The effect of dopamine on the intraocular pressure and pupil of the rabbit eye

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1 The effect of dopamine on the intraocular pressure and pupil of the rabbit eye Richard P. Shannon,* Alden Mead, and Marvin L. Sears The presence of a dopamine-specific receptor that can influence intraocular pressure in the rabbit eye is suggested by these experiments: intravitreal, systemic, or topically administered dopamine solutions can produce a decrease in intraocular pressure that is dose-dependent. In high doses the decrease in intraocular pressure is accompanied by mydriasis, an alpha-adrenergic pupillary response. This alpha-response may be caused by dopamine as a direct, but weak alpha-stimulator or by displacement of norepinephrine onto the receptor from its storage site. At lower doses the decrease in intraocular pressure occurs in the absence of mydriasis and in spite of blockade of beta-adrenergic receptors. Further, a specific dopaminergic blocker, haloperidol, prevents this decrease in intraocular pressure. The decreases are small, but reproducible and suggest that dopamine can influence the adrenergic regulation of intraocular pressure. -Lhe role of dopamine as the adrenergic neurotransmitter in the mammalian extrapyramidal system has been described. 1 ' 3 Other dopamine systems have unique pharmacological properties that distinguish them from alpha- and beta-adrenergic receptors and from nicotinic and muscarinic From the Department of Ophthalmology and Visual Science, Yale University School of Medicine, New Haven, Conn "The work was done while Mr. Shannon was on a summer fellowship in our department in 1974 and This work was supported in part by United States Public Health Service No. EY-00237, EY-00785, The Connecticut Lions Eye Research Foundation, Inc., and Research to Prevent Blindness, Inc. Submitted for publication Nov. 18, Reprint requests: Dr. M. L. Sears, Department of Ophthalmology and Visual Science, Yale University School of Medicine, 333 Cedar St., New Haven, Conn cholinergic systems. Specific dopamine receptors have been found in the isolated perfused canine pancreas, 40 and in the renal and mesentric circulation. 7 ' 8 The results obtained in these organ systems suggested a need for study of dopamine in the eye. Dopamine has long since been identified as the most prominent catecholamine in the retina (see review 9 ). Circumstantial evidence bearing on the role of dopamine as a modulator of intraretinal synaptic transmission has accumulated. 10 Dopamine has also been shown to activate adenyl cyclase in the whole calf or rat retina. 11 It has also been reported that 1-dopa and dopamine cause mydriasis in the normally innervated iris of the human eye The purpose of this study was to learn whether dopamine has any effect on the intraocular pressure of the rabbit, and to decide what sort of receptors are activated by dopamine.

2 372 Shannon, Mead, and Sears Investigative Ophthalmology May 1976 Materials and methods Animals. Male albino New Zealand rabbits weighing between 2 and 3 kilograms were used. Intraocular pressure. Intraocular pressure was measured with a pneumotonometer calibrated against water. The animals were gently restrained in a canvas wrap and two drops of 0.1 per cent ophthalmic solution were administered prior to each measurement. All measurements were taken by the same individual. Pupil size. Pupil size was estimated with a millimeter ruler by measuring horizontal and vertical diameters under constant low-level laboratory illumination. Ganglionectomy. Sympathetic denervation was accomplished by the surgical removal of the cervical ganglion. The surgery was performed under 50 mg. (20 mg. per kilogram) pentobarbitol anesthesia administered through the marginal ear vein. Denervated animals were used for study usually eight days after ganglionectomy. Blood pressure. Cannulation of the right common carotid artery was performed under pentobarbitol anesthesia (20 mg. per kilogram). The cannula was attached by a system of stopcocks to a pressure transducer (Hewlett Packard, Series 947), and amplified and recorded on a Sanborn system. One-hundred to three-hundred units of sodium heparin were used initially to prevent coagulation. Glaucoma rabbits. A colony of these with glaucoma of two year's duration was available. 15 Drug administration. Dopainine (intravitreal). Intravitreal injections of 3-hydroxytyramine HC1, dopamine (Sigma Chemical Company) solutions were given to the conscious animal while the animal was restrained in a close fitting canvas wrap. Two drops of 0.5 per cent ophthalmic solution were applied to the equatorial area of the sclera and the globe gently proptosed. A 30-gauge needle attached to a Hamilton No. 701 microsyringe was carefully introduced into the center of the vitreous body through the superior temporal quadrant. Ten microliters of a 0.1 per cent, 1.0 per cent, and 10 per cent solutions of dopamine were injected into experimental eyes. Ten microliters of 0.9 per cent normal saline was injected into the vitreous body in contralateral eyes as a control. Pressure and pupil measurements were then recorded every 30 minutes for an eight hour period. Dopamine (systemic). 3-Hydroxytyramine was administered systemically by constant intravenous infusion into a marginal ear vein. The rate of infusion was 250 /xl per minute for 20 minutes via a 23-gauge butterfly infusion set (Abbott Laboratories). The concentrations administered were 2 /xg per kilogram per minte, 5 jug per kilogram per minute, 10 jug per kilogram per minute, and 20 fig per kilogram per minute. Between 100 and 300 units of sodium heparin (Bel-Mar Laboratories, Inc.) were employed to prevent coagulation over the 20-minute period. Pressure and pupil measurements were recorded every 30 minutes for a four hour period. Dopamine (topical). One hundred microliters of a 10 per cent solution of dopamine were applied in a double-blind fashion to one cornea of each animal. One hundred microliters of 0.9 per cent normal saline served as a control in the contralateral eye. Pressure and pupil measurements were recorded every five minutes for a 45-minute period after topical administration. Haloperidol. Intramuscular injections of 1.5 mg. of Haldol (McNeil Laboratories, Inc.) were given two hours prior to intravitreal administration of dopamine. Eye pressure and pupil diameter were then measured every 30 minutes for six hours. Propranolol. Propranolol HC1 (Sigma Chemical Company) was administered by intravenous injection of a dosage of 5 mg. per kilogram two hours prior to the intravitreal injections of dopamine. Solutions were freshly prepared each day in sterile 0.9 per cent normal saline and subjected to Millipore filtration (1.2 JM). Statistics. Results are expressed as the mean ± S.E.M. Student's t-test was employed to determine the level of significance. Results Blood pressure. Transient changes in blood pressure were induced by intravenous administration of low and high doses of dopamine in amounts of 100 /xg and 1 mg. and by intravenous haloperidol (1.5 mg.). The effects of even the largest doses of dopamine and haloperidol on blood pressure were of very short duration. Two hours were therefore considered sufficient to allow for pretreated animals to reach steady-state in the ocular tests reported. Topical dopamine. The effect of the topical application of 100 pi of 10 per cent dopamine on the intraocular pressure is shown in Fig. 1. In the normal and glaucomatous animals, a significant decrease (P < 0.01) is observed in the eye treated with dopamine. Fig. 1 shows that there is a steady decrease in outflow pressure over the first 30 minutes before it begins to return to baseline levels. Topical dopamine also caused a significant ipsilateral k.

