[Skip to Content]
Access to paid content on this site is currently suspended due to excessive activity being detected from your IP address 23.23.54.109. Please contact the publisher to request reinstatement.
[Skip to Content Landing]
Download PDF
Irides of 2 Dutch-belted rabbits. Top, Eyes of a rabbit 6 months after surgical removal of the right unilateral superior cervical ganglion. The right eye (A) was judged to be lighter in color than the left eye (B) by 6 masked observers. Bottom, Eyes of a rabbit 6 months after the surgical removal of both superior cervical ganglia. The left eye (D) had been treated with vehicle and the right eye (C) with 0.005% latanoprost, once or twice daily for 6 months after the surgery. The right eye was judged to be darker than the left eye by 6 masked observers.

Irides of 2 Dutch-belted rabbits. Top, Eyes of a rabbit 6 months after surgical removal of the right unilateral superior cervical ganglion. The right eye (A) was judged to be lighter in color than the left eye (B) by 6 masked observers. Bottom, Eyes of a rabbit 6 months after the surgical removal of both superior cervical ganglia. The left eye (D) had been treated with vehicle and the right eye (C) with 0.005% latanoprost, once or twice daily for 6 months after the surgery. The right eye was judged to be darker than the left eye by 6 masked observers.

Effects of Unilateral Latanoprost Treatment on Iris Color in 11 Rabbits Following Bilateral Superior Cervical Ganglionectomy (Group 2)*
Effects of Unilateral Latanoprost Treatment on Iris Color in 11 Rabbits Following Bilateral Superior Cervical Ganglionectomy (Group 2)*
1.
Camras  CB Prostaglandins and the treatment of the glaucomas. Ritch  RShields  MBKrupin  Teds.The Glaucomas. St Louis, Mo Mosby–Year Book Inc1996;1449- 1461
2.
Camras  CBThe US Latanoprost Study Group, Comparison of latanoprost and timolol in patients with ocular hypertension and glaucoma: a 6-month, masked, multicenter trial in the United States. Ophthalmology. 1996;103138- 147Article
3.
Camras  CBAlm  AWatson  PGStjernschantz  JThe Latanoprost Study Groups, Latanoprost, a prostaglandin analog, for glaucoma therapy: efficacy and safety after 1 year of treatment in 198 patients. Ophthalmology. 1996;1031916- 1924Article
4.
Alm  AStjernschantz  JThe Scandinavian Latanoprost Study Group, Effects on intraocular pressure and side-effects of 0.005% latanoprost applied once daily, evening or morning: a comparison with timolol. Ophthalmology. 1995;1021743- 1752Article
5.
Watson  PGStjernschantz  JThe Latanoprost Study Group, A 6-month, randomized, double-masked study comparing latanoprost with timolol in open-angle glaucoma and ocular hypertension. Ophthalmology. 1996;103126- 137Article
6.
Wistrand  PStjernschantz  JOlsson  K The incidence and time-course of latanoprost-induced iridial pigmentation as a function of eye color. Surv Ophthalmol. 1997;41(suppl 2)S129- S138Article
7.
Johnstone  MA Hypertrichosis and increased pigmentation of eyelashes and adjacent hair in the region of the ipsilateral eyelids of patients treated with unilateral topical latanoprost. Am J Ophthalmol. 1997;124544- 547
8.
Wand  M Latanoprost and hyperpigmentation of eyelashes. Arch Ophthalmol. 1997;1151206- 1208Article
9.
Selén  GStjernschantz  JResul  B Prostaglandin-induced iridial pigmentation in primates. Surv Ophthalmol. 1997;41(suppl 2)S125- S128Article
10.
Tomita  YMaeda  KTagami  H Melanocyte-stimulating properties of arachidonic acid metabolites: possible role in postinflammatory pigmentation. Pigment Cell Res. 1992;5357- 361Article
11.
Eagle  RC  Jr Iris pigmentation and pigmented lesions: an ultrastructural study. Trans Am Ophthalmol Soc. 1988;88581- 687
12.
Wilkerson  CLSyed  NAFisher  MRRobinson  NLWallow  IHLAlbert  DM Melanocytes and iris color: light microscopic findings. Arch Ophthalmol. 1996;114437- 442Article
13.
Imesch  PDBindley  CDKhademian  Z  et al.  Melanocytes and iris color: electron microscopic findings. Arch Ophthalmol. 1996;114443- 447Article
14.
Calhoun  FP Causes of heterochromia iridis with special reference to paralysis of the cervical sympathetic. Am J Ophthalmol. 1919;2255- 269
15.
Calhoun  FP A consideration of the causes of heterochromia iridis, with special reference to a paralysis of the cervical sympathetic. Trans Am Ophthalmol Soc. 1919;16277- 312
16.
Laties  AM Ocular melanin and the adrenergic innervation to the eye. Trans Am Ophthalmol Soc. 1974;72560- 605
17.
Bito  LZ Prostaglandins: a new approach to glaucoma management with a new, intriguing side effect. Surv Ophthalmol. 1997;41(suppl 2)S1- S14Article
18.
Diesenhouse  MCPalay  DANewman  NJTo  KAlbert  DM Acquired heterochromia with Horner syndrome in 2 adults. Ophthalmology. 1992;991815- 1817Article
19.
Odin  LO'Donnell  FE  Jr Adrenergic influence on iris stromal pigmentation: evidence for alpha-adrenergic receptors. Invest Ophthalmol Vis Sci. 1982;23528- 530
20.
Camras  CBPodos  SM The role of endogenous prostaglandin in clinically-used and investigational glaucoma therapy. Bito  LZStjernschantz  Jeds.The Ocular Effects of Prostaglandins and Other Eicosanoids. New York, NY Alan R Liss Inc1989;312459- 475
21.
Camras  CBZhan  G-LOhia  SOhia  EWang  Y-L The role of prostaglandins and neuropeptide Y in the reduction of intraocular pressure 1 day after sympathectomy in rabbits. Invest Ophthalmol Vis Sci. 1993;34(suppl)932
22.
Laties  AMLerner  AB Iris colour and relationship of tyrosinase activity to adrenergic innervation. Nature. 1975;255152- 153Article
23.
Bito  LZMatheny  APCruickshanks  KJNondahl  DMCarino  OB Eye color changes past childhood: the Louisville Twin Study. Arch Ophthalmol. 1997;115659- 663Article
24.
Dryja  TPAlbert  DM Lack of adrenergic influence on the pigmentation of iris nevus cells. Arch Ophthalmol. 1980;981996Article
Laboratory Sciences
August 1998

Prostaglandin-Induced Iris Color DarkeningAn Experimental Model

Author Affiliations

From the Department of Ophthalmology, University of Nebraska Medical Center, Omaha (Drs Zhan, Toris, Camras, and Wang); and the Department of Ophthalmology, Columbia University, New York, NY (Dr Bito). Drs Bito and Camras are consultants to Pharmacia and Upjohn, the company that developed latanoprost. Dr Bito has a proprietary interest in the use of prostaglandins for the medical management of glaucoma through patents he has assigned to Columbia University, New York, NY, according to the policies of the US Public Health Service, National Institutes of Health, Bethesda, Md, which supported the research program of his laboratory in this field between 1970 and 1989.

Arch Ophthalmol. 1998;116(8):1065-1068. doi:10.1001/archopht.116.8.1065
Abstract

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 µL) 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.

RECENT REPORTS demonstrate that with long-term latanoprost treatment, some patients have darkening of iris color16 and eyelashes.4,7,8 In those patients exhibiting iris color changes, the baseline iris color was described as green-brown, yellow-brown, or blue/gray-brown, with a darker color around the sphincter than in the periphery. Latanoprost treatment caused darkening of the lighter-colored peripheral iris, resulting in a more uniformly brown color. Iris color darkening was not observed in any patient who had a uniformly blue or brown iris color at baseline.16

The precise mechanism of this iris color darkening is unknown. Iris color darkening occurs in cynomolgus monkey eyes treated with relatively high doses of latanoprost, a prostaglandin (PG) F2α analog, or with any of the naturally occurring PGs.9 Arachidonic acid products that are known to stimulate growth and possibly melanogenesis in normal human epidermal melanocytes in tissue culture include PGE2, PGD2, leukotriene B4, leukotriene C4, leukotriene E4, thromboxane B2, and 12-hydroxyeicosatetraenoic acid.10 On the other hand, PGE1, PGF, and 6-keto-PGF do not exhibit a stimulatory effect.10 Iris color is determined by the amount of melanin (melanosomes) within iris stromal melanocytes, not by the number of melanocytes.1113 A uniformly blue iris has as many stromal melanocytes as a dark brown iris. Prolonged PG treatment has been shown to increase the eumelanin content of the melanocytes, but not the number of melanocytes in monkey irides.9

