Nonpigmented Rabbits Research Articles

Retinal injury thresholds for 532, 578, and 630 nm lasers in connection to photodynamic therapy for choroidal neovascularization

The purpose of this study was to explore the retinal injury thresholds in rabbits and evaluate the influence of retinal pigmentation on threshold irradiance at laser wavelengths of 532, 578, and 630 nm which might be involved in hypocrellin B (HB) and hematoporphyrin monomethyl ether (HMME) photodynamic therapy (PDT) for choroidal neovascularization (CNV). The eyes of pigmented and non-pigmented rabbits were exposed to 532, 578, and 630 nm lasers coupled to a slit lamp biological microscope. The exposure duration was 100 seconds and the retinal spot size was 2 mm throughout the experiment. The minimum visible lesions were detected by funduscopy at 1 and 24 hours post exposure. Bliss probit analysis was performed to determine the ED50 thresholds, fiducial limits and probit slope. In pigmented rabbits, the 24-hour retinal threshold irradiances at 532, 578, and 630 nm were 1,003, 1,475, and 1,720 mW/cm(2) , respectively. In non-pigmented rabbits, the 24-hour threshold irradiances were 1,657, 1,865, and 15,360 mW/cm(2) , respectively. The ED50 for 24-hour observation differed very little from the ED50 for 1-hour observation. The non-pigmented rabbits required a ninefold increase in threshold irradiance at 630 nm comparing to the pigmented rabbits. This study will contribute to the knowledge base for the limits of laser irradiance in application of HB or HMME PDT for CNV.

A comparative study on ocular damage induced by 1319nm laser radiation

High energy loss of 1,319 nm laser due to pre-retinal water absorption makes the ocular axial length more critical, while the relative low absorbance of melanin makes retinal pigmented epithelium less contributing, to retinal damage threshold. However, both have never been illustrated experimentally. Here we determined and compared the retinal damage thresholds at this wavelength in three species with different axial lengths and retinal pigmentations. The corneal damage threshold was also determined for further comparative analysis. The retinal damage thresholds of albino rat, non-pigmented and pigmented rabbit, and the corneal damage threshold of non-pigmented rabbit were determined for 0.2 and/or 0.4-second exposure durations. The incident beam diameter on cornea was 5 mm for rabbit retinal and 2 mm for rat retinal and rabbit corneal lesion. Minimum visible lesions were examined 1- and 24-hour post-exposure. Probit analysis was used to establish the estimated damage threshold for 50% of exposures (ED(50) ). The direct transmittance of pre-retinal eye media was used for further comparative analysis. The retinal ED(50) of albino rat for 0.2 seconds, non-pigmented rabbit for 0.2 seconds, 0.4 seconds, pigmented rabbit for 0.4 seconds was 8.8, 12.1, 22.5, 18.5 J/cm(2) (0.28, 2.4, 4.4, 3.6 J in total intraocular energy (TIE)). The corneal ED(50) was 86.1 J/cm(2) . Under the condition of 5 mm beam diameter and 0.4-second exposure duration, the calculated retinal ED(50) of human was higher, while that of rhesus monkey was lower than the corneal ED(50) . For 1,319 nm laser radiation, the ocular axial length has great, while the retinal pigmentation has only slight influence on retinal damage threshold. The relative lower direct transmittance of human eye media makes the retina more resistant to laser lesion, which should be considered when deriving human maximum permissible exposure (MPE).

Relationship between Intensity of Reflected Light and Temperature Increase: Assessment of Fundus Pigmentation for Transpupillary Thermotherapy

To develop a new device for measuring reflected light during diode-laser irradiation in transpupillary thermotherapy (TTT) and to assess the correlation between fundus pigmentation, the measured intensity of reflected light, and the increase in fundus temperature. We developed a device to measure reflected light by modifying a slit-lamp-mounted 810-nm diode-laser delivery system used clinically for TTT. The diode laser was used to irradiate test charts with various degrees of reflectance in in vitro experiments, and the fundus of nonpigmented or pigmented rabbits in in vivo experiments; then, the intensity of the reflected light and the temperature increase at the target were measured. The retinal sections were also examined histologically. There was a significant negative correlation between the intensity of the reflected light and the temperature increase in the target that depended on the degree of reflectance of the charts or the pigmentation of the rabbit eyes. On histopathologic examination, the extent of the changes in the irradiated retina after TTT was clearly different between pigmented and nonpigmented rabbits. Correlations between fundus pigmentation, the intensity of reflected light during diode-laser irradiation, and the temperature increase in TTT were demonstrated in vitro and in vivo. Our results suggest that measurement of the intensity of reflected light should be helpful for modulating the laser power in TTT according to the degree of fundus pigmentation.

