Desensitization of the isolated bovine iris sphincter muscles by pilocarpine

We investigated the effect of pilocarpine on the bovine iris sphincter. This muscle contracted in a dose-dependent manner from 2 × 10^–6 M, with a maximal response at 3 × 10^–3 M. The ED₅₀ values was (1.6 ± 0.3) × 10^–4M. Pilocarpine generated a desensitization while desensitization was not significant in the case of other cholinergic agents such as acetylcholine (Ach) and carbachol. Desensitization was profoundly increased in the presence of Ach, neostigmine or eserine: the responses to the second and third trials of pilocarpine were reduced to approximately 8–10% or 30% of the corresponding first responses. Pilocarpine reportedly releases transmitters and alters choline-acetyltransferase activity.These results taken together suggest that either variable Ach synthesis, inhibitory transmitter release or possible toxic action in high concentrations may be involved in the pilocarpine-induced responses of the bovine iris sphincter muscle. The desensitization and parital agonist-antagonist action of pilocarpine could not explain the characteristics of the bovine iris sphincter muscle.     

Effect of somatostatin and galanin on isolated rabbit iris sphincter and dilator muscles

The neuropeptides somatostatin and galanin are present in the iris and may modulate pupil diameter. We examined the effects of somatostatin and galanin on isolated rabbit iris dilator and sphincter smooth muscles that were mounted in an organ bath. An isometric transducer recorded changes in tension in response to electric field stimulation (100 Hz, 0.3 m sec in duration, 10 V in strength) delivered by a pair of platinum plate electrodes. The dilator muscle response to field stimulation was not changed by either peptide, even at the highest concentrations examined. The sphincter muscle response consisted of two components: a fast component mediated by acetylcholine and slow component mediated by substance P. Both somatostatin and galanin attenuated the cholinergic component in a dose-dependent manner (from 0.3 nm to 0.1 μm) but had no effect on responses mediated by substance P. Galanin was more effective (attenuation of 43% at 0.1 μm) compared with somatostatin (attenuation of 16% at 0.1 μm) in reducing the cholinergic response. Neither peptide affected the contraction induced by acetylcholine (1 mm). Therefore both peptides inhibited cholinergic transmission in the sphincter muscle, although the degree of inhibition by each was different. We conclude that somatostatin and/or galanin may induce mydriasis by attenuating cholinergic neurotransmitter release.     

Effects of acetylcholine and carbachol on the iris sphincter

We compared the effects of acetylcholine and carbachol on iris sphincters. The muscle strips (mostly bovine tissues) were isolated and incubated in a 0.2ml organ bath. Results revealed an approximately ten-thousandfold difference in potency between carbachol and acetylcholine, in the bovine iris. Acetylcholine and/or endogenous chemical agents were spontaneously released from bovine tissue. Acetylcholinesterase activity in this tissue strongly inhibited the acetylcholine action, in order to protect the nerve terminals from random depolarization due to the released acetylcholine. Among several species (bovine, rabbit, hamster), Ach action was the least in the bovine iris sphincter muscle.     

Response of bovine intra-ocular muscles to transmural stimulation in the presence of various prostaglandins

Effects of prostaglandins (PGs) E1, E2 and F2 alpha on the electrically stimulated and non-stimulated smooth muscles of bovine intra-ocular tissues were investigated in vitro. PGs E1, E2 and F2 alpha contracted the sphincter and these contractions were not antagonized by either cholinergic or adrenergic blocking agents. PGF2 alpha was much weaker than PGE1 or PGE2. PGs in high concentrations produced an irreversible contracture of the sphincter muscle. In contrast, the dilator muscle contracted with PGF2 alpha, but not PGE1 or PGE2. The ciliary muscle did not respond to these PGs or to indomethacin. Electrical field stimulation with short pulses produced excitation of the intrinsic nerves. The responses produced were not altered in amplitude by the application of any dose of PGs or indomethacin, although indomethacin did lower the tone of the sphincter. PGs did not influence the amplitude of the acetylcholine-induced responses of the intra-ocular muscles. These results indicate that in each of the bovine intra-ocular muscles, PGs play a minor role in neuromuscular transmission both prejunctionally and postjunctionally. Thus, PG-induced contractions of the bovine intra-ocular muscles are considered to occur by a direct action on the muscle rather than on the nerves.     

