Ciliary Muscle Strips Research Articles

Serotonin-2B/2C Receptors Mediate Bovine Ciliary Muscle Contraction: Role in Intraocular Pressure Regulation.

To study the pharmacological profile of the serotonin (5-hydroxytryptamine [5-HT]) receptor subtype mediating contractions in bovine isolated ciliary muscles. Ciliary muscle strips were isolated from bovine eyeballs and mounted in organ baths containing aerated (95% O2, 5% CO2) Krebs buffer solution maintained at 37°C. Each muscle strip was attached at 1 end to a Grass Force-displacement Transducer connected to a Polyview Computer System for recording changes in isometric tension. After an equilibration period, ciliary muscle strips were exposed to selective agonists and antagonists of 5-HT receptors. Both selective and nonselective agonists for 5-HT produced concentration-dependent contractions of isolated ciliary muscles with the following rank order of potency: BW723C86>α-methyl-5-HT>MK-212>>8-hydroxy-DPAT>quipazine>R-DOI>>5-HT>>tryptamine. The selective 5-HT2 receptor antagonists, M-100907 (5-HT2A), RS-127445 (5-HT2B), and RS-102221 (5-HT2C), produced noncompetitive inhibition of the contractile effects of selective agonists yielding antagonist potency (pKB) values of 251 ± 27.2 nM (n = 4), 52.5 ± 6.3 nM (n = 4), and 79.4 ± 9.5 nM (n = 4), respectively. On the basis of the profile of activity of selective agonists and antagonists, we conclude that the 5-HT2B and 5-HT2C receptor subtypes appear to be the predominant serotonin receptors that mediate the contractile action of this amine in bovine isolated ciliary muscles.

Addition of urea and thiourea to electrophoresis sample buffer improves efficiency of protein extraction from TCA/acetone-treated smooth muscle tissues for phos-tag SDS-PAGE.

Phosphorylation analysis by using phos-tag technique has been reported to be suitable for highly sensitive quantification of smooth muscle myosin regulatory light chain (LC20 ) phosphorylation. However, there is another factor that will affect the sensitivity of phosphorylation analysis, that is, protein extraction. Here, we optimized the conditions for total protein extraction out of trichloroacetic acid (TCA)-fixed tissues. Standard SDS sample buffer extracted less LC20 , actin and myosin phosphatase targeting subunit 1 (MYPT1) from TCA/acetone treated ciliary muscle strips. On the other hand, sample buffer containing urea and thiourea in addition to lithium dodecyl sulfate (LDS) or SDS extracted those proteins more efficiently, and thus increased the detection sensitivity up to 4-5fold. Phos-tag SDS-PAGE separated dephosphorylated and phosphorylated LC20 s extracted in LDS/urea/thiourea sample buffer to the same extent as those in standard SDS buffer. We have concluded that LDS (or SDS) /urea/thiourea sample buffer is suitable for highly sensitive phosphorylation analysis in smooth muscle, especially when it is treated with TCA/acetone.

Force-inhibiting effect of Ser/Thr protein phosphatase 2A inhibitors on bovine ciliary muscle.

Ciliary muscle is a smooth muscle characterized by a rapid response to muscarinic receptor stimulation and sustained contraction. Although it is evident that these contractions are Ca2+-dependent, detailed molecular mechanisms are still unknown. In order to elucidate the role of Ser/Thr protein phosphatase 2A (PP2A) in ciliary muscle contraction, we examined the effects of okadaic acid and other PP2A inhibitors on contractions induced by carbachol (CCh) and ionomycin in bovine ciliary muscle strips (BCM). Okadaic acid inhibited ionomycin-induced contraction, while it did not cause significant changes in CCh-induced contraction. Fostriecin showed similar inhibitory effects on the contraction of BCM. On the other hand, rubratoxin A inhibited both ionomycin- and CCh-induced contractions. These results indicated that PP2A was involved at least in ionomycin-induced Ca2+-dependent contraction, and that BCM had a unique regulatory mechanism in CCh-induced contraction.