The role of pigmentation on the development of chloroquine retinopathy in rabbits

In a prospective study using sex-matched pigmented and non-pigmented rabbits, the role of pigmentation or melanin on chloroquine-induced retinal or ocular toxicity was determined. Results showed that chloroquine produced a series of ocular toxicity ranging from loss of pupillary and corneal reflex to severe retinopathy. The minimal dose at which these ocular toxicities develop on pigmented rabbits was found to be 5 mg/kg daily for 30 days or a cumulative dose of 225 mg of chloroquine. Furthermore, the cumulative effect of the drug depends on the body weight of the animal, total dose administered and duration of therapy. It is concluded that the effect of chloroquine on pigmentation depends on total amount of drug used, which would predispose to retinal pathology. Keywords : Chloroquine, pigmentation, melanin, retinopathy, retinotoxic West African Journal of Pharmacology and Drug Research Vol. 21 (1&2) 2005: pp. 55-59

The effect of indocyanine green pretreatment on the parameters of transscleral diode laser thermotherapy-induced threshold coagulation of the ciliary body.

To study the effect of indocyanine green (ICG) pretreatment on threshold parameters of transscleral diode laser thermotherapy-induced threshold coagulation of the ciliary body. The procedure was termed 'cyclothermotherapy' based on the long duration (15-60 seconds) of diode laser application. The right eyes of nine young adult New Zealand white rabbits underwent transscleral cyclothermotherapy (TCT, Group 1), TCT following ICG pretreatment (Group 2), and external manipulation of the ciliary body alone (Group 3). Rabbits were sacrificed after 24 hours; specimens were evaluated with gross examination and light microscopy. Thresholds were 30 J/cm2 (TCT) and 4.5 J/cm2 (TCT with ICG). Widespread structural damage was seen in the ciliary processes and the ciliary body in Groups 1 and 2. In Group 3, external manipulation of the ciliary body caused hemorrhage and structural damage confined to the ciliary processes. ICG pretreatment reduced the energy necessary to cause a threshold lesion with TCT in nonpigmented rabbits.

Transpupillary thermotherapy threshold parameters: funduscopic, angiographic, and histologic findings in pigmented and nonpigmented rabbits.

To evaluate the effect of pigmentation on threshold fluence levels, needed to produce visible and angiographic lesions, of transpupillary thermotherapy (TTT) in rabbits. Six pigmented and nine nonpigmented rabbits underwent TTT with an 810-nm diode laser coupled to a slit-lamp biomicroscope using a spot size of 2 or 3 mm. The power ranged from 80 to 200 mW with 1 to 3 minutes of laser exposure for pigmented rabbits and 750 to 1800 mW with 1 minute of exposure for albino rabbits. These parameters were also evaluated after compression of the globe using the contact lens to induce blanching of the optic nerve head. After the experiment, the eyes were enucleated under deep anesthesia, and the animals were killed immediately. In pigmented rabbits, the threshold fluence with the 2-mm spot size was 229 J/cm2 without compression and 153 J/cm2 with compression. With the 3-mm spot size, the threshold decreased from 200 to 150 mW as the duration of exposure lengthened (2 or 3 minutes), increasing the fluence from 170 to 382 J/cm2. In nonpigmented rabbits, the threshold fluence with the 2-mm spot size was 2,865 J/cm2 without compression and 2,674 J/cm2 with compression. With the 3-mm spot size, the threshold fluence of 1,528 J/cm2 was not affected by compression. Histopathologic studies showed transretinal damage at the lowest levels necessary to achieve angiographic evidence of a treatment lesion or a barely visible funduscopic lesion at the time of treatment. Nonpigmented rabbits required more than a 12-fold increase in total TTT fluence compared with pigmented rabbits with the 2-mm spot size and a ninefold increase with the 3-mm spot size. Inner and outer retinal damage was seen histopathologically at these levels.

Transpupillary thermotherapy threshold parameters: effect of indocyanine green pretreatment.

