The Ciliary Vasculature and Its Perturbation with Drugs and Surgery*
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THE CILIARY VASCULATURE AND ITS PERTURBATION WITH DRUGS AND SURGERY* BY E. Michael Van Buskirk, MD INTRODUCTION THE CILIARY BODY AND ITS VASCULATURE SERVE A WIDE VARIETY OF COM- plex functions in the eye. Three structural and functional components of the ciliary body can be identified: (1) the pars plana; (2) the pars plicata; and (3) the ciliary muscle. Approximately 70 major ciliary processes, interdigi- tated between an equal number of smaller minor processes, comprise the highly vascular pars plicata that is covered by a complex bilayered epithe- lium. Through an array ofintercellularjunctions, the ciliary processes form the inflow side of the blood-aqueous barrier and are responsible for the secretion of aqueous humor. 1-7 As such, they keep the globe inflated with aqueous humor, and provide nutrition and humoral mediators to the avas- cular structures of the anterior segment. The ciliary muscle controls the accommodative mechanism, provides structural support for the peripheral uvea by its attachment to the scleral spur, and modulates outflow ofaqueous humor by means of traction on the trabecular meshwork. 8-11 Physiologic studies of this small, complex tissue demonstrate a similar diversity ofits vascular perfusion with evidence ofvascular autoregulation and for regional changes in blood flow in response to pharmacologic or neurogenic stimuli. 12-20 Advances in photofluorometric measurement of aqueous humor formation in the undisturbed and experimentally altered eye are now rapidly advancing knowledge about factors affecting the formation of aqueous humor. 21-25 However, the interplay of ciliary body perfusion and aqueous formation cannot be understood entirely without a thorough understanding of the microvasculature and microcirculation of the ciliary processes and the surrounding uveal tissues. Recent emphasis on clinical alteration of the ciliary body with aqueous suppressant drugs, *From the Department ofOphthalmology, the Oregon Health Sciences University, Portland, Oregon. Supported in part by grant EY05231 (National Eye Institute), Core grants RRC0163 and RR05694 (National Institutes of Health), and a Heed Foundation Fellowship. TR. AM. OPHTH. Soc. vol. LXXXVI, 1988 Ciliary Vasculature 795 with cyclodestructive surgery, and for support ofintraocular lens implants by this complex tissue, has made study ofthe ciliary vasculature especially relevant. This thesis examines methods for examination ofthe ciliary vasculature, while minimizing artifactual disturbance ofthe tissue. The precise anatom- ic details of the ciliary vasculature are presented in primates and are compared to other nonprimate mammalian eyes. The effects on the ciliary vasculature of commonly used ocular therapeutic modalities are also examined, including pharmacologic manipulation, cyclodestruction, and intraocular lens implantation. METHODS All of the studies for this project involved preparation of methacrylate luminal castings ofthe ocular microvasculature by intra-arterial injection of casting media. This technique, de6cribed by Batson26 in 1955, was evalu- ated intensively and modified,to minimize perturbation ofthe general and ocular physiology prior to injection ofthe plastic casting media. The initial studies of the anterior uveal angioarchitecture employed injection of commercially available Batson's compound no. 17.27 Although it is ex- tremely viscous, carefully injected Batson's compound no. 17 produces complete castings ofmicrovascular beds to the level ofthe finest capillaries, with detailed relief replication of the vascular endothelial nuclear imprint on the surface ofthe casting (Figs 1 and 2).28 Examination ofthese imprints permits differentiation of arteries with their spindle-shaped palisaded nuclei from veins with rounded randomly distributed nuclei (Fig 2). Batson's methacrylate media provides dry rigid castings that are easily handled and dissected to reveal details ofthree-dimensional architecture at a microscopic level. Moreover, plastic compounds resist high voltage and allow scrutiny by scanning electron microscopy (SEM). INITIAL STUDIES In the initial studies, whole ocular microvascular luminal methacrylate castings were prepared by simultaneous bilateral carotid artery injection of Batson's compound no. 17 in anesthetized cynomolgus monkeys.27 The injection medium was prepared according to the manufacturer's recom- mendations. The animals were intratracheally intubated and anesthetized with halothane. The major blood vessels in the neck were isolated through a mid-line incision, the carotid arteries were cannulated with appropriately sized catheters and the jugular veins were incised. Batson's compound was hand-injected simultaneously into the carotid arteries as rapidly as possi- 796 Van Buskirk _ r ... s~~~~~~~~~~..A H! ^uS- FIGURE 1 Gold-palladium coated methacrylate casting ofcanine ocular vasculature. ble, using multiple 10 cc syringes until clear Batson's compound egressed from the cut end ofthe incisedjugular veins. To facilitate filling ofthe ocular vessels, the intraocular pressure was reduced to ambient pressure by incising the cornea with a razor blade fragment immediately prior to injection. After polymerization of the injection material, the eyes were enucleated and fixed in formalin. The whole, methacrylate injected, fixed eyes were immersed and macer- ated in 6 M potassium hydroxide for at least 24 hours at 500 to 55°C, digesting away all tissue. They were then gently tumbled in running tap water to remove all residual tissue and air dried, leaving a rigid methacry- late casting of the entire ocular vasculature (Fig 1). The dried, tissue-free castings were hemisected at the equator and the anterior portions were mounted on a stub for further dissection and for SEM. Dissection ofOcular Castings The ocular castings presented a complexity of intertwined vessels that required careful and orderly methods of dissection. The ciliary processes lie internal to the rich ciliary muscle vasculature rendering them relatively inaccessible to examination without complex microdissection methods.29 Ciliary Vasculature 797 FIGURE 2 Scanning electron micrograph of contiguous artery and vein, showing palisaded spindle- shaped arterial nuclear imprints and rounded venous nuclei (x 600). The hemisected casting ofthe anterior ocular segment was draped over a wooden disc, 2 mm thick and 8 mm in diameter, to preserve its spherical shape while mounted on the stub. The equatorial cut edge was secured to a SEM stub with silver paint. The entire specimen was then coated with gold palladium in a sputter coater and examined with an AMR 1000 scanning electron microscope. SEM of these preparations permitted identification of the anterior ciliary arteries, with their interconnections with the episcleral and per- forating arteries entering the bulk ofthe ciliary muscle. By using a hooked insect dissection needle and fine scissors, the ciliary muscle capillary bed and draining veins were removed layer-by-layer without disturbing the underlying choroidal veins and the ciliary processes vessels using binocular magnification of a dissecting microscope. Subsequent SEM revealed the interconnections of the perforating anterior and long posterior ciliary arteries and their supply to the iris and ciliary processes. To examine the angioarchitecture of the individual ciliary processes, three to four contiguous processes were separated from the main casting with fine scissors and were mounted together on a separate stub. These 798 Van Buskirk specimens were then returned to the scanning electron microscope where the processes could be scanned from various angles by manipulating the microscope stage. Because the arteriolar, capillary, and venular intercon- nections were at the upper limit of resolution for dissecting microscope, scanning electron micrograph stereoscopic pairs were employed to deter- mine and record otherwise undetectable anatomic relationships. An initial photograph of the casting was obtained at low magnification (50 to 200 x) and repeated after tilting the stage by 6 degrees to provide a stereomicro- graph. Examination of these stereoscopic pairs of photographs revealed superficial arteriolar connections to the ciliary processes, and also indi- cated additional connections deeper within the specimen. After recording the superficial layer, individual capillaries and arterioles were cut away using a pneumatically powered ultramicroscissors that was specifically developed for this purpose. It was mounted on the arm ofa micromanipula- tor and brought into the microscopic field without disruption ofthe casting. Thus, the entire casting was examined, recorded, and microdissected layer-by-layer to reveal the complex interconnections within and between the ciliary processes. IMPROVED CASTING METHODS Ocular microvascular castings, conventionally prepared with Batson's com- pound no. 17, produced fine luminal replication ofthe ocular microvascula- ture suitable for detailed determination of the ocular angioarchitecture. Once these baseline structural details were understood, methods for vascular modulation in response to physiologic, pharmacologic, or surgical influences were planned. However, because of the high viscosity of the material, high injection pressures, hand-applied to catheters in each carotid artery, were required for prolonged periods of time in order to obtain complete filling ofdistal ocular capillary beds. To study the effects of the unilateral ocular effects of drugs or operations, it was necessary to assure equal