3 Volume 15 Number 5 Dopamine effect on intraocular pressure 373 Normal 100 o Gloucomatout Minutes After Topical Administration Of 10% Oopamine Fig. 1. Twenty to thirty minutes after topical dopamine there occurs, a 20 per cent decrease in outflow pressure in the normal eye and in the eye of rabbits with impaired outflow facility. Table I. Intraocular pressure after intravitreal dopamine in six animals Time (hours) Control 1 mg. Experiment ± ± ± ± ± ± ± ± ± 0.3 Values represent the mean ± S.E.M. Control ± ± ± ± 0.4 Dosage WO vg Experiment 18.9 ± ± ± ± ± 0.4 Control 18.5 ± ± ± ± ± ng Experiment ± ± ± ± 0.4 mydriasis which occurs within 10 minutes and lasts for several hours. Intravitreal dopamine. Figs. 2 and 3 show the effects of an intravitreal injection of dopamine on intraocular pressure and pupil diameter. At all doses the difference in intraocular pressure between the control eye, injected with saline, and the experimental eye, treated with 10 /d of a dopamine solution, was statistically significant (P < 0.01). A significant increase in pupil diameter in response to intravitreal dopamine was found only after an injection of 10 /x\ of a 10 per cent solution. Doses of 1 per cent and 0.1 per cent had a significant effect on intraocular pressure but had no effect on pupil size over the eight hour period. Table I shows that the onset of a statistically significant difference in intraocular pressure (P < 0.01) was first observed between three and four hours for concentrations of 10 per cent and 1 per cent. A significant decrease in intraocular pressure at a concentration of 0.1 per cent was first observed at five hours and lasted two hours. With concentrations of 1 per cent and 10 per cent, the intraocular pressure decrease in the eye treated with dopamine persisted for eight and twelve hours, respectively. Fig. 3 demonstrates the time course of the pupil response to intravitreal injections of 10 /J of 10 per cent dopamine. A pronounced unilateral mydriasis is observable 45 minutes after the injection and the pupil remains dilated over an eight hour period. Fig. 4 is the dose-response curve of the

4 374 Shannon, Mead, and Sears Investigative Ophthalmology May \00fiQ mg Hours After Dopamine Fig. 2. After intravitreal dopamine a progressive dose related decrease in outflow pressure occurs Hours After Injection Fig. 3. An effective increase in pupil diameter was only noted after intravitreal injection of 1.0 mg. of dopamine. intraocular pressure at a time when the decrease was maximum. Systemic dopamine. Table II demonstrates the intraocular pressure changes after systemic administration of dopamine. A significant decrease in intraocular pressure is observed after intravenous infusion of dopamine. The time course is shown on Fig. 5. There was no significant change in pupil size after relatively large doses of systemic dopamine (20 /*g per kilogram per minute

5 Volume 15 Number 5 Dopamine effect on intraocular pressure 375 intravitreal x intravenous o topical UJ 2 LOG DOPAMINE Fig. 4. The dose-related decrease in intraocular pressure for three modes of administration is shown. All routes produce small but reproducible decreases of a comparable nature, but the intravitreal and intravenous routes are quite similar in their effect. Table II. Intraocular pressure after intravenous dopamine Intraocular pressure(mm. Hg) Time (hours) Baseline ng/kg./min ±0.3 (4) 17.5 ± ± ± ± ng/kg./min. \ 19.5 ±0.6 (4) 17.8 ± ± ± ± ng/kg./min ±0.5 (4) 16.6 ± ± ± ± ng/kg./min ±0.4 (4) 15.6 ± ± ± ± » about 1,000 /xg). The time course of the intraocular pressure decrease after systemic dopamine indicates that as the dose is increased the onset of the decrease in intraocular pressure occurs more rapidly and is more prolonged in duration. The doseresponse relationship for the maximum effect after systemic administration of dopamine is depicted in Fig. 4. At all concentrations the intraocular pressure started toward pretreatment levels within four hours after systemic infusion. The time course of the intraocular pressure change in the glaucomatous is similar to that in the normal eye. As the dosage of dopamine is increased, the onset of a significant decrease in intraocular pressure occurs more rapidly and is of a longer duration. In all cases, with the exception of the largest concentration, the intraocular pressure begins to return to pretreatment levels within four hours after infusion. Propranolol pretreatment. Table III describes the effect of propranolol, 5 mg. per kilogram, on the dopamine-induced reduction of intraocular pressure via intravitreal injection. Propranolol does not prevent the decrease in intraocular pressure seen after administration of dopamine, but the maximum response occurs a little later, between three and five hours compared with two hours in the untreated. As would be expected with a low dose of dopamine, there was no significant difference in pupil size between the control and treated eye. When mydriasis occurred with higher doses, the beta-blocker had no effect on the alpha stimulus.

6 376 Shannon, Mead, and Sears Investigative Ophthalmology May t 2/^g/kg/min 10/*g/kg/min Hours After 20 Minute Infusion Fig. 5. Intravenous dopamine infusions produce dose-related decreases in intraocular pressure. Decrease in outflow pressure reaches a maximum at about two hours. After a 20-minute infusion, a 20 to 30 per cent decrease in outflow pressure is apparent. Table III. Intraocular pressure after 100 fxg intravitreal dopamine two hours after pretreatment with intravenous propranolol Time Baseline Control 16.9 ±0.6 (6) 16.8 ± ± ± ± ± ± ± 0.5 Treated 17.6 ± 0.5 (6) 14.5 ± ± ± ± ± ± ± 0.8 Effects of haloperidol. Animals were pretreated with the specific dopaminergic receptor blocker, haloperidol, intramuscularly in doses of 1.5 mg. Fig. 6 illustrates the difference between the per cent decrease in outflow pressure in response to 500 fxg of intravitreal dopamine in normal animals (no haloperidol) and animals pretreated with haloperidol. Two hours after haloperidol administration, 10 ju.1 unilateral injections of dopamine were delivered into one vitreous body. Ten microliters of normal saline served as a contralateral control. Table IV shows that there was no significant difference in intraocular pressure in the control eye and the eye treated with dopamine. Mydriasis in response to the 500 jug of dopamine was not blocked by haloperidol, however, and persisted for four hours. Fig. 7 shows the absence of any dopamine-induced effect on pressure, but the persistence of mydriasis in response to 500 /tg of dopamine after intravenous injection of haloperidol. Effect of denervation on responses to dopamine. Eight days after unilateral denervation, both eyes received intravitreal injections of 10 /xl of varying concentrations of dopamine. One hundred micrograms of dopamine produced slight and equal reductions in intraocular pressure in denervated and normally innervated eyes [17.8 ± 0.3 (6) to 15.8 ± 0.6 (6)] in the former, and [17.9 ±0.4 (6) to 16.5 ± 0.3 (6)] in the latter, while pupil size was unaffected. Larger doses, e.g., 500 /xg, also produced equal but larger reductions (than 100 /xg) in intraocular pressure in both denervated and normally innervated eyes (6) [18.3 ± 0.3 to 15.2 ±

7 Volume 15 Number 5 Dopamine effect on intraocular pressure 377 ( T I I ^ { haloparidol pr«-tr«atmtnt _ \ \ \ T \T T 50 - no pr«-tr«atm«nt i i Hours After Dopamine Fig. 6. Pretreatment of rabbits with haloperidol prevents the decrease in intraocular pressure and outflow pressure seen with intravitreal injection of 100 jug of dopamine (see Table IV). i I I Table IV. Intraocular pressure and pupil size after intravitreal injection of 10 pi of a 5 per cent solution of dopamine. Pretreatment with haloperidol, 1.5 mg. was done two hours before Time (hours) Baseline Intraocular pressure (mm. Hg) Left Right 16.4 ± 0.5 (8) 16.2 ± ± ± ± ± ± ±0.4 (8) 17.0 ± ± ± ± ± ± 0.4 Pupil diameter (mm.) Left Right 5.3 ±0.2 (8) 5.5 ± ± ± ± ± ± ±0.2 (8) 6.3 ± ± ± ± ± ± 0.3 All values are expressed as the mean ± S.E.M. (n = 8). The left eye served as the saline control eye. The right eye received dopamine. 0.5 and 18.3 ± 0.3 to 15.0 ± 0.5] and also provoked mydriasis in both. Discussion Intravitreal, systemic, or topical administration of dopamine causes a decrease in intraocular pressure that is dose-related. Dopamine administered by intravitreal injection in amounts greater than 300 /xg, decreased intraocular pressure and produced mydriasis. At lower doses this alphaadrenergic pupillary response is not observed but significant decreases in intraocular pressure still occur. It appears then that more than one receptor can account for the effect of dopamine. For example, intravenous doses of 1 to 2 fig per kilogram per minute dopamine dilate mesenteric and renal blood vessels through stimulation of a dopamine-specific receptor. At higher doses in the range of 2 to 10 fig per kilogram per minute, dopamine also stimulates beta-adrenergic receptors in the heart. At doses greater than 10 fig per kilogram per minute, however, dopamine has been shown to produce generalized vasoconstriction through stimulation of alpha-adrenergic receptors. 7 To decide in a preliminary way whether the action of dopamine on intraocular

8 378 Shannon, Mead, and Sears Investigative Ophthalmology May r Solin* 18 - >: o ^. Q Q ->^ Dopomin* ~ 8 E L Dopamin* 57-0> E o ^ 6 ^ ^ ^ ~~~*~ m Salin* 5 - i 1 1 I Hours After Dopamine Fig. 7. In the haloperidol-treated animal intravitreal dopamine cannot reduce intraocular pressure but mydriasis still occurs. pressure at low to moderate doses was mediated through a dopamine-specific receptor or via beta-adrenergic reception, intravitreal dopamine was administered unilaterally in moderate doses, (100 fig),** to animals pretreated with propranolol. Propranolol may be an effective betablocker when given intravenously to rabbits in doses of 5 mg per kilogram. 16 In the presence of beta-blockade, the decrease in intraocular pressure after dopamine (Table III) still occurred and suggests the existence of a dopamine-specific receptor independent of either an alpha- or betaadrenergic response. This conclusion is supported by the finding that haloperidol, a specific dopamine receptor blocker, significantly diminished the intraocular pressure decrease after intravitreal dopamine (Table IV, Fig. 6). The action of 1-dopa or dopamine on the human iris causing alpha-mediated pupil One hundred micrograms in the vitreous is about 10"» molar final concentration assuming a volume of 4 ml. foi vitreous. dilation was attributed to dopamine displacing norepinephrine from adrenergic nerve terminals. 12 This view is probably correct because in human Horner's syndrome such dilation did not occur. 14 In our studies, the pronounced mydriasis observed in the denervated eye after 500 (xg of intravitreal dopamine must be due to the direct action of dopamine itself on the dilator pupillae muscle. This observation confirms Van Rossum's 17 finding that dopamine is a weak alpha-stimulator, effective only in high doses. Intravitreal dopamine in lower doses causes mydriasis in the normally innervated eye, but not in the denervated eye, because dopamine can still work by displacing norepinephrine. No mydriasis in the denervated eye occurs because without norepinephrine stores only direct stimulation of alphareceptors would work, and therefore higher concentrations for weak nonsupersensitive receptors are required. It can therefore be concluded that the action of dopamine at alpha-adrenergic sites may be a displace-

9 Volume 15 Number 5 Dopamine effect on intraocular pressure 379 ment effect in moderate doses, but indicates that dopamine in very large doses can stimulate alpha-adrenergic receptors directly. Dopamine in moderate doses produced the same decrease in intraocular pressure and mydriasis in denervated and normally innervated eyes. Thus, dopaminergic effects do not exhibit the enhanced effects associated with supersensitivity denervation. Significant decreases in intraocular pressure in glaucomatous eyes with impaired outflow facility were found after systemic and local administration of dopamine. This would indicate, although not prove, that dopamine can exert its effect on pressure by reducing inflow. Further studies are of course indicated to decide whether dopamine can affect either inflow or outflow of aqueous, or both. It is necessary to employ high doses of dopamine by a systemic route to obtain mydriasis, an alpha-adrenergic response. At lower doses, in the absence of mydriasis, the decreased intraocular pressure after systemic dopamine could be considered a beta-adrenergic function. Since the decrease in intraocular pressure occurred in the propranolol-treated animal but not in the haloperidol animal, a specific dopaminergic receptor is likely. The presence of a dopamine-specific response in the normal eye, in the absence of an alpha- or beta-adrenergic response, elicited via the blood circulation, taken together with the reduction in pressure (in steady-state) in eyes with impaired outflow, raises the possibility that such reception has a regulatory function, perhaps on aqueous humor formation. In this connection, it should be recalled that uptake of 1-dopa by the ciliary epithelium was demonstrated by Ehinger and Falck 18 using fluorescence microscopy. These extra neuronal uptakes were weak by comparison with neuronal uptakes for norepinephrine and epinephrine. Whether these nonneuronal uptakes represent some specific functional capacity in the tissue is not certain but taken together with the current work suggests a possible locus of action for dopamine. REFERENCES 1. Hornykiewicz, O.: Dopamine (3-hydroxytyramine) and brain function, Pharmacol. Rev. 18: 925, Iverson, L.: Dopamine receptors in the brain, Science 188: 1084, Miller, R., Horn, S., and Iverson, L.: The action of neuroleptic drugs on dopaminestimulated adenosine cyclic 3',5'-monophosphate production in rat neostriatum and limbic forebrain, Molec. Pharmacol. 10: 759, Hashimoto, K., Satoh, S., and Takeuchi, O.: Effects of dopamine on the pancreatic secretion in the dog, Br. J. Pharmacol. 43: 739, Furuta, Y., Hashimoto, K., Ishii, Y., et al.: Modification by drugs of the secretogogue effect of dopamine on the pancreas, Br. J. Pharmacol. 51: 225, Furuta, Y., Hashimoto, K., Iwatsuki, K., et al.: Effects of enzyme inhibitors of catecholamine metabolism and of haloperidol on the pancreatic secretion induced by 1-dopa and by dopamine in dogs, Br. J. Pharmacol. 47: 77, McNay, J. L., and Goldberg, L. I.: Comparison of the effects of dopamine, isoproterenol, norepinephrine and bradykinin on renal and femoral blood flow, J. Pharm. Exp. Ther. 151: 23, Goldberg, L. I.: Cardiovascular and renal actions of dopamine: potential clinical applications, Pharmacol. Rev. 24: 1, Sears, M. L.: Catecholamines in relation to the eye. In: Handbook of Physiology, Section on Endocrinology, Astwood, E., and Creep, R., editors. American Physiological Society, 1975, chap. 35, pp Kramer, S. G.: Dopamine: a retinal neurotransmitter, INVEST. OPHTHALMOL. 10: 438, Brown, J. H., and Makman, M.: Stimulation by dopamine of adenylate cyclase in retinal homogenates and of adenosine-3',5'-cyclic monophosphate formation in the intact retina, Proc. Nat. Acad. Sci. 49: 539, Spiers, A. S., and Calne, D. B.: Action of dopamine on the human iris, Br. Med. J. 4: 333, Spiers, A. S. D., Calne, D. B., Vakil, S. D., et al.: Action of thymoxamine on mydriasis induced by levodopa and dopamine, Br. Med. J. 2: 438, Weintraub, M. I., Gaasterland, D., and Van Woert, M. H.: Pupillary effects of levodopa

10 380 Shannon, Mead, and Sears investigative Ophthalmology May 1976 therapy. Development of anisocoria in latent 17. Van Rossum, J. M.: Different types of sympa- Horner's syndrome, N. Engl. J. Med. 283: thomimetic alpha-receptors, J. Pharm. Phar- 102, macol. 17: 202, Sears, D. E., and Sears, M. L.: Blood aqueous 18. Ehinger, B., and Falck, B.: Cellular uptake of barrier and alpha-chymotrypsin glaucoma in some amino acids and amines in vitro into rabbits, Am. J. Ophthalmol. 77: 378, rabbit and monkey anterior eye segment pre- 16. Langham, M. E., and Diggs, E. M.: Beta- parations, Exp. Eye Res. 10: 352, adrenergic responses in the eyes of rabbits, primates and man, Exp. Eye Res. 19: 281, 1974.