To evaluate the effect of combined treatment with systemic indocyanine green (ICG) on threshold fluence levels of transpupillary thermotherapy (TTT) in rabbits. Four pigmented rabbits and 13 nonpigmented rabbits were studied. TTT was performed on normal rabbit choriocapillaris using an 810-nm diode laser via slit-lamp biomicroscope delivery through a Goldmann macular lens. Laser spot size, power, and duration of laser exposure were varied to achieve a range of TTT fluences for threshold testing in both albino and pigmented rabbit fundi. Intravenous ICG pretreatment at doses of 0.41 to 10 mg/kg was initiated at varying times before TTT treatment. After the experiment, the eyes were enucleated under deep anesthesia, the animals were killed, and the eyes were prepared for light microscopy. When intravenous ICG pretreatment was employed, there was a dose-dependent decrease in the TTT fluence threshold as compared with known threshold values. At threshold fluences, histopathologic sections revealed damage to all layers of the retina in addition to choriocapillaris damage. Intravenous ICG pretreatment can be used to lower the TTT threshold fluence and irradiance required to create angiographically visible lesions in the normal rabbit choriocapillaris. Damage was seen in all layers of the retina and choriocapillaris at threshold levels when TTT was used alone or in combination with ICG pretreatment.

Biodegradable calcium phosphate nanoparticles as a new vehicle for delivery of a potential ocular hypotensive agent.

The purpose of this study was to determine the efficacy of a newly prepared formulation containing biodegradable calcium phosphate nanoparticles (CAP) and 7-hydroxy-2-dipropyl-aminotetralin (7-OH-DPAT) in pigmented and non-pigmented rabbits using the surrogate end points of intraocular pressure (IOP) and aqueous flow rate. IOP (mmHg) was measured by utilizing a manometrically calibrated Mentor pneumatonometer. Rates of aqueous humor flow were measured with a Fluorotron Master by estimating the dilution rate of fluorescein. In non-pigmented rabbits, the ocular hypotension induced by topical administration of 7-OH-DPAT (75 microg) with CAP (115 microg) was more pronounced and sustained than that of 7-OH-DPAT without CAP. Furthermore, IOP-lowering effects of topically administered 7-OH-DPAT (125 microg) alone were markedly diminished in pigmented rabbits compared to non-pigmented rabbits. However, topical application of 7-OH-DPAT formulated with CAP produced significant dose-related (37.5, 75, 125 microg) reductions of IOP accompanied by suppression of aqueous humor flow rates in pigmented rabbits. It is postulated that 7-OH-DPAT in vehicle without CAP binds to pigments in the anterior segment of the pigmented rabbit's eyes, and this binding limits the 7-OH-DPAT's action. Pretreatment with raclopride, a dopamine D2/D3 receptor antagonist, reduced the ocular hypotensive effect induced by 7-OH-DPAT in vehicle containing CAP thereby supporting the role for dopamine D2/D3 receptors in modulating IOP. It is concluded that CAP, as a delivery system, enhances activity by 7-OH-DPAT in pigmented rabbit eyes suggesting that CAP is potentially useful for achieving controlled and targeted drug delivery for treatment of ocular diseases.

Ocular pANCA antigens are expressed in nonpigmented ciliary body epithelium and are conserved in multiple mammalian species

PURPOSE: pANCA marker autoantibody is expressed by a subset of patients with anterior uveitis. A recombinantly isolated pANCA monoclonal antibody, Fab 5–3, identifies ocular expression of corresponding pANCA antigens in human ciliary body and retina. In this study, Fab 5–3 was used to explore pANCA antigen expression in ocular tissues of multiple mammalian species and identify the ciliary body cell type expressing the pANCA antigen. METHODS: Ocular tissues were obtained from several mammalian species and evaluated for expression of the pANCA (Fab 5–3) antigen(s) using immunohistochemistry and Western analysis of tissue extracts. Additionally, primary cultures of nonpigmented and pigmented rabbit ciliary body epithelium were analyzed for pANCA expression using immunofluorescence and Western analysis. RESULTS: Ocular pANCA (Fab 5–3) antigen expression was observed by immunohistochemistry only in the cytoplasm of retinal ganglion cells and ciliary body epithelium. Retinal antigen expression was conserved in all species examined. Ciliary body expression was observed in human, rabbit, rat, and mouse, but not in pig or cow. Antigen expression in the rabbit ciliary body was restricted to the nonpigmented layer as defined in primary cultures of nonpigmented and pigmented ciliary body epithelium. Immunoreactive proteins in both the human and rabbit included a 32–33 kDa doublet (histone H1), and novel 80 and 100 kDa proteins. CONCLUSIONS: This study identifies ocular pANCA antigen expression in multiple mammalian species localized to the retinal ganglion cell layer and the non-pigmented ciliary body epithelium. The present study also establishes novel 80 and 100 kDa proteins which may correspond to the cytoplasmic antigens detected in situ and can be further characterized biochemically and immunologically using small animal model systems.

Vitreoretinal surgery through multifocal intraocular lenses compared with monofocal intraocular lenses in fluid-filled and air-filled rabbit eyes

Purpose To compare vitrectomy procedures and visualization of posterior segment structures through multifocal silicone intraocular lenses (IOLs) with the same procedures through monofocal silicone IOLs in rabbit eyes. Design Experimental study. Participants Twelve eyes of six rabbits. Methods Each rabbit eye underwent phacoemulsification of the lens and posterior chamber implantation of a silicone multifocal or silicone monofocal IOL. The type of IOL (monofocal vs. multifocal) implanted in the first eye of each rabbit was randomly decided. The fellow eye then received the other IOL type. Vitrectomy procedures were performed through the IOLs by using a flat contact lens (part 1) or wide-angled contact lens (part 2) for visualization through fluid-filled (parts 1 and 2) and air-filled (part 2) eyes. Main outcome measures Image quality, stereopsis, and contrast were subjectively graded on a scale of 0 (none) to 4 (excellent) for each eye by each surgeon. Results In part 1, image quality averaged 4 for the monofocal IOL and 3.6 for the multifocal IOL. Stereopsis averaged 4 for the monofocal IOL and 4 for the multifocal IOL. Contrast averaged 4 for the monofocal IOL and 3.9 for the multifocal IOL. Vitrectomy with retinal surface maneuvers was successfully performed in both pigmented and nonpigmented rabbit eyes through both IOL types. In part 2, image quality, stereopsis, and contrast were rated as 4 for both multifocal and monofocal silicone IOLs. Air-fluid exchange was performed without difficulty. Image quality, stereopsis, and contrast were rated as 4 for air-filled eyes. Conclusions Visualization of posterior segment structures through multifocal silicone IOLs was sufficient for retinal surface maneuvers during vitrectomy procedures in both fluid-filled and air-filled rabbit eyes.

Threshold Power Levels for NPe6 Photodynamic Therapy

To determine the threshold power levels for producing retinal and choroidal vascular occlusion using mono-L-aspartyl chlorin e6 (NPe6) photodynamic therapy; to evaluate its efficacy with longer intervals between photosensitizer injection and laser application; to determine the elapsed time between light application and appearance of angiographic changes. Pigmented and nonpigmented rabbits were injected intravenously with 2 mg/kg of NPe6 before laser irradiation of the retina-choroid. Group 1 was treated at increasing power levels; fluorescein angiograms were obtained at each fluence. Group 2 animals were exposed to laser irradiation at 5 minutes, and 1 and 3 hours postinjection to determine (by fluorescein angiography 24 hours post-treatment) if increasing the interval affected outcome. Group 3 animals underwent fluorescein angiography at 30 minutes, 1 hour, 2 hours, and 24 hours posttreatment to document the time between laser application and subsequent vessel closure. Choroidal vessel occlusion was angiographically evident in all lesions at fluences of > or = 2.65 J/cm2 in pigmented rabbits and at > or = 0.88 J/cm2 in nonpigmented rabbits. Lesion diameter decreased as the time between injection and treatment increased. Vessel occlusion was documented at least 2 hours after treatment. Choroidal vessel occlusion can occur at very low fluence.

Problems with and pitfalls of photodynamic therapy

ObjectiveTo delineate the various factors that may influence the outcome of photodynamic therapy of the retina and choroid. DesignExperimental animal study. AnimalsPigmented and nonpigmented rabbits; rhesus monkeys. InterventionThe hydrophilic photosensitizer, mono-l-aspartyl chlorin e6, which is maximally activated at 664 nm, was studied after intravenous injection into pigmented and nonpigmented rabbits and rhesus monkeys. Laser light was supplied by a red diode laser coupled to a modified slit-lamp biomicroscope and delivered to the ocular fundus after passing through a standard fundus contact lens. Standard photodynamic parameters were used. The effects of fundus pigmentation, intraocular pressure, spot focus and defocus, region of fundus treated, equivalent fluence, and retreatment were observed in the different animal species. Main outcome measuresSlit-lamp biomicroscopy, fluorescein angiography, light and transmission electron microscopy. ResultsFundus pigmentation appeared to be a factor only at the lowest fluence level tested, where only 4 of 12 lesions attempted in pigmented fundi were noted on fluorescein angiography, compared with 12 of 12 lesions in albino rabbits. At normal intraocular pressures and a given fluence, 10 of 10 lesions were fully manifested on fluorescein angiography, compared with 4 of 10 at 30 mmHg and 0 of 10 at pressures sufficient to blanch the optic nerve (>60 mmHg). For laser spots either focused or defocused, there were 6 of 6 lesions that were fully manifested on fluorescein angiography for each of the parameters. Lesions treated in the fovea resulted in larger spots on fluorescein angiography. The fluence of 5 mW for 10 seconds resulted in a larger lesion on angiography than the equivalent fluence of 10 mW for 5 seconds. Areas of retreatment in rabbits demonstrated more thinning of the neurosensory retina and loss of photoreceptor outer segments and nuclei than corresponding areas receiving one treatment. ConclusionsPhotodynamic therapy results varied, depending on intraocular pressure, region of fundus treated, ocular pigmentation, and the total time of exposure to the photosensitizer. Retreatment resulted in progressive thinning of the neurosensory retina with loss of photoreceptor outer segments and nuclei in the rabbit eye.

Blood Velocity in an Experimental Iris Tumor

Greene strain melanoma was implanted into the irides of eight nonpigmented rabbits to evaluate the blood flow in tumor vasculature. Conventional scanning laser ophthalmoscopy was used in conjunction with fluorescent microsphere angiography (FMA). Changes were documented on SVHS videotape for later analysis. Individual microsphere movement was tracked through the tumor vessels. Subsequently, blood velocity measurements were taken. The tumor vessels were poorly organized and inefficient. Tumor blood velocity was up to 2.5 times slower compared with normal blood velocity in the unaffected iris of the same eye of the same rabbit (P = .05). Tumor blood flow could be qualitatively visualized in real time in the liver rabbit model. The ability to visualize fluorescent microspheres within the poorly organized tumor vasculature coupled with the reduced blood velocity in the tumor helps to explain the success of hyperthermic tumoricidal therapy, and may allow for development of more efficient and selective drug delivery systems and tumoricidal agents.

Fluorometric Measurement of Light Absorption by the Rabbit Cornea

A bench specular microscope, modified to perform as a scanning microfluorometer, was used to make simple and precise measurements of the absorption of visible light by the isolated rabbit cornea. The tissue was scanned while identical FITC dextran solutions were perfused over both the surfaces, and the measured fluorescence levels in the solutions on the two sides were compared. The alteration in level corresponds to the difference between the absorption of the cornea and that of an equal thickness of the solution, which was determined in a separate experiment. The absorption of light at 500 nm ranged from 0-8% with a mean of 3·3% ( n = 20; S.E. = 0·52). Pigmented and non-pigmented rabbits did not show any clear difference in absorption. These results suggests that light absorption is not a major source of error in anterior segment fluorophotometry.

Different Iris Coloration and Uptake of a Fluoroquinolone Agent into the Iris Ciliary Body of Rabbit Eyes

In order to know the correlation of a drug with high affinity to melanin in the iris ciliary body with the amount of melanin contained in the iris ciliary tissues, an in vivo study was performed on pigmented rabbits with brown or blue irises and albino rabbits. The melanin concentrations in the iris ciliary bodies of pigmented rabbits with brown and blue irises and nonpigmented rabbit eyes with whitish irises were determined. The mean concentrations of melanin in the eyes with brown-, blue- and whitish-colored irises were 47.43, 34.73 and below 0.2 microgram/mg tissue, respectively. The amount of sparfloxacin uptake in the iris ciliary bodies of eyes with brown, blue and whitish irises administered as 40 microliters of 0.3% solution 5 times every 5 min was 15.46 +/- 2.07, 8.53 +/- 1.87 and 1.9 micrograms/g, respectively. The results may be useful for intraocular drug dynamics studies.

