The affinity, intrinsic activity and selectivity of a structurally novel EP2 receptor agonist at human prostanoid receptors.

BACKGROUND AND PURPOSE: Prostanoid EP2 receptor agonists exhibit several activities including ocular hypotension, tocolysis and anti-inflammatory activity. This report describes the affinity and selectivity of a structurally novel, non-prostanoid EP2 receptor agonist, PGN-9856, and its therapeutic potential. EXPERIMENTAL APPROACH: The pharmacology of a series of non-prostanoid EP2 receptor agonists was determined according to functional and radioligand binding studies, mostly using human recombinant prostanoid receptor transfectants. The selectivity of PGN-9856, as the preferred compound, was subsequently determined by using a diverse variety of non-prostanoid target proteins. The therapeutic potential of PGN-9856 was addressed by determining its activity in relevant primate cell, tissue and disease models. KEY RESULTS: PGN-9856 was a selective and high affinity (pKi ≥ 8.3) ligand at human recombinant EP2 receptors. In addition to high affinity binding, it was a potent and full EP2 receptor agonist with a high level of selectivity at EP1 , EP3 , EP4 , DP, FP, IP and TP receptors. In cells overexpressing human recombinant EP2 receptors, PGN-9856 displayed a potency (pEC50 ≥ 8.5) and a maximal response (increase in cAMP) comparable to that of the endogenous agonist PGE2 . PGN-9856 exhibited no appreciable affinity (up 10 µM) for a range of 53 other receptors, ion channels and enzymes. Finally, PGN-9856 exhibited tocolytic, anti-inflammatory and long-acting ocular hypotensive properties consistent with its potent EP2 receptor agonist properties. CONCLUSIONS AND IMPLICATIONS: PGN-9856 is a potent, selective and efficacious prostanoid EP2 receptor agonist with diverse potential therapeutic applications: tocolytic, anti-inflammatory and notably anti-glaucoma.

Pharmacotherapies for glaucoma.

Glaucoma is a group of progressive optic neuropathies in which the axons in the optic nerve are injured, retinal ganglion cell numbers are reduced and vision is gradually and permanently lost. The only approved and effective way to treat glaucoma is to reduce the intraocular pressure (IOP). This is usually accomplished by surgical and/or pharmacological means. Drugs designed to reduce IOP target one or more of the parameters that maintain it. These parameters (collectively known as aqueous humor dynamics) are the production rate of aqueous humor, the pressure in the episcleral veins and the drainage of aqueous humor through the trabecular or uveoscleral outflow pathways. Intraocular pressure lowering drugs can be classified as inflow or outflow depending on whether they reduce aqueous humor inflow into the anterior chamber or improve aqueous humor outflow from the anterior chamber. Inflow drugs, like ß adrenergic antagonists and carbonic anhydrase inhibitors, reduce the rate of aqueous humor production. Outflow drugs, like prostaglandin analogs, cholinergic agonists and sympathomimetics, increase the rate of drainage through the uveoscleral outflow pathway and/or increase the facility of outflow through the trabecular meshwork. Some drugs have mixed inflow/outflow effects. This review summarizes the pharmacological treatments for glaucoma in use today and some new drugs showing potential for use in the future.

Acute effects of insulin on aqueous humor flow in patients with type 1 diabetes.