Action of biologically active peptides on monkey iris sphincter and dilator muscles

Biologically active peptides modulate pupillary responsiveness in many non-primate mammals. We examined the action of seven different peptides on iris sphincter and dilator muscles of rhesus monkey. Iris sphincter and dilator muscle preparations from rhesus monkeys were mounted in an organ bath, and tension changes were recorded by an isometric transducer. Electrical field stimulation (100Hz, 0.3 msec, 10V) was applied through a pair of platinum plate electrodes. Monkey iris sphincter and dilator muscles produced simple cholinergic and adrenergic excitatory responses respectively to electrical field stimulation. Strong field stimulation did not elicit slow Substance P (SP) mediated contractions like those in rabbit iris sphincter. Exogenously applied pituitary adenylate cyclase-activating peptide (PACAP) enhanced in a concentration-dependent manner (0.3 nM-0.1 microm) the sphincter response to field stimulation, while neuropeptide Y (NPY) and somatostatin (SRIF) attenuated it. These three peptides did not affect sphincter contractions induced by acetylcholine, and therefore were acting at presynaptically. SP, calcitonin gene-related peptide (CGRP), vasoactive intestinal polypeptide (VIP) and galanin (GAL) had no effect (at 0.1 microm) on iris sphincter. None of seven exogenously applied peptides had an effect on monkey iris dilator muscle. The innervation of primate irises may be relatively simple compared to non-primates because each of the peptides in this study can modulate miosis or mydriasis in non-primate mammals. Future studies will be expected on the functional significance of species differences in iridial innervation.     

Growth of the sphincter muscle and neuroepithelia of the iris and ciliary body in rat: Autoradiographic study

Cell proliferation in the sphincter muscle and neuroepithelia of the iris and ciliary body in pigmented 1–11-day old rats was studied using pulse and continuous labelling with [³H]-thymidine. The duration of the M phase was calculated by the colchicine method. Labelling indices were the highest between the third and fifth days and did not exceed 28·3% in the ciliary body, 10·8% in the iris and 14·1% in the sphincter muscle. This level of labelled nuclei was observed in the sphincter muscle up to the 8th day while in the neuroepithelia labelling indices start to decrease after the fifth day. The cell cycle time (T) and population doubling time (T₂) in the 5- and 8-day old rats were calculated.Three characters of the iris and ciliary body cells proliferation call for attention: (1) cell populations with a longer cell cycle (neuroepithelia of the ciliary body) multiply more intensively than those with a shorter cell cycle (neuroepithelia of the iris) due to the larger number of cells involved in proliferation, (2) cell populations multiplying with an equal intensity (both neuroepithelia of the iris, sphincter muscle) have different structures of the cell cycles mainly owing to the variation of G₁ phase, (3) the M phase is considerably prolonged by comparison with that of the neural retina. These peculiarities of proliferation were considered as a way of cell adaptation to the development of functional entity of the anterior part of the eye during the period of the most intensive growth of its components.     

Effects of prostaglandin F2alpha, latanoprost and carbachol on phosphoinositide turnover, MAP kinases, myosin light chain phosphorylation and contraction and functional existence and expression of FP receptors in bovine iris sphincter

