA M. BHEITFELLEM. The comparative effect on pilocarpine flux across rabbit cornea induced by two hydrogel polymer lenses containing ci/ual doses was quantitated in a transport chamber. This closed system featured continuous flow of a tear analog but excluded variables of the internal eye influencing concentration. Flux induced by both lenses increased linearly with time. At 240 minutes total flux was a whole order greater than that induced bij the same pilocarpine dose in free fluid. Analysis of pilocarpine in tear analog effluent showed the flux to be independent of the available dose retained in the hydrogel polymer lens, suggesting that eorneul transport of pilocarpine to the aqueous may involve mediation by a carrier system. Clinical studies1"' and animal experiments1' r> have demonstrated enhanced transcorneal pilocarpine flux induced by presoaked hydrogel polymer lenses. The mechanism of this enhancement has February 1975 been uncertain. In addition, exact measurements of the effect of these polymers relative to that of ordinary drop administration or constant flow delivery have been unavailable, since aqueous concentration in the living eye after topical administration of any type is variably influenced by tear elution and multiple intraocular events. The latter include chamber inflow and outflow, preferential uptake of the drug by tissue in the internal eye," isomerization and hydrolysis, "• s anil, in the case of radioisotope determinations, interchange of isotope with solvent." This report describes <|iiantitation of the effect on pilocarpine flux across isolated rabbit cornea of two well-known hydrogel polymer lenses. A transport chamber system was used which incorporates the ellect of tear How but excludes the complex variables of pilocarpine concentration in the living internal eye. It is a closed system in that all administered pilocarpine can be accounted for. The intention was to compare relative flux efficiency of the two polymer lenses at various points in time. It was anticipated that some inference as to mechanism of transport enhancement relative to equal dose "drop" administration might be available from this comparison. The transport chamber and llux data lor single drop administration ol pilocarpine were described in a previous publication.1" All parameters reported, including flush and control runs in advance of the pilocarpine administration, were held constant for experiments now described. However, single fluid pilocarpine instillations were replaced by delivery by presoaked hydrogel polymer lens buttons. Each of these was trephined when fully hydrated with pilocarpine solution. The experiment was designed so that the total available pilocarpine close in each lens button was equivalent to the "secondary" dose in the upper chamber in the previously described single-fluid dose experiments."' Lens buttons were presoaked in 30 to 50 ml. 16.0 per cent pilocarpine—HCI in saline for 48 hours with a change of solution at 24 hours. Each button was blotted in a standard manner, then eluted for a minimum of 1 week in 100 ml. of saline at 4° C. The buttons were then matched on the basis of pilocarpine capacity independent of weight or thickness. Matching buttons were accumulated until a "pool" ol prematched buttons was available. Each lens button from this "pool" was used randomly for determination at various time intervals. Long and short intervals were rim sequentially. From time to time, a group of lenses were taken Ironi the "pool" and re-tested by full soaking and elution for constancy of piloearpine-HCI space. Buttons were discarded if any evidence of fraying or mechanical damage was present. Downloaded From: http://iovs.arvojournals.org/pdfaccess.ashx?url=/data/journals/iovs/933292/ on 06/11/2017