Neurohumoral input, including sympathetic innervation, is known to influence iris color.1416 Sympathetic innervation plays a role in the development of normal pigmentation of the iris at birth.17 A hallmark of unilateral congenital Horner syndrome is iris heterochromia, with a lighter-colored iris on the side of the sympathetic deficiency. Less well known is the observation that acquired Horner syndrome also can produce a lightening of iris color,16,18 suggesting that sympathetic innervation may be required to maintain normal iridial pigmentation, even in adults. Unilateral sympathectomy in rabbits, regardless of age, is known to produce heterochromia, with a lighter-colored iris on the sympathectomy side.1416 Also, topical application of an α-adrenergic blocker results in iris heterochromia in young rabbits.19

An interaction between PGs, adrenergic drugs, and the sympathetic nervous system has been reported previously. Cyclooxygenase inhibitors suppress the ocular hypotensive effects of adrenergic agents20 and suppress the reduction in intraocular pressure normally seen following acute superior cervical ganglionectomy.21 In toxicology studies, none of the hundreds of pigmented rabbits with intact sympathetic innervation of the iris that were treated with high doses of latanoprost for more than a year demonstrated darkening of iris color (Johan Stjernschantz, MD, PhD, Pharmacia & Upjohn, Kalamazoo, Mich, unpublished data, 1994). This study examines the relationship of sympathetic denervation and latanoprost treatment to the development of iris hyperpigmentation in a rabbit model.

MATERIALS AND METHODS

Twelve Dutch-belted female rabbits underwent unilateral superior cervical ganglionectomy (SCGx) at age 1 to 3 months (group 1). They were anesthetized with an injection of ketamine hydrochloride (20 mg/kg), xylazine hydrochloride (8 mg/kg), and acepromazine maleate (1 mg/kg), and SCGx as performed with the aid of a dissecting microscope. The occurrence of miosis postoperatively was initial evidence that sympathetic denervation had been achieved. Six months after surgery, the adequacy of the SCGx was tested pharmacologically with topical 10% cocaine hydrochloride; 1% hydroxyamphetamine hydrobromide; and 0.25% phenylephrine hydrochloride.

Color photographs of the iris of each eye were taken at 1- to 2-month intervals for 10 months. Four or 5 photographs of each eye were taken in a darkened room using a fundus camera. Flash intensity, magnification, F-stop, exposure time, and film type were consistent for all photographs. Slides of the right and left eyes of each rabbit were projected side-by-side in a darkened room. Four to 6 masked observers were asked to choose the darker of the 2 irides by indicating ++ for very certain, + for certain, and − for undecided. For each rabbit, 4 or 5 pairs of slides were judged at each time interval. The ratings from each rabbit at all time intervals were pooled and the degree of certainty of heterochromia between contralateral eyes of rabbits among the observers was rated as follows: strong (very certain was the unanimous opinion of all observers); moderate (1 or 2 observers might have disagreed with the others, or there might have been a slight inconsistency at some intervals during the course of treatment); mild (although there was a trend toward heterochromia, considerable variability existed); or no consistently distinguishable difference between eyes.

A separate group of 12 rabbits underwent bilateral SCGx at age 1 month (group 2). One rabbit died on the day following surgery. Four days after surgery, 1 drop (about 20 µL) of 0.005% latanoprost, was topically applied twice daily on weekdays and once daily on weekends to one randomly assigned eye and its vehicle to the contralateral eye in a masked fashion for 6 to 9 months in the 11 remaining rabbits. Photographs were taken and evaluated as described above. Treatment codes were broken only after a final assessment of heterochromia had been established at the end of the treatment period. All animal experimental procedures were approved by the university's Institutional Animal Care and Use Committee.

RESULTS

At 8 to 10 months after unilateral SCGx, 11 of the 12 rabbits in group 1 showed definite heterochromia, with the lighter-colored iris on the SCGx side, as determined unanimously by the masked observers (Figure 1). The heterochromia was observed as soon as 2 weeks after SCGx in the youngest rabbits.

Of the 11 rabbits in group 2 that underwent bilateral SCGx and unilateral latanoprost treatment, 10 showed some degree of heterochromia after 6 months of treatment: 5 rabbits, strong degree of certainty; 3 rabbits, moderate; and 2 rabbits, mild (Table 1). The darker eye was found to be the latanoprost-treated eye in 9 of the 10 rabbits (Figure 1). The only rabbit that received latanoprost in the eye thought to be lighter in color received a heterochromia rating with a mild degree of certainty. The 1 rabbit that did not show heterochromia was the only 1 of the 11 with an incomplete SCGx, as demonstrated by pupillary testing with cocaine hydrochloride, hydroxyamphetamine hydrobromide, and phenylephrine hydrochloride.

COMMENT

The heterochromia produced after unilateral SCGx in these pigmented rabbits (group 1) confirmed the results of previous studies1416,22 and is consistent with the heterochromia that is observed clinically in patients with Horner syndrome, especially when it occurs congenitally. Iris color undergoes changes during aging17,23 and appears to be influenced by neurohumoral factors. The lightening of iris color in some patients who develop acquired Horner syndrome as adults suggests that sympathetic tone contributes to maintenance of iris color in adulthood.16,18

The interaction between PGs, the sympathetic nervous system, and adrenergic agonists has been previously delineated. In clinical and/or animal studies, cyclooxygenase inhibitors prevented the ocular hypotensive effects of adrenergic agents (reviewed by Camras and Podos,20 1989) and inhibited the intraocular pressure decrease after acute superior cervical ganglionectomy.21 With intact sympathetic innervation of the iris, high doses of latanoprost for more than a year do not result in darkening of iris color in pigmented rabbits (Johan Stjernschantz, MD, PhD, Pharmacia & Upjohn, Kalamazoo, unpublished data, 1994). However, the results of the present study suggest that PGs may act in the absence of sympathetic tone by stimulating melanogenesis to darken the color of irides that are lighter following sympathectomy.

Iris stromal melanocytes in the iris periphery are preferentially innervated by the sympathetic nerve endings, whereas those around the sphincter of the iris tend to be cholinergically innervated.16 Iris freckles/nevi are not sympathetically innervated and therefore do not fade in Horner syndrome.24 If PGs were substituting for deficient sympathetic innervation, the peripheral rather than more central iris stroma would be expected to darken, which is consistent with the clinical observations.16 Furthermore, latanoprost-induced darkening of nevi/freckles would not be expected and in fact does not occur. Among the hundreds of eyes treated with latanoprost that have been carefully evaluated photographically in clinical studies, none of the iris freckles/nevi have demonstrated any change. Even large iris nevi observed at baseline do not change during the course of 1 year of treatment with latanoprost.16

Darkening of the iris after topical latanoprost treatment does not appear to be due to proliferation of melanocytes.9 In our experimental rabbit model, histopathologic studies are being performed to determine whether the iris color darkening is due to proliferation of melanocytes or an increase in melanin synthesis within the melanocytes. The model of sympathetically denervated rabbit eyes may be useful to further study the mechanism of PG-induced iris color change.

Back to top
Article Information

Accepted for publication April 16, 1998.

This study was supported in part by a grant from Pharmacia & Upjohn, Kalamazoo, Mich; by a challenge grant from Research to Prevent Blindness Inc, New York, NY; and by the Gifford Laboratory funds, University of Nebraska Medical Center, Omaha.

Presented in part at the 72nd annual meeting of the Association for Research in Vision and Ophthalmology, Fort Lauderdale, Fla, May 17, 1995. This article was submitted in part by Dr Camras as a small portion of a thesis to the American Ophthalmologic Society, Durham, NC.

Reprints: Carl B. Camras, MD, Department of Ophthalmology, University of Nebraska Medical Center, 600 S 42nd St, Omaha, NE, 68198-5540 (Dr Camras).