A potential role for myosin light chain kinase (MLCK) in regulating intraocular pressure and outflow function has recently been reported in living monkey eye and rabbit eye. There is little information about the effects of the ocular hypotensive agents, prostaglandin F2alpha (PGF2alpha) and latanoprost on this signaling pathway in ocular tissues. The aim of this study was to determine the agonist activity of PGF2alpha, latanoprost and carbachol (CCh) on the MLCK pathway in isolated bovine iris sphincter and furthermore to investigate the existence of the FP receptor in this tissue. In the present studies on the MLCK pathway four signal transduction mechanism assays were employed, phosphoinositide (PI) turnover, p42/p44 MAP kinase phosphorylation and activation, MLC phosphorylation and contraction. In the studies on the existence of the FP receptor in the bovine iris sphincter, the pharmacology and expression of the FP receptor protein, using a polyclonal anti-FP-receptor antibody and Western blot analysis, were determined. The data obtained on the MLCK pathway showed that the three agonists stimulated the biochemical and pharmacological responses in a concentration and time-dependent manner and that the order of potency and efficacy is PGF2alpha>latanoprost>CCh. The EC50 values in the PI turnover, MAP kinase phosphorylation, MLC phosphorylation and contraction assays were for PGF2alpha: 9, 42, 200 and 140 nM, respectively, for latanoprost: 13, 59, 250 and 828 nM, respectively, and for CCh: 22, 200, 630 and 910 nM, respectively. Wortmannin, a selective inhibitor of MLCK, dose-dependently inhibited MLC phosphorylation and contraction induced by PGF2alpha, demonstrating a close relationship between activation of the MLCK pathway and contraction. The pharmacological studies showed that in the concentration range of 1 nM to 10 microM, the FP-receptor agonists caused concentration-response curves with the following order of potencies: 17-phenyl trinor PGF2alpha (bimatoprost acid)>PGF2alpha>cloprostenol>latanoprost>latanoprost acid>bimatoprost amide>fluprostenol. Immunoblot analysis of the FP receptor demonstrated expression of the prostaglandin FP receptor protein in this smooth muscle. These results clearly indicate that the MLCK signaling pathway is involved in the FP-receptor function of the bovine iris sphincter and furthermore demonstrate that functional FP receptors exist and are expressed in this tissue.     

Ghrelin as a novel locally produced relaxing peptide of the iris sphincter and dilator muscles

Ghrelin is a recently described acylated peptide, which works as a somatosecretagogue and has described effects on the smooth, skeletal and cardiac muscle. We examined the production and effects of ghrelin on relaxation of the iris muscles. Contractile effects of 1-5 human ghrelin (frGhr, 10(-9)-6 x 10(-5)M) and 1-5 human des-octanoyl-ghrelin (d-frGhr; 10(-9)-6 x 10(-5)M) were tested on iris rabbit sphincter (n=11 frGhr; n=7 d-frGhr), dilator (n=6 frGhr; n=6 d-frGhr) and rat sphincter (n=6 frGhr; n=8 d-frGhr) precontracted muscles. On rabbit sphincter the effect of frGhr was also tested in presence of: i) L-NA (10(-5)M; n=7); ii) indomethacin (10(-5)M; n=7); iii) DLys(3)GHRP6 (10(-4)M; n=6); and iv) apamin+carybdotoxin (10(-6)M; n=6). Furthermore, on rabbit dilator the effect of frGhr was tested in presence of DLys(3)GHRP6 (10(-4)M; n=7). Finally, ghrelin mRNA production was assessed by "in situ" hybridization in Wistar rat eyes (n=8). In all muscles, frGhr promoted a concentration-dependent relaxation, maximal at 6 x 10(-5)M, 1.5-3 min after its addition, decreasing tension by 34.1+/-12.1%, 25.8+/-4.8% and 52.1+/-10.3% in the rabbit sphincter, dilator and rat sphincter, respectively. In the rabbit sphincter the relaxing effects of frGhr were: (i) enhanced in presence of DLys(3)GHRP6 (118.1+/-21.1%); (ii) blunted by indomethacin; and (iii) not altered by apamin+carybdotoxin (36.4+/-14.4%) or L-NA (52.4+/-11.4%). Relaxing effects of d-frGhr in rabbit (43.3+/-5.2%) and rat (77.1+/-15.3%) sphincter muscles were similar to those of frGhr. In rabbit dilator muscle, d-frGhr did not significantly alter active tension and the relaxing effect of frGhr was blunted by GHSR-1a blockage. Ghrelin mRNA was identified in iris posterior epithelium. In conclusion, ghrelin is a novel, locally produced, relaxing agent of iris dilator and sphincter muscles, an effect that is mediated by GHSR-1a in the former, but not in the latter. Furthermore, in the sphincter it seems to be mediated by prostaglandins, but not by NO or K(Ca) channels.     

The response of the isolated iris sphincter muscle of various mammalian species to substance P

Contractile responses of the isolated iris sphincter muscle from the rabbit, cow, pig, cat, dog, baboon and man to substance P were compared under isotonic conditions using carbachol as a reference standard. Iris preparations from the rabbit, pig and cow showed strong and consistent responses to substance P whilst those from the cat, dog, baboon and human eyes did not contract.     

The different contributions of prostaglandins (E1, E2, F2alpha,D2) to the tone and neurogenic response of bovine iris sphincter muscle

The function and sites of action of prostaglandins (PGs) vary in different animal species and tissues. In this study the influence of PGs (E₁, E₂, F_2alpha, D₂) on muscle tone and nerve-mediated contraction was investigated in isolated bovine iris sphincter muscles.None of these PGs exogenously applied influenced the neuromuscular transmission. By contrast, after treatment with indomethacin, all PGs tested contracted the muscle much more than in the absence of indomethacin and under these conditions the PGs potentiated responses to cholinergic nerve stimulation. Their ED₅₀ were (2.2 ± 0.2) × ^–7 M for PGE₁, (6.7 ± 0.3) × ^–8M for PGE₂, and (7.3 ± 0.4) × ^–8 M for PGF_2alpha. PGE₁ acted both on nerves and the muscle cell. PGE₂ had its influence mostly via nerves. Whereas PGF_2alpha was less potent in the absence of indomethacin, PGF_2alpha had much more potent action primarily on nerves and partly on muscles after treatment with indomethacin. High concentrations of PGD₂ had both pre- and post-junctional action with accompanying weak contraction of the muscle. Thus the degrees of pre- and post-junctional involvement were different from one another.There is a possibility that the application of these PGs alone masked the role of such endogenous agents. In order to understand and clarify the site and action of PGs, pretreatment with indomethacin may be useful in the iris muscle. In conclusion, PGs modulate cholinergic activity in the bovine iris sphincter muscles, as well as regulate the muscle tone.     

The musculature and pupillary response of the Great Horned Owl iris

There is considerable confusion in the literature regarding the nature of the musculature of the avian iris. The most commonly held view is that both the sphincter and dilator are striated. The iris of the Great Horned Owl (Bubo virginianus) has a complex iridial musculature consisting of three circumferential components (a myoepithelium, smooth muscle and striated muscle) and two radial components (a well-developed myoepithelium and a few striated fibers). On the basis of the anatomy and relative development of these components, and a quantitative analysis of the pupillary reflex, it is proposed that the circumferential striated muscle is the primary pupillary constrictor and radial myoepithelium is the primary dilator. The annular band of smooth muscle may play an important role in maintaining pupillary size.     

Artisan虹膜夹持型人工晶状体植入术及后房型人工晶状体睫状沟缝线固定术治疗无晶状体眼的比较

目的:比较Artisan虹膜夹持型人工晶状体植入术及后房型人工晶状体睫状沟缝线固定术治疗无晶状体眼的疗效及并发症。方法:2007-03/2009-03我院住院患者中连续24例24眼无后囊膜支持的无晶状体眼患者,随机分为两组。一组11眼行Artisan虹膜夹持型人工晶状体植入术,另一组13眼选择后房型人工晶状体睫状沟缝线固定术。观察手术前及手术后1d;1wk;1mo的裸眼视力(visual acuity,VA)、最佳矫正视力(best corrected visual acuity,BCVA)、眼压(intraocularpressure,IOP)、角膜内皮细胞计数(corneal endothelial cells,CECs)。结果:两组间比较,术前VA,BCVA,CECs差异无统计学意义,术后BCVA,CECs差异无统计学意义。Artisan组手术后VA优于术前BCVA,差异有统计学意义。睫状沟缝线固定组手术后VA与手术前BCVA差异无统计学意义。两组手术前后IOP差异无统计学意义。结论:Artisan虹膜夹持型人工晶状体植入术与后房型人工晶状体睫状沟缝线固定术都是治疗无晶状体眼有效方法。两者比较,Artisan虹膜夹持型人工晶状体植入术手术操作相对简单,组织损伤小,更加安全,是治疗无后囊膜支持的无晶状体眼的比较理想的治疗方法。     