PURPOSE: Previous studies reported reduced aqueous humor flow through the anterior segment of the eye in patients with type 1 diabetes. This study investigates whether reduced flow is the result of the diabetic state or of alterations in glucose or insulin concentrations. METHODS: A cross-sectional study, involving patients with type 1 diabetes and healthy controls, measured aqueous flow at different insulin concentrations. Eleven patients with type 1 diabetes (hemoglobin A1C = 7.0 +/- 0.3% [mean +/- SEM], normal < 6.5) with no microvascular complications and 17 controls were prospectively studied. Controls were studied fasting and during a hyperinsulinemic-euglycemic clamp (insulin 2 mU/kg per minute). Patients with type 1 diabetes were similarly studied during two euglycemic clamp procedures (insulin 0.5 and 2.0 mU/kg per minute). Aqueous flow was measured by fluorophotometry. Pulsatile ocular blood flow and intraocular pressure were measured with a Langham flow probe. RESULTS: Control subjects had no change in aqueous flow during fasting and hyperinsulinemic conditions (3.0 +/- 0.1 vs 2.8 +/- 0.1 microl per minute). In the patients with type 1 diabetes, aqueous flow was not decreased with hyperinsulinemia, compared with the low insulin state (P =.7). Compared with control subjects, patients with type 1 diabetes had lower aqueous flow during hyperinsulinemia (2.4 +/- 0.1 microl per minute, P =.03) and at lower insulin conditions (2.6 +/- 0.1 microl per minute, P <.05). No differences in intraocular pressure or pulsatile ocular blood flow were noted between groups or between insulin states within groups. CONCLUSIONS: Aqueous flow is decreased in patients with type 1 diabetes under euglycemic conditions of high and relatively low insulin concentrations, despite the absence of microvascular complications.

Potential mechanism for the additivity of pilocarpine and latanoprost.

PURPOSE: To determine the ocular hypotensive mechanism underlying the additivity of latanoprost and pilocarpine. METHODS: This randomized, double-masked study included 30 patients with ocular hypertension on no ocular medications for at least 3 weeks. On each of six visits to the clinic, measurements were taken of aqueous flow and outflow facility by fluorophotometry, intraocular pressure by tonometry, and episcleral venous pressure by venomanometry. Uveoscleral outflow was calculated. Clinic visits were scheduled on baseline day; on day 8 of four times daily pilocarpine (2%) to one eye and vehicle to the other; on day 8 of continued pilocarpine/vehicle treatment plus latanoprost (0.005%) once daily to both eyes; after a 3-week washout period; on day 8 of once-daily latanoprost to one eye and vehicle to the other; and on day 8 of continued latanoprost/vehicle treatment plus pilocarpine four times a day to both eyes. Drug-treated eyes were compared with contralateral vehicle-treated eyes and with baseline day by paired t tests. Combined pilocarpine and latanoprost-treated eyes were compared with individual drug-treated eyes and with baseline day using the Bonferroni test. RESULTS: Compared with baseline, pilocarpine reduced intraocular pressure from 18.9 to 16.2 mm Hg (P =.001) and increased outflow facility from 0.18 to 0.23 microl per minute per mm Hg (P =.03). No other parameters were affected. Adding latanoprost further reduced intraocular pressure to 13.7 mm Hg (P <.001) and increased uveoscleral outflow from 0.82 to 1.36 microl per minute (P =.02). Latanoprost alone reduced intraocular pressure from 17.6 to 14.3 mm Hg (P <.0001) and increased uveoscleral outflow from 0.89 to 1.25 microl per minute (P =.05). Adding pilocarpine to the latanoprost treatment further reduced intraocular pressure to 12.7 mm Hg (P <.001) and increased outflow facility from 0.21 to 0.30 microl per minute per mm Hg (P =.03). CONCLUSIONS: Latanoprost and pilocarpine predominantly increase uveoscleral outflow and outflow facility, respectively, when given alone. These drugs are additive because pilocarpine does not inhibit the uveoscleral outflow increase induced by latanoprost.

Aqueous humor dynamics in monkeys with laser-induced glaucoma.