Signaling cross-talk between cannabinoid and muscarinic systems actives Rho-kinase and increases the contractile responses of the bovine ciliary muscle

The aim of the present study was to evaluate the role of a possible interaction between cannabinoid and muscarinic systems, both widely expressed in the ocular structure and involved in the control of bovine ciliary muscle contractility and intraocular pressure modulation. The ciliary muscle strips isolated by bovine eyes were exposed cumulatively to anandamide in the presence and in the absence of carbachol (5nM), in a miograph system for isometric recording. The experiments were also conducted in the presence of AM251 (100nM), 4-DAMP (100nM), Pertussis toxin (500ng/ml), U73122 (0.1 and 1μM), chelerythrine (1 and 10μM) and Y27632 (1 and 10μM). Contractile responses were expressed as the percentage of 10μM carbachol-induced contraction. The anandamide-induced contraction on bovine ciliary muscle strips was enhanced by the previous stimulation of Gq-protein-coupled muscarinic M3 receptors with carbachol. The contractile response to anandamide plus carbachol was affected by different inhibitors such as Pertussis toxin, phospholipase C, protein kinase C and Rho-kinase. The key results of the present study show that sequential activation of muscarinic M3 receptors and cannabinoid CB1 receptors produce synergistic contractile effects of the bovine ciliary muscle by involving the activation of Rho-kinase and protein kinase C.

Nifedipine enhances the relaxant effects of cyclo-oxygenase inhibitors on the bovine ciliary muscle.

The inhibition of cyclo-oxygenase (COX) enzymes and the blockade of Ca (2+) channels play an important role in the regulation of smooth muscle relaxation. This study was designed to investigate the relaxant effects of celecoxib, DFU (5,5-dimethyl-3-(3-fluorophenyl)-4-(4-methylsulphonyl)phenyl-2(5H)-furanone), and indomethacin, cyclo-oxygenase (COX-1 and -2) inhibitors, in the absence or presence of a nifedipine, L-type Ca(2+) channel blocker, on bovine ciliary muscle. Ciliary muscle strips (n = 12) were mounted in organ baths and tested for changes in isometric tension in response to celecoxib, DFU, and indomethacin. The relaxant effects of celecoxib, DFU, and indomethacin on carbachol-induced contractions in the presence or absence of nifedipine were investigated. Celecoxib (10(-7)-10(-4) M), DFU (10(-7)-10(-4) M), indomethacin (10(-7)-10(-4) M), and nifedipine (10(-7)-10(-4) M) inhibited the carbachol-induced contractions in a concentration-dependent manner. The E(max) value of indomethacin was significantly higher than the E(max) values of celecoxib and DFU in ciliary muscle (P < 0.05), with no significant change in pD(2) values (P > 0.05). The relaxation responses by celecoxib, DFU, and indomethacin were significantly increased in the presence of nifedipine (10(-6) M). There were no significant differences between pEC50 and values of celecoxib, DFU, and indomethacin in the absence of nifedipine (10(-6) M) (P > 0.05), but E(max)values were significantly increased (P < 0.05). These results suggest that the celecoxib, DFU, and indomethacin cause relaxation in ciliary muscle precontracted with carbachol. Blockade of calcium channels with nifedipine in ciliary muscle may increase the relaxant effect of celecoxib, DFU, and indomethacin. The topical or systemic use of celecoxib, DFU, and indomethacin with nifedipine can cause blurred near vision due to ciliary muscle relaxation, and in ocular pain conditions caused by ciliary spasm, the pain can be decreased more easily by combined use of these drugs.

Evidence for the Involvement of Cannabinoid CB1Receptors in the Bimatoprost-Induced Contractions on the Human Isolated Ciliary Muscle

To evaluate the bimatoprost effects in the isolated human ciliary muscle and to assess how these response can be modulated by AL8810 and SR141716A. In a myograph system (isometric force measurement), ciliary muscles were exposed cumulatively to PGF(2alpha), latanoprost, travoprost, bimatoprost, and anandamide (0.1 nM-10 microM). Experiments were also conducted in the presence of AL8810 (FP receptor antagonist; 100 nM) or SR141716A (CB(1) receptor antagonist; 10-100 nM). Contractions were expressed as the percentage of 10 microM carbachol-induced contractions. In quiescent tissues, concentration-response curves for bimatoprost, anandamide, PGF(2alpha,) latanoprost, and travoprost were constructed. Bimatoprost showed an important contractile effect on isolated human ciliary muscle strips (E(max) = 125% +/- 0.09%); the maximal effect was higher than that obtained with carbachol. Contractions were inhibited by SR141716A (10 and 100 nM) and AL8810 (100 nM). This study showed evidence of direct interaction of bimatoprost with the contractility of the human ciliary muscle through interaction with cannabinoid CB(1) receptor and prostanoid FP receptors.

Relaxant effects of β-adrenoceptor agonist formoterol and BRL 37344 on bovine iris sphincter and ciliary muscle

We investigated the relaxant effect of β 2-adrenoceptor agonist formoterol and β 3-adrenoceptor agonist BRL 37344 on bovine iris sphincter and ciliary muscle and measured cAMP and cGMP levels. Iris sphincