Alteration in vitreal structure following C3F8 injection

Abstract

Although the use of intraocular perfluorocarbon gases has been demonstrated to be a clinically useful procedure for retinal compression and diagnostic procedures, the effect of the gas expansion to 90% ocular volume causes the displacement of the vitreous. This investigation examines the morphological changes in the vitreous of 36 rabbit eyes following C3F8 intravitreal gas injection, by light (LM), transmission (TEM) and scanning (SEM) electron microscopy. Lincoff et al have demonstrated that C3F8gas exhibits a fourfold expansion of its volume at 3-4 days after injection into the vitreous. Following expansion the gases disappear logarithmically with the volume of the bubble following first order decay kinetics. For the rabbit, the C3F8 half-life is six days, with the total reabsorption of the gas in 40 days. To determine whether initial structural changes following gas injection eventually resolved, eyes were studied at 3, 6, 8, 16, 41 and 64 days post injection.Four pigmented and 14 albino mature (3-4 Kg) rabbits were anesthetized and given topical proparacaine anesthetic prior to injection into the inferior vitreous cavity with 0.1, 0.3 or 0.4 cc filtered C3F8. An indirect ophthalmoscope was used to direct the 30 gauge needle to a position 2-3 mm posterior to the limbus. In each case, the contralateral eye was used as the normal control. Animals were euthanized with intravenous T61 solution and the eyes immediately enucleated and frozen in liquid nitrogen. While still submerged in liquid nitrogen the tissue was placed in a deep freeze and allowed to equilibrate to −80°C for 24 hrs. Eyes were cryosectioned at −20°C and the sections placed on prechilled glass slides, cover slips, sapphire wafers or pyrolytic graphite planchets. For LM histochemistry, the frozen sections were placed in 95% ethanol 10 min and stained using a Papanicolaou method. To identify collagen, Van Gieson's picric acid fuchsin and 2.5% aniline blue were used. Proteoglycans were stained with pH 1 or pH 1.5 alcian blue or 0.5% ruthenium red. Some cryosections were freeze dried or fixed in 3% glutaraldehyde, dehydrated and dried by the critical point method (CO2) for SEM examination of an AMRay 1000A equipped with a lanthanum hexaboride gun at Brookhaven National Laboratory, Upton, NY. Tissue for routine TEM was fixed immediately following enucleation in 3% glutaraldehyde in 0.1 M cacodylate buffer (pH 7.4), post fixed in 1% aqueous OsO4, dehydrated in acetone and embedded in Epon 812. To identify and retain proteoglycans, some eyes were fixed in 2% glutaraldehyde in 0.1 M cacodylate containing 0.01% ruthenium red. Both unstained and stained (Ur-acetate and Pb-citrate) plastic thin sections were examined at 80 kV on a Philips 300.