This study determines the effects of laser-induced glaucoma on aqueous humor dynamics of 18 cynomolgus monkeys. Baseline measurements of 12 monkeys included intraocular pressure (IOP) by pneumatonometry, aqueous flow by fluorophotometry and outflow facility by tonography. Beginning 4 to 14 days later, the trabecular meshwork of one eye was treated repeatedly with laser photocoagulation until elevated IOP was induced. Thirty-six to 75 days after the last laser treatment, all measurements were repeated. Between 1.7 and 11.4 years after laser treatment, the same 12 monkeys plus 6 additional monkeys underwent IOP and aqueous flow measurements. In addition, outflow facility was determined with fluorophotometry, and uveoscleral outflow was both calculated (n=18) and measured with an intracameral tracer (n=7). In glaucoma eyes compared to control eyes (n=12), IOP was increased (p<0.04) by at least 8 mmHg at Time 1 (1 to 3 months) or Time 2 (3 to 4 years) after laser treatment; aqueous flow was reduced (p=0.0007) by 46% at Time 1 but returned to baseline levels at Time 2; tonographic outflow facility was reduced (p=0.0008) by 71% at Time 1. In lasered eyes compared to control eyes, fluorophotometric outflow facility was reduced (p=0.0008; n=18) by 63%, and uveoscleral outflow was increased (p<0.05), whether calculated or measured with tracers at least 1 year after laser treatment. The increased IOP in monkeys with laser-induced glaucoma was caused by a sustained reduction in outflow facility. The uveoscleral outflow increase was not enough to prevent the rise in IOP.

Efficacy and adverse effects of medications used in the treatment of glaucoma.

With the advent of several new topically active medications for glaucoma therapy, intraocular pressure (IOP) can be reduced to target levels in more patients before resorting to surgery. Some of these newer agents have a number of advantages over some of the older medications, several of which are seldom used now. The topically active carbonic anhydrase inhibitors are better tolerated than oral formulations, which are infrequently used despite their greater efficacy compared with the topical formulations. The alpha2-adrenergic agonists effectively reduce IOP with few systemic adverse effects. The prostaglandin analogues are even more effective and well tolerated when applied once daily without known systemic adverse effects. The variety of glaucoma medications forces the physician to be selective with various combinations before proceeding with surgery. This article critically reviews the literature pertaining to the newer glaucoma medications, thereby providing guidelines to make rational choices from among the available options.

Acute versus chronic effects of brimonidine on aqueous humor dynamics in ocular hypertensive patients.

PURPOSE: To report the acute vs chronic effects of brimonidine, a selective alpha2-adrenergic receptor agonist, on aqueous humor dynamics in ocular hypertensive patients. METHODS: Brimonidine 0.2% was given topically twice daily for 29 days to one eye each of 28 ocular hypertensive volunteers in a randomized double-masked study. The fellow eye was similarly treated with vehicle. Aqueous flow (Fa) and outflow facility (Cfl) were determined with fluorophotometry. Intraocular pressure, outflow facility (Cton), and episcleral venous pressure (Pev) were measured with pneumatonometry, tonography, and venomanometry, respectively. Uveoscleral outflow (Fu) was calculated from intraocular pressure, Fa, Pev, and Cfl values. All measurements were taken on baseline day, day 8, and day 29 of treatment. Intraocular pressure and Fa only were measured after instillation of 1 drop of brimonidine on day 1. RESULTS: When measured 3 hours after instillation on days 1, 8, and 29 of treatment, brimonidine significantly (P < .001) reduced intraocular pressure by at least 5.0 +/- 0.7 mm Hg (mean +/- SEM) compared with baseline day, and by 2.7 +/- 0.5 mm Hg compared with the vehicle-treated contralateral control eyes. The greatest decrease (6.0 +/- 0.6 mm Hg) was observed at 3 hours after the first drop. Aqueous flow was reduced by 29% (P < .001) after the first application but was not significantly different from baseline when measured at day 29 of treatment. Uveoscleral outflow was increased 60% at day 8 (P < .06) and day 29 (P < .05) compared with baseline. There was no significant difference in outflow facility or episcleral venous pressure at day 8 or day 29 of treatment. CONCLUSIONS: The brimonidine-induced reduction in intraocular pressure in humans is associated initially with a decrease in aqueous flow, and after chronic treatment with an increase in uveoscleral outflow.

Effects of topical epinephrine on aqueous humor dynamics in the cat.

The purpose of this study was to investigate, in cats, the effects of topical epinephrine on aqueous humor dynamics as measured by the non-invasive method of fluorophotometry and by other methods. Measurements were carried out on 12 cats before and after one week of twice daily treatment with 2% epinephrine hydrochloride to one eye. Aqueous flow and outflow facility were determined using fluorophotometry. Uveoscleral outflow was calculated from these results and was evaluated with anterior chamber perfusion of FITC-dextran. Outflow facility also was measured by tonography. Epinephrine-treated eyes, compared with their baseline values, showed a 31% reduction in intraocular pressure (P<0.001), a 23% reduction in aqueous flow (P<0.05), a 60% increase in fluorophotometric outflow facility (P<0.05), and a 43% increase in tonographic outflow facility (P<0.05). Treated eyes, compared with contralateral control eyes, showed a 27% reduction in IOP (P<0.005), a 25% reduction in aqueous flow (P<0.005), a 38% increase in fluorophotometric outflow facility (P<0.05), and a 34% increase in tonographic outflow facility. When evaluated by both fluorophotometry and FITC-dextran tracer methods, epinephrine had no significant effect on uveoscleral outflow. It was concluded that, in cats treated with topical epinephrine twice daily for a week, a reduction in intraocular pressure is induced by an increase in outflow facility and decrease in aqueous flow.

Aqueous humor dynamics in the aging human eye.

PURPOSE: Healthy subjects were recruited to identify normal, age-associated changes in intraocular pressure and aqueous humor dynamics. METHODS: Normal healthy subjects from two age groups were enrolled in the study: (1) those from 20 to 30 years of age (n = 51) and (2) those 60 years of age and older (n = 53). Intraocular pressure was measured by pneumatonometry, tonographic outflow facility by pneumatonography, and episcleral venous pressure by venomanometry. Aqueous flow and outflow facility were determined by a fluorophotometric technique. Uveoscleral outflow and anterior chamber volume were calculated. Results from the older group were compared with those from the younger group by means of unpaired, two-tailed t tests. RESULTS: Compared with the younger group, the older group showed significant differences as follows: smaller anterior chamber volume (160+/-39 vs. 247+/-39 microl; mean +/- SD; P< .00001), reduced aqueous flow (2.4+/-0.6 vs. 2.8+/-0.8 microl/minute; P = .002), and reduced uveoscleral outflow (1.10+/-0.81 vs. 1.52+/-0.81 microl/minute; P = .009). CONCLUSIONS: In the healthy aging eye, there is a reduction in the production of aqueous humor and a reduction in its drainage through the uveoscleral outflow pathway.

Prostaglandin-induced iris color darkening. An experimental model.

OBJECTIVES: To determine the role of sympathetic innervation and the effect of topical prostaglandin therapy on iris color in pigmented rabbits. METHODS: Twelve Dutch-belted rabbits underwent unilateral superior cervical ganglionectomy (SCGx) at age 1 to 3 months. A second group of 11 rabbits underwent bilateral SCGx at age 1 month and were treated once or twice daily for 6 to 9 months with 1 drop (about 20 microL) of latanoprost, 0.005%, to one eye and its vehicle to the contralateral eye. Standardized color photographs of the iris of each eye were taken at 1- to 2-month intervals for 6 to 10 months and evaluated by 4 to 6 observers in a masked fashion. RESULTS: At 8 to 10 months after unilateral SCGx, 11 of 12 rabbits showed definite heterochromia, with the lighter-colored iris on the SCGx side. Of the 11 rabbits that underwent bilateral SCGx and unilateral latanoprost treatment, 9 showed heterochromia at 6 to 9 months, with the darker-colored iris on the latanoprost-treated side. CONCLUSIONS: These results demonstrate that sympathetic innervation is required for age-related, physiologic darkening of iris color in rabbits, that prostaglandins may compensate for sympathetic denervation to produce darkening in SCGx eyes, and that this model may be useful to study prostaglandin-induced iris color change.

Bunazosin reduces intraocular pressure in rabbits by increasing uveoscleral outflow.

The mechanism of the ocular hypotensive effect of bunazosin hydrochloride (an alpha1-adrenergic antagonist) and the possible intermediary role of prostaglandins were studied in New Zealand albino rabbits. Aqueous flow, outflow facility and uveoscleral outflow were determined by fluorophotometry, and intraocular pressure (IOP) was measured by pneumatonometry on the fourth day of twice daily topical treatment with 0.1% bunazosin. Uveoscleral outflow was measured with a tracer infusion technique at 1 to 2 hours after one dose of 0.1% bunazosin. Total outflow facility was measured by a two-level constant-pressure infusion method before and at one hour after one dose of 0.1% bunazosin. The effect of topically applied cyclooxygenase inhibitors, including 0.25% indomethacin and 0.03% flurbiprofen, on the IOP reduction after bunazosin was evaluated. At 3 hours after the seventh consecutive dose given twice-daily, bunazosin significantly (P<0.001) reduced IOP to 13.4+/-0.8 mm Hg (mean +/- SEM) from a baseline of 19.6+/-1.1 mm Hg. Indomethacin significantly inhibited the IOP reduction after one dose of bunazosin, whereas flurbiprofen did not (repeated measures ANOVA). Bunazosin significantly increased uveoscleral outflow (P<0.05) and total outflow facility (P<0.02), but not fluorophotometric outflow facility or aqueous flow. It is concluded that, in rabbits, 0.1% bunazosin reduces IOP predominantly by increasing uveoscleral outflow. The role of prostaglandins in this effect is equivocal.

Effects of exogenous prostaglandins on aqueous humor dynamics and blood-aqueous barrier function.

Topical prostaglandins (PGs) are very effective at reducing intraocular pressure (IOP) in a variety of animals and in humans with relatively few side effects. The mechanisms of action of several PGs, their prodrugs and analogues have been studied in rabbits, cats, monkeys and humans. PGF2 alpha and its analogues evaluated in monkeys include PGF2 alpha-tromethamine salt, PGF2 alpha -isopropylester (-IE), S-1033, PhXA34, PhDH100A and latanoprost (PhXA41). Aqueous flow and outflow facility are either increased or remain unchanged by these agents. PGF2 alpha-IE, PHXA34, PhDH100A and latanoprost increase uveoscleral outflow, accounting for most of the IOP reduction. PGA2 in cats increases aqueous flow and outflow facility, but it reduces IOP primarily by stimulating uveoscleral outflow. The PGD2 analogue BW245C is unique in that it is the only PG that decreases aqueous flow. Mechanistic studies in humans have been performed with PGF2 alpha -IE, unoprostone, PhXA34 and latanoprost. In two clinical studies with latanoprost, a significant increase in uveoscleral outflow was found which, as in animals, accounts for most of the IOP reduction. A slight but inconsistent increase in outflow facility may also be involved. The doses tested had minimal effects on the permeability of the blood-aqueous barrier (BAB). In vitro studies of human tissue have been conducted to elucidate the PG effect on outflow facility and uveoscleral outflow. Studies of isolated human anterior segment preparations show that PGE2 increases outflow facility whereas PGF2 alpha has no measurable effect on this parameter. Studies of human ciliary muscle cells in tissue culture indicate that PGs may directly modulate extracellular matrix metabolism, which may be related to the increased uveoscleral drainage. This review summarizes in vitro and in vivo studies of the effects of PGs on aqueous humor dynamics and BAB integrity in humans, cats and monkeys.

Effects of brimonidine on aqueous humor dynamics in human eyes.

OBJECTIVE: To evaluate the mechanism by which brimonidine, a selective alpha 2-adrenergic agonist, lowers intraocular pressure (IOP) in humans. SUBJECTS: Twenty-one volunteers with ocular hypertension. METHODS: Brimonidine tartrate (0.2%) was given topically twice daily for 1 week to one eye in a randomized, double-masked study. The fellow eye was similarly treated with brimonidine vehicle. Before (baseline) and after 1 week (day 8) of dosing, IOP, aqueous flow, episcleral venous pressure, and tonographic outflow facility were directly measured. Fluorophotometric outflow facility and uveoscleral outflow were calculated. Brimonidine-treated eyes were compared with vehicle-treated contralateral control eyes and with baseline measurements after 1 week of dosing. RESULTS: Brimonidine significantly (P < .001, Student's two-tailed t test) reduced IOP mean +/- SE of 4.7 +/- 0.7 and 4.2 +/- 0.4 mm Hg compared with the baseline day and with the vehicle-treated contralateral control eyes, respectively. Compared with the baseline day, aqueous flow was reduced by 20% (P = .002) and uveoscleral outflow was increased (P = .04). A slight contralateral decrease in IOP of 1.2 +/- 0.6 mm Hg (P = .05) and in aqueous flow of 12% (P = .05) was noted. No significant difference was seen in the outflow facility values or episcleral venous pressure compared with the baseline day or with the contralateral control eye. CONCLUSIONS: The brimonidine-induced reduction in IOP in humans is associated with a decrease in aqueous flow and an increase in uveoscleral outflow. The decrease in IOP and aqueous flow in the contralateral control eye on day 8 compared with the baseline day suggests a mild contralateral effect.

Prostaglandin A2 increases uveoscleral outflow and trabecular outflow facility in the cat.

Prostaglandins (PG) are very effective ocular hypotensive agents. It is generally agreed that these drugs reduce intraocular pressure primarily by increasing uveoscleral outflow. They may also increase trabecular outflow facility though available evidence is less convincing. It has been hypothesized that PGs may increase facility of uveoscleral outflow in addition to their other mechanisms, but this has not yet been tested. To help clarify the ocular hypotensive mechanism of action of a derived PG of the A type, cats were treated twice daily for one week with PGA2 (0.01%) to one eye and vehicle to the other. Measurements were made of aqueous flow and outflow facility with fluorophotometry and of intraocular pressure with pneumatonometry. From these values, uveoscleral outflow was calculated. In addition, total outflow facility, uveoscleral outflow, and uveoscleral outflow facility were determined with invasive methods. PGA2 significantly reduced IOP by a mean of at least 4.7 mmHg in all experiments with all P-values less than 0.01. Compared with contralateral vehicle-treated control eyes, uveoscleral outflow in the treated eye was significantly (P < 0.05) increased by at least 50% using two different methods of measurement. Compared with baseline day, PGA2 significantly (P < or = 0.05) increased aqueous flow by 1.8 microliters min-1, fluorophotometric outflow facility by 0.36 microliter min-1 mmHg-1 and fluorophotometric uveoscleral outflow by 2.0 microliters min-1. Total outflow facility was not significantly different comparing treated with contralateral control eyes. Facility of uveoscleral outflow was < or = 0.02 microliters min-1 mmHg-1 for both control and treated eyes. It is concluded that PGA2 decreases IOP in cats by increasing uveoscleral outflow and trabecular outflow facility as measured with fluorophotometry. A significant increase in aqueous flow reduces the ocular hypotensive effect.

Effects of apraclonidine on aqueous humor dynamics in human eyes.

PURPOSE: The mechanism by which apraclonidine, an alpha 2-adrenergic agonist, lowers intraocular pressure (IOP) was evaluated in humans. METHODS: In a randomized, double-masked, placebo-controlled study, 0.5% apraclonidine was given topically twice daily for 1 week to one eye in each of 21 ocular hypertensive volunteers. The other eye was treated similarly with vehicle. Before and after 1 week of treatment, aqueous flow, uveoscleral outflow, fluorophotometric outflow facility, intraocular pressure, tonographic outflow facility, episcleral venous pressure, and outflow pressure were either directly measured or mathematically calculated. Values were compared in treated versus contralateral control eyes and on baseline versus day 8 of treatment. RESULTS: When compared with both contralateral control eyes and baseline day, fluorophotometric outflow facility in the apraclonidine-treated eyes increased by 0.09 to 0.10 microliter/minute/mmHg (P < 0.04), IOP decreased by 3.1 to 5.2 mmHg (P < 0.0001), and outflow pressure decreased by 3.3 to 4.2 mmHg (P < 0.0001). When compared with baseline day only, aqueous flow in the apraclonidine-treated eyes decreased by 0.3 microliter/minute (P < 0.04), and episcleral venous pressure decreased by 1.0 mmHg (P < 0.001). Episcleral venous pressure also decreased in the control eyes compared with baseline day by 1.3 mmHg (P < 0.001). When compared with contralateral control eyes only, uveoscleral outflow in the apraclonidine-treated eyes decreased by 0.47 microliter/minute (P < 0.03). Tonographic outflow facility showed no change when compared with either contralateral control eyes or baseline values. CONCLUSIONS: The apraclonidine-induced reduction in intraocular pressure was associated with an increase in fluorophotometric outflow facility, decrease in aqueous flow and decrease in episcleral venous pressure compared to baseline. The lack of a significant difference in aqueous flow and episcleral venous pressure between treated and contralateral control eyes may represent a contralateral drug effect.

Morphology of ganglion cells in the neotenous tiger salamander retina.

The morphology of retinal ganglion cells in the neotenous tiger salamander (Ambystoma tigrinum) was analyzed with the aid of morphometric techniques to determine the diversity of cell types and to evaluate the widely held notion that this form of Ambystoma has a simple retina, with little variance among its cell morphologies. Single-cell staining was achieved through retrograde labeling with horseradish peroxidase injected around the optic nerve sheath followed by a period of several days before tissue processing; 83 well-labelled cells with axons were studied in detail with light microscopy and a computer-aided reconstruction system. Five different morphological cell classes were devised based on broad morphometric criteria such as the dendritic area of influence; the number, length, and complexity of dendritic branches; and the amount of overlap between neighboring dendrites. These classes included small simple, small complex, medium simple, medium complex, and large cells. In addition, a class of cells with numerous varicosities among the dendrites was separately analyzed. These swellings did not stain for catecholamines. Based on optical determinations of the dendritic sublamination pattern within the inner plexiform layer, presumed On-Off cells are present in all subclasses, whereas On cells predominate in the smaller cell groups. Presumed Off cells are well represented in the large field units, although the small total number of cells in this latter class leads to uncertainty regarding the significance of this observation. The diversity of ganglion cell morphology revealed in the present study argues against the assumption that the neotenous tiger salamander has a simple retina, with a relatively invariant set of ganglion cells. On the contrary, it appears that this aquatic form shows morphological diversity in the retinal ganglion cell population rivaling that reported for other vertebrates, including mammals. A functional role for the different cell classes is briefly considered.

Effects of PhXA41, a new prostaglandin F2 alpha analog, on aqueous humor dynamics in human eyes.

PURPOSE: PhXA41, a new phenyl-substituted analog of a prostaglandin F2 alpha (PGF2 alpha) prodrug (13,14-dihydro-17-phenyl-18,19,20-trinor-prostaglandin F2 alpha-1-isopropyl ester), is an effective ocular hypotensive agent in patients with glaucoma. To understand its mechanism of action, various components of aqueous humor dynamics were examined after topical application to human eyes. METHODS: In a randomized, double-masked, placebo-controlled study, PhXA41 (0.006%) was given topically twice daily for 1 week to one eye each of 22 volunteers with normotension or ocular hypertension. The other eye was similarly treated with vehicle. Intraocular pressure (IOP) was measured by pneumatonometry and tonographic outflow facility by pneumatonography. Aqueous flow and outflow facility were determined either directly or indirectly by a fluorophotometric technique, and uveoscleral outflow was calculated secondarily. Comparison of values obtained in treated versus contralateral control eyes and on baseline versus day 8 of treatment were made. RESULTS: Compared with baseline measurements, PhXA41 significantly (P < 0.001) reduced IOP by 5.5 +/- 0.6 mmHg (mean +/- standard error of the mean) as measured 3 hours after the last dose on the eighth day of treatment. Aqueous flow, tonographic outflow facility, and fluorophotometric outflow facility were not changed by PhXA41. However, uveoscleral outflow was significantly greater in the PhXA41-treated eyes (0.87 +/- 0.22 microliter/minute) compared with either the contralateral vehicle-treated eyes (0.14 +/- 0.30; P < 0.02) or baseline measurements (0.39 +/- 0.20 microliter/minute; P < 0.05). CONCLUSIONS: PhXA41 decreases IOP in humans by increasing uveoscleral outflow without significantly affecting other parameters of aqueous humor dynamics.

Extravascular albumin concentration of the uvea.

The hypothesis that uveal vessels absorb fluid was tested by measuring the albumin in extravascular uveal tissues and in plasma. From these results the effective albumin concentration was calculated in both rabbits and monkeys. Three separate methods were used to measure uveal albumin, and the results of these were compared. In method 1, the intravenous fluorescein isothiocyanate (FITC)-albumin concentration found in the uvea 5 min after injection (intravascular tracer) was subtracted from that found 2 hr after injection (intravascular plus extravascular tracer) to determine the extravascular albumin concentration. In method 2, intravenous FITC-albumin was followed by vascular washout after a 2-hr equilibration period to determine extravascular uveal albumin. In method 3, the endogenous extravascular albumin concentration of uveal tissues was measured with an enzyme-linked immunosorbent assay (ELISA) after vascular washout. The effective albumin concentration was determined by dividing the data in methods 1, 2, and 3 by the extravascular albumin space volume. The effective albumin concentration in monkey (as percentage of plasma) was, for methods 1, 2, and 3: iris 2, 3, and 4%; pars plicata 14, 12, and 7%; pars plana 2, 10, and 12%; and choroid 2, 12, and 10%, respectively. In rabbit, the extravascular albumin concentrations were: iris 10, 21, and 7%; pars plicata 69, 26, and 39%; pars plana 41, 46, and 10%; and choroid 88, 30, and 26%, respectively. These findings are lower than previously reported in rabbits, yet are consistent with previous estimates in monkeys. These results support the hypothesis that uveal vessels are capable of fluid absorption, since a large colloid osmotic gradient exists across the vessel wall.

Hydrostatic pressure of the suprachoroidal space.

The hydrostatic pressure of the suprachoroidal space was measured in 18 cynomolgus monkey eyes by one of two methods: (1) direct cannulation, or (2) silicone sponge implantation. The intraocular pressure (IOP) and suprachoroidal pressure were monitored simultaneously with the IOP being held at various levels between 5 and 60 mm Hg. In eyes with direct cannulation, at an IOP of 15 mm Hg, the pressure in the anterior suprachoroidal (supraciliary) space was 0.8 +/- 0.2 mm Hg (n = 6, mean +/- SE) below the IOP, but the posterior suprachoroidal pressure was 3.7 +/- 0.4 mm Hg (n = 8) below the IOP. The suprachoroidal pressure in eyes with silicone sponge implant was 4.7 +/- 0.6 (n = 7) mm Hg below the IOP. A change in IOP produced a corresponding change in the supraciliary space pressure. However, the pressure difference between the anterior chamber and the posterior suprachoroidal space increased at higher IOP. This pressure differential is the driving force for uveoscleral outflow.

Functional recovery of retinal pigment epithelial damage in experimental retinal detachment.

The integrity of the RPE barrier function in retinal detachment was studied in vitro. The retinal pigment epithelium (RPE)-choroid tissue was isolated from cynomolgus monkey eyes with acute (less than 1 hr), subacute (1-2 weeks), and chronic (8-20 months) retinal detachments, and clamped between Ussing-type chambers. Electrical characteristics and choroid-to-retina permeability to carboxyfluorescein were determined. In the HEPES-buffered bathing solution, transepithelial potential difference and resistance in eyes with acute retinal detachments (0.2 mV and 134 ohm-cm2, respectively) were significantly lower than subacute (7.9 and 350) and chronic (10.4 and 348) retinal detachments. Furthermore, the permeability was increased five-fold in acute retinal detachments with respect to subacute and chronic retinal detachments, indicating a breakdown of the RPE barrier in acute retinal detachment. No statistical difference was found between subacute and chronic retinal detachments. In this animal model, RPE barrier function is destroyed at the onset of retinal detachment, but recovers in a week or two, and is maintained in the chronic stage. Histological examination revealed that RPE recovery was accomplished by RPE proliferation and hyperplasia.