Effects of acetylcholine and norepinephrine on incorporation of [32P]orthophosphate into phospholipids of rabbit iridial processes and iris smooth muscle

We have investigated: (a) phospholipid composition, inclusive of higher inositides, of rabbit iridial processes and iris smooth muscles; (b) ³²P_i incorporation into their respective phospholipids, and (c) the effects of muscarinic cholinergic and adrenergic agonists and antagonists on ³²P labelling of phospholipids of the iridial processes and iris smooth muscle. (1) Phosphatidylcholine, phosphatidylethanolamine, their respective plasmalogens and sphingomyelin, were found to be the major phospholipids in the iridial processes and iris smooth muscle. They constituted about 85% of the total phospholipids of these ocular tissues. (2) Both iridial processes and iris smooth muscles rapidly incorporated ³²P_i and [1-¹⁴C]-arachidonic acid into their respective phospholipids, however this incorporation amounted to only 20% of that found for the whole iris-ciliary body. This could suggest a metabolic interrelationship between the iridial processes and the smooth muscle of the iris. (3) Addition of acetylcholine and norepinephrine to the iridial processes and iris smooth muscle increased ³²P labelling of phosphatidic acid and phosphatidylinositol of the tissues. The increase in phospholipid labelling was higher in the iridial processes as compared to the iris smooth muscle. The effect of acetylcholine was blocked by atropine and that of norepinephrine was blocked by phentolamine and prazosin but not by yohimbine. This suggests that the observed effects of these neurotransmitters on phospholipid phosphorylation in the iridial processes and iris smooth muscle are mediated through muscarinic cholinergic and α₁-adrenergic receptors, respectively. The data presented provide additional support for the concept that in the iris-ciliary body the neurotransmitter-induced ³²P labelling of phosphoinositides is probably linked to the functional activities of this tissue.     

Bimatoprost and prostaglandin F(2 alpha) selectively stimulate intracellular calcium signaling in different cat iris sphincter cells

Bimatoprost is a synthetic analog of prostaglandin F(2 alpha) ethanolamide (prostamide F(2 alpha)), and shares a pharmacological profile consistent with that of the prostamides. Like prostaglandin F(2 alpha) carboxylic acid, bimatoprost potently lowers intraocular pressure in dogs, primates and humans. In order to distinguish its mechanism of action from prostaglandin F(2 alpha), fluorescence confocal microscopy was used to examine the effects of bimatoprost, prostaglandin F(2 alpha) and 17-phenyl prostaglandin F(2 alpha) on calcium signaling in resident cells of digested cat iris sphincter, a tissue which exhibits contractile responses to both agonists. Constant superfusion conditions obviated effective conversion of bimatoprost. Serial challenge with 100 nM bimatoprost and prostaglandin F(2 alpha) consistently evoked responses in different cells within the same tissue preparation, whereas prostaglandin F(2 alpha) and 17-phenyl prostaglandin F(2 alpha) elicited signaling responses in the same cells. Bimatoprost-sensitive cells were consistently re-stimulated with bimatoprost only, and prostaglandin F(2 alpha) sensitive cells could only be re-stimulated with prostaglandin F(2 alpha). The selective stimulation of different cells in the same cat iris sphincter preparation by bimatoprost and prostaglandin F(2 alpha), along with the complete absence of observed instances in which the same cells respond to both agonists, strongly suggests the involvement of distinct receptors for prostaglandin F(2 alpha) and bimatoprost. Further, prostaglandin F(2 alpha) but not bimatoprost potently stimulated calcium signaling in isolated human embryonic kidney cells stably transfected with the feline- and human-prostaglandin F(2 alpha) FP-receptor and in human dermal fibroblast cells, and only prostaglandin F(2 alpha) competed with radioligand binding in HEK-feFP cells. These studies provide further evidence for the existence of a bimatoprost-sensitive receptor that is distinct from any of the known prostaglandin receptor types.     

