Olli Oksala

Nitric oxide and cyclic GMP enhance aqueous humor outflow facility in rabbits

Purpose. Nitric oxide (NO) may control intraocular pressure (IOP)-regulating mechanisms physiologically and in ocular diseases such as glaucoma. The aim of the present study was to clarify whether an increase in aqueous humor outflow facility could explain the IOP-lowering effect of the NO/cyclic GMP pathway we recently described. Methods. Test compounds were administered to anesthetized rabbits (New Zealand White, n = 6) intracamerally (5µl) in the following doses: nitrosocaptopril 12.3µg, captopril 10.9µg, sodium nitroprusside (SNP) 13.1µg and 8-Brc-GMP 22.3µg. Outflow facility (C) was determined by the two-level constant pressure infusion method. Outflow facility, C 1 and C 2, was measured at lower and higher pressure levels, respectively. Results. Outflow facility was increased after treatment with SNP (increase in C in the experimental eye as compared to the control eye C 1 80% and C 2 74%), nitrosocaptopril (C 1 69% and C 2 64%) and 8-Br-cGMP (C 1 35% and C 2 33%). Captopril had no effect on outflow facility (C 1 -12% and C 2 2%). Blood pressure was not affected by the drugs. Conclusions. We conclude that enhancement of outflow facility by nitrosocaptopril, SNP and 8-Br-cGMP, their second messenger derivative, at least partly explains the IOP-lowering effect of NO releasing compounds.

Activities of Angiotensin-Converting Enzymes ACE1 and ACE2 and Inhibition by Bioactive Peptides in Porcine Ocular Tissues

PURPOSE An active local renin-angiotensin system (RAS) has recently been found in the human eye. The aim of the present study was to compare the activities of central RAS enzymes (ACE1 and 2) in porcine ocular tissues, morphologically and physiologically close to the human eye. In addition, the effects of three ACE-inhibitory tripeptides on these enzymes were evaluated. METHODS Enucleated fresh porcine eyes were used. Activities of ACE1 and ACE2 and their inhibition by bioactive tripeptides (Ile-Pro-Pro, Val-Pro-Pro, Leu-Pro-Pro) as well as by a standard ACE-inhibitor captopril were assayed in the vitreous body, the retina and the ciliary body using fluorometric detection methods. RESULTS Activity of ACE1 as well as ACE2 was found in all tissues evaluated. ACE1 activity was markedly higher in the ciliary body (3.7 +/- 0.7 mU/mg protein) than in retina (0.2 +/- 0.02 mU/mg), whereas ACE2 activities in the ciliary body (0.2 +/- 0.02 mU/mg) and retina (0.2 +/- 0.01 mU/mg) were at the same level. In the vitreous body ACE1 activity (8.2 +/- 0.31 nmol/min/mL) was manifold compared to that of ACE2 (0.1 +/- 0.02 nmol/min/mL). The tripeptides inhibited ACE1 at one-thousandth of the concentration needed to inhibit ACE2. All peptides studied evinced about equal inhibitory activities. CONCLUSION To our knowledge the present findings constitute the first evidence of ACE2 activity in the ciliary and vitreous bodies, in addition to previously described activity in the retina. The known favorable effects of ACE2 products vs. those of ACE1 suggest a counterbalancing interaction of these two enzyme homologues in physiological regulation of ocular circulation and pressure and possible protective role in certain ophthalmic disorders such as glaucoma and diabetic retinopathy.

Angiotensin (1-7) Reduces Intraocular Pressure in the Normotensive Rabbit Eye

PURPOSE In the present study the effects of exogenous angiotensin II and its breakdown metabolite angiotensin (1-7) on the intraocular pressure (IOP) and on aqueous humor dynamics in normotensive rabbit eye were evaluated. METHODS Male New Zealand White rabbits with normal IOP were used for intravitreous and topical administration of the test compounds. IOP was measured in conscious rabbits by pneumatonometer after topical anesthesia. Outflow measurements were made with a two-level constant pressure method in anesthetized animals. RESULTS Angiotensin (1-7) administered intravitreously reduced IOP within 1 to 5 hours (P < 0.05). This effect was abolished by the selective angiotensin (1-7) antagonist A-779, and partially by the selective angiotensin II type 2 receptor antagonist PD123319. When olmesartan, an angiotensin II type 1 receptor blocker, was administered simultaneously with angiotensin (1-7), no antagonism was seen. Intravitreous administration of CGP42112 A, an angiotensin II type 2 receptor agonist, and angiotensin II did not significantly influence IOP, nor did topical administration of these compounds alter IOP. Angiotensin II significantly reduced outflow facility (P < 0.01) dose dependently, whereas angiotensin (1-7) had no effect. CONCLUSIONS Angiotensin (1-7) is a biologically active vasodilatory and antiproliferative heptapeptide, and its vascular effects counteract those of angiotensin II. It reduces intraocular pressure possibly by a selective Mas receptor, without changing aqueous humor outflow facility in the normotensive rabbit eye.

Does the renin-angiotensin system also regulate intra-ocular pressure?

The renin-angiotensin-aldosterone system is known to play an essential role in controlling sodium balance and body fluid volumes, and thus blood pressure. In addition to the circulating system which regulates urgent cardiovascular responses, a tissue-localized renin-angiotensin system (RAS) regulates long-term changes in various organs. Many recognized RAS components have also been identified in the human eye. The highly vasoconstrictive angiotensin II (Ang II) is considered the key peptide in the circulatory RAS. However, the ultimate effect of RAS activation at tissue level is more complex, being based not only on the biological activity of Ang II but also on the activities of other products of angiotensinogen metabolism, often exerting opposite effects to Ang II action. In recent studies, orally administered angiotensin II type 1 receptor blockers and angiotensin-converting enzyme inhibitors lower intra-ocular pressure (IOP), likewise topical application of these compounds, the effect being more prominent in ocular hypertensive eyes. Based on previous findings and our own experimental data, it can strongly be suggested that the RAS not only regulates blood pressure but is also involved in the regulation of IOP.

Is There a Relationship Between Blood Pressure and Intraocular Pressure? An Experimental Study in Hypertensive Rats

Purpose: Evaluation of relation between blood pressure (BP) and intraocular pressure (IOP) in two hypertensive rat strains: spontaneously hypertensive rats (SHR) and double transgenic (dTGR) (harboring human renin and angiotensinogen genes) rats, and in their normotensive control Wistar Kyoto and Sprague Dawley rats, respectively. Methods: Each rat strain was divided into medicated and non-medicated groups. Medicated rats were treated orally with an angiotensin II receptor type 1 blocker. IOP was measured using a specific rebound tonometer and BP by a tail-cuff method. Both parameters were determined in conscious animals every second week. For comparison, at the end of the study, IOP was measured in conscious and anesthetized rats. Results: The baseline IOP was higher in hypertensive rats vs their normotensive controls. Eight weeks of treatment with an angiotensin type 1 receptor blocker did not prevent a slight increase in IOP, although it abolished the development of hypertension in SHR. The markedly elevated IOP was reduced in medicated and non-medicated dTGR animals during the short follow-up period. General anesthesia reduced IOP significantly. Conclusion: The results suggest a positive relation between BP and IOP in hypertensive rats.

Localization of calcitonin gene-related peptide binding sites in the eye of different species

The localization of calcitonin gene-related peptide (CGRP) binding sites in the eye of monkey, pig, cat and guinea pig was studied by autoradiography. Specific binding of CGRP was found in ciliary muscle, ciliary processes and limbal conjunctiva in all tested species. Furthermore, specific binding sites of CGRP was found in the choroidea of monkey, pig and guinea pig, in the iris of pig, cat and guinea pig, in the retina of pig and in the anterior chamber angle of cat. The number of specific binding sites varied depending on the tissue and species. The present study shows that there are specific binding sites of CGRP in the eye of monkey, pig, cat and guinea pig. CGRP binding sites found in vascular system of ciliary body, choroidea and iris further demonstrates the role of CGRP as a vasoregulatory peptide. Binding sites in the ciliary muscle, in the limbal conjunctiva and in the chamber angle area may indicate a role in the regulation of ciliary muscle tone, epithelial cell regeneration and aqueous humour outflow.

Angiotensin receptors in the eyes of arterial hypertensive rats

Purpose:  The aim of the present study was to determine whether the eye tissues of arterial hypertensive rats evince expression of angiotensin receptors (AT1 and AT2) as well as the novel Mas receptor, whose endogenous ligand is vasorelaxing Angiotensin (1–7) [Ang (1–7)].