Biomicroscopic and Histopathologic Observations After Corneal Laser Photocoagulation in a Rabbit Model of Corneal Neovascularization

Corneal neovascularization complicates many anterior segment diseases. Corneal laser photocoagulation using yellow light (577 nm) has been shown to reduce corneal neovascularization. No histopathologic studies of the effects of this treatment on the eye have been reported, however. Target (cornea) and nontarget (iris, lens, retina, and choroid) ocular tissue were studied 1, 24, and 48 h and 5 days after yellow dye corneal laser photocoagulation in a rabbit model of corneal neovascularization. Biomicroscopic examination of the corneas revealed intracorneal hemorrhage in five of 24 (21%) eyes of nonpigmented rabbits. Faint lenticular opacities were observed in two eyes of pigmented rabbits 24 h after laser treatment. Histopathologic examination revealed increased cellularity (neutrophils) (p < 0.005) in the cornea, increasing from 1 h after treatment, peaking 24 h later, and persisting past 5 days. Distortion of the corneal lamellae by red blood cells occurred in eyes in which intracorneal hemorrhage developed. These results indicate that corneal laser photocoagulation using yellow light is a relatively safe procedure for reducing corneal neovascularization.

Comparative Effects of Alpha-2 and DA-2 Agonists on Intraocular Pressure in Pigmented and Nonpigmented Rabbits

Alpha-2 and DA2 adrenoceptor agonists produce ocular hypotension in cats, rabbits, monkeys and humans. In this study, the effects of topical, unilateral administration of medetomidine (MED), an alpha-2 agonist, and Ha-118 (HA), a DA2 agonist, were compared on intraocular pressure (IOP) in normal, unilaterally sympathectomized (SX), and ocular hypertensive (oral water loaded) Dutch Belted (DB) and New Zealand White (NZW) rabbits. MED (75 micrograms) produced bilateral ocular hypotensive effects in both DB and NZW normal rabbits; however, HA (250 micrograms) lowered IOP unilaterally in NZW rabbits only. MED (25 micrograms) inhibited the rise in IOP caused by oral water loading in both DB and NZW rabbits; HA, on the other hand, was effective only in NZW rabbits. Topical bilateral pretreatment with metoclopramide, a DA2 antagonist, inhibited the ocular antihypertensive effect of HA in NZW rabbits. The IOP lowering effects of MED were absent in the SX eyes of DB and NZW rabbits, but hypotensive responses to MED were present in normal (contralateral) eyes of both strains. In contrast, HA was effective only in the treated, normal eyes of SX NZW rabbits. These data support pharmacodynamic roles for alpha-2 and DA2 receptors in modulating IOP. The lack of activity by HA in DB rabbit eyes suggests a possible absence of DA2 receptors or excessive binding of HA to pigment in the anterior segment of DB rabbit eyes.

Binding of 14C-imipramine by pigmented and non-pigmented tissues

When pigmented and non-pigmented rabbit irides were incubated with various concentrations of 14C-imipramine at equilibrium (120 min), the accumulation of the drug by the pigmented iris was 1.5 times as great as that by the non-pigmented iris. The accumulated drug is lost from both types of irides in a complex fashion. However, even after 120 min of washing, the differences in accumulation remain nearly constant. When accumulation of the drug in the non-pigmented iris was analyzed by discontinuous sucrose density gradient, it was observed that the drug was bound mainly by the low density sucrose fractions where the synaptosomes separate. On the contrary, in the pigmented iris approximately 70% of the drug was found in the melanin-containing fraction. The homogenate from the substantia nigra accumulated 1.5 times more than that from the human brain cortex. The affinity of the drug for bovine iris melanin granules and the synthetic L-dopa melanin was 9.9 × 10 5 M −1 and 3.8 × 10 3 M −1, respectively. On the rabbit iris sphincter muscles, imipramine was evaluated for antimuscarinic effects. The apparent dissociation constants, K B values, for the antagonist in the non-pigmented and pigmented iris were 1.7 × 10 −7 M and 3.8 × 10 −6 M, respectively. The low antimuscarinic activity in the pigmented iris is attributed to the loss of teh drug to the pigment. On this basis, relevancy of the drug binding by pigmented tissues to the effects of this tricycli drugs is discussed.