Short-term desensitization of prostaglandin F2 alpha receptors increases cyclic AMP formation and reduces inositol phosphates accumulation and contraction in the bovine iris sphincter

The effect of short-term prostaglandin (PG) desensitization on PGF2 alpha receptor-mediated inositol phosphates accumulation, 1,2-diacylglycerol production, measured as phosphatidic acid (PA), myosin light chain (MLC) phosphorylation, cAMP formation and contraction was investigated in bovine iris sphincter smooth muscle. We have found that incubation of the sphincter with 25 microM PGF2 alpha for 45 min leads to: (a) significant loss in sensitivity of the tissue to PGF2 alpha receptor-stimulated inositol phosphates accumulation, PA production, MLC phosphorylation and contraction, and (b) significant increase in both basal and PGF2 alpha-stimulated cAMP formation. These changes are probably not due to reduction in phospholipid synthesis because there were no detectable differences in basal phospholipid labeling, either from 3H-inositol or from 32P, between normal and desensitized muscles. Preincubation of the sphincter in the absence of PGF2 alpha for 45 min did not lead to alterations in the biochemical-pharmacological responsiveness of the control muscle to PGF2 alpha. Our results suggest that desensitization of PG receptors in the iris sphincter occurs by a receptor-specific process. The PG receptor mediating contraction (IP3-Ca2+) is selectively susceptible to desensitization, in contrast with the receptor mediating smooth muscle relaxation (cAMP). These findings add further support to the developing hypothesis that there are functional and biochemical reciprocal interactions between the IP3-Ca2+ and cAMP messenger systems in the iris of the mammalian eye.     

The effect of sodium iodate on the ciliary body/iris in rabbits

In an attempt to damage the secretory mechanism for ascorbic acid, the effect of the oxidizing agent sodium iodate on the ciliary body has been studied. A decrease in intraocular tension and an inconstantly occurring mydriasis was observed. Histological sections revealed oedema of the ciliary processes mainly localized in the iridial portion, and an increase in vascular permeability of the ciliary body was demonstrated by the carbon labelling technique. Apart from remarkably few endothelial fenestrae, the iridial vessels also showed some intercellular gaps in the endothelial wall. The ascorbic acid concentration was low both in aqueous humour and serum leaving the aqueous/serum ratio roughly unchanged, and the aqueous protein was only slightly raised. All changes were transient phenomena being reversed about 14 days after sodium iodate administration. It is concluded that the reduced ascorbic acid concentration in the aqueous humour reflects the low serum level, and that the secretory mechanism in the ciliary body is essentially unaltered after sodium iodate injection.     

Mathematical and computer simulation modelling of intracameral forces causing pupil block due to air bubble use in Descemet's Stripping Endothelial Keratoplasty: the mechanics of iris buckling

Background: Descemet's Stripping Endothelial Keratoplasty has been associated with a steep learning curve. Angle closure post Descemet's Stripping Endothelial Keratoplasty has been reported, either because of air posterior to the iris causing iridocorneal adhesions, or by air anterior to the iris causing pupillary block. The mechanics of floppy iris syndrome and pupil block have not been discussed. Methods: We evaluated the various forces competing within the anterior chamber via mathematical modelling and computational simulation, and considered the influence of floppy iris on pupil block glaucoma. Results: Energy formulae suggest a critical pressure value will maintain normal anterior chamber relationships, above which abnormal iris buckling may occur. This mechanical instability can be influenced intraoperatively by abnormal iris properties and intracameral forces (such as air). This critical value is lowered if the patient has a floppy iris (because of a lower elastic modulus, a mechanical measure of iris rigidity). To demonstrate this mathematical concept, a 3‐D computational model was built. Simulations show that, as intracameral pressure increases, the iris ring can buckle into predictable modes of shapes. Conclusion: This model shows how iris buckling could occur with an intracameral air bubble leading to posterior iris displacement and mechanical pupil block. It also shows that abnormal iris behaviour in IFIS is consistent with the expected predicted buckling of an elastic disc.     

囊袋张力环和虹膜拉钩在白内障手术中的应用进展

在白内障手术中合理应用手术辅助器械能够有效提高手术安全性,减少手术并发症,提高患者视觉质量。囊袋张力环及虹膜拉钩是白内障手术中最主要的手术辅助器械,近年来得到了广泛的应用。本文就其材料与设计的发展、单独或联合在白内障手术中的应用作一综述。