Expression of Cytochrome P450 (CYP) Enzymes in Human Nonpigmented Ciliary Epithelial Cells: Induction of CYP1B1 Expression by TCDD

PURPOSE Cytochrome P450 (CYP) enzymes metabolize endogenous compounds such as steroid hormones, fatty acids, and xenobiotics, including drugs and carcinogens. Expression of CYP enzymes in ocular tissues is poorly known. However, mutations in the CYP1B1 gene have been linked to congenital glaucoma. The aim of the present study was to investigate the expression and regulation of cytochrome P450 enzymes in a human nonpigmented ciliary epithelial cell line. METHODS Expression of mRNAs for major xenobiotic metabolizing CYPs in families 1-3 and regulatory factors involved in the induction of CYPs was studied using reverse transcriptase-polymerase chain reaction. For induction studies, the cells were treated with dexamethasone or 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) for 24 hours. RNA and immunoblotting analysis were used to study CYP induction. Transcriptional regulation of CYP1B1 gene was studied by transient transfection of reporter gene constructs. RESULTS mRNAs of CYP1A1, CYP1B1, and CYP2D6 and of the regulatory factors aryl hydrocarbon receptor (AHR), aryl hydrocarbon receptor nuclear translocator, and glucocorticoid receptor were expressed in the human nonpigmented ciliary epithelial cell line. CYP1B1 mRNA was strongly and dose dependently induced by TCDD. CYP1B1 protein was detected only after TCDD treatment of the human nonpigmented ciliary epithelial cells. CYP1B1 promoter was activated by TCDD. The major drug-metabolizing enzymes CYP1A2, CYP2Cs, and CYP3As were not detected in these cells, and dexamethasone treatment had no effect on CYP expression. CONCLUSIONS TCDD potently induces CYP1B1 mRNA in human nonpigmented ciliary epithelial cells, suggesting the involvement of an AHR-mediated pathway in the regulation of ciliary CYP1B1 expression.

Ocular irritative response to YAG laser capsulotomy in rabbits: Release of calcitonin gene-related peptide and effects of methysergide

The Neodymium (Nd):YAG laser is commonly used in ophthalmology mainly for the posterior capsulotomy in patients with secondary cataract after extracapsular cataract extraction. A frequent side-effect following different kinds of YAG laser treatments is an acute increase in the intraocular pressure (IOP). The present study addresses the role of calcitonin gene-related peptide (CGRP) in the ocular irritative response following YAG laser anterior capsulotomy in rabbits. The YAG laser anterior capsulotomy caused an irritative response in the eye, which consisted of an increase in the IOP, miosis and breakdown of the blood-aqueous barrier. Following YAG laser capsulotomy, CGRP-immunoreactivity was found in the aqueous humour in different molecular weight forms as revealed by gel-permeation chromatography. One of the peaks coeluted with synthetic human CGRP. Methysergide attenuated the increase in the IOP and disruption of the blood-aqueous barrier, but not the miosis, following YAG laser anterior capsulotomy. The present study demonstrates the release of CGRP into the aqueous humour following YAG laser capsulotomy, and suggests that CGRP is partly causing the increase in IOP and disruption of the blood-aqueous barrier in this irritative response.

Effect of Simultaneous CGRP and PGF2α on the Outflow Facility in the Rabbit Eye

Both calcitonin gene-related peptide and prostaglandins have an influence on the intraocular pressure in the eye. In this study, the effects of low intracameral doses (20 ng and 50 ng) of calcitonin gene-related peptide, prostaglandin F2α, and simultaneous dosing of both, on the outflow facility were studied in the rabbit. Calcitonin gene-related peptide increased the outflow facility at both 20 and 50 ng doses, while prostaglandin F2α increased it only at a 50 ng dose. Further, simultaneous administration of both at 50 ng doses increased the outflow facility and showed a slight additive effect with these two compounds. The results further indicate that these compounds have a different mechanism of action. Calcitonin gene-related peptide seems to increase the conventional outflow (trabecular outflow), while prostaglandin F2α tends to increase the unconventional outflow (uveoscleral outflow).