References
1.
Camras  CB Prostaglandins and the treatment of the glaucomas. Ritch  RShields  MBKrupin  Teds.The Glaucomas. St Louis, Mo Mosby–Year Book Inc1996;1449- 1461
2.
Camras  CBThe US Latanoprost Study Group, Comparison of latanoprost and timolol in patients with ocular hypertension and glaucoma: a 6-month, masked, multicenter trial in the United States. Ophthalmology. 1996;103138- 147Article
3.
Camras  CBAlm  AWatson  PGStjernschantz  JThe Latanoprost Study Groups, Latanoprost, a prostaglandin analog, for glaucoma therapy: efficacy and safety after 1 year of treatment in 198 patients. Ophthalmology. 1996;1031916- 1924Article
4.
Alm  AStjernschantz  JThe Scandinavian Latanoprost Study Group, Effects on intraocular pressure and side-effects of 0.005% latanoprost applied once daily, evening or morning: a comparison with timolol. Ophthalmology. 1995;1021743- 1752Article
5.
Watson  PGStjernschantz  JThe Latanoprost Study Group, A 6-month, randomized, double-masked study comparing latanoprost with timolol in open-angle glaucoma and ocular hypertension. Ophthalmology. 1996;103126- 137Article
6.
Wistrand  PStjernschantz  JOlsson  K The incidence and time-course of latanoprost-induced iridial pigmentation as a function of eye color. Surv Ophthalmol. 1997;41(suppl 2)S129- S138Article
7.
Johnstone  MA Hypertrichosis and increased pigmentation of eyelashes and adjacent hair in the region of the ipsilateral eyelids of patients treated with unilateral topical latanoprost. Am J Ophthalmol. 1997;124544- 547
8.
Wand  M Latanoprost and hyperpigmentation of eyelashes. Arch Ophthalmol. 1997;1151206- 1208Article
9.
Selén  GStjernschantz  JResul  B Prostaglandin-induced iridial pigmentation in primates. Surv Ophthalmol. 1997;41(suppl 2)S125- S128Article
10.
Tomita  YMaeda  KTagami  H Melanocyte-stimulating properties of arachidonic acid metabolites: possible role in postinflammatory pigmentation. Pigment Cell Res. 1992;5357- 361Article
11.
Eagle  RC  Jr Iris pigmentation and pigmented lesions: an ultrastructural study. Trans Am Ophthalmol Soc. 1988;88581- 687
12.
Wilkerson  CLSyed  NAFisher  MRRobinson  NLWallow  IHLAlbert  DM Melanocytes and iris color: light microscopic findings. Arch Ophthalmol. 1996;114437- 442Article
13.
Imesch  PDBindley  CDKhademian  Z  et al.  Melanocytes and iris color: electron microscopic findings. Arch Ophthalmol. 1996;114443- 447Article
14.
Calhoun  FP Causes of heterochromia iridis with special reference to paralysis of the cervical sympathetic. Am J Ophthalmol. 1919;2255- 269
15.
Calhoun  FP A consideration of the causes of heterochromia iridis, with special reference to a paralysis of the cervical sympathetic. Trans Am Ophthalmol Soc. 1919;16277- 312
16.
Laties  AM Ocular melanin and the adrenergic innervation to the eye. Trans Am Ophthalmol Soc. 1974;72560- 605
17.
Bito  LZ Prostaglandins: a new approach to glaucoma management with a new, intriguing side effect. Surv Ophthalmol. 1997;41(suppl 2)S1- S14Article
18.
Diesenhouse  MCPalay  DANewman  NJTo  KAlbert  DM Acquired heterochromia with Horner syndrome in 2 adults. Ophthalmology. 1992;991815- 1817Article
19.
Odin  LO'Donnell  FE  Jr Adrenergic influence on iris stromal pigmentation: evidence for alpha-adrenergic receptors. Invest Ophthalmol Vis Sci. 1982;23528- 530
20.
Camras  CBPodos  SM The role of endogenous prostaglandin in clinically-used and investigational glaucoma therapy. Bito  LZStjernschantz  Jeds.The Ocular Effects of Prostaglandins and Other Eicosanoids. New York, NY Alan R Liss Inc1989;312459- 475
21.
Camras  CBZhan  G-LOhia  SOhia  EWang  Y-L The role of prostaglandins and neuropeptide Y in the reduction of intraocular pressure 1 day after sympathectomy in rabbits. Invest Ophthalmol Vis Sci. 1993;34(suppl)932
22.
Laties  AMLerner  AB Iris colour and relationship of tyrosinase activity to adrenergic innervation. Nature. 1975;255152- 153Article
23.
Bito  LZMatheny  APCruickshanks  KJNondahl  DMCarino  OB Eye color changes past childhood: the Louisville Twin Study. Arch Ophthalmol. 1997;115659- 663Article
24.
Dryja  TPAlbert  DM Lack of adrenergic influence on the pigmentation of iris nevus cells. Arch Ophthalmol. 1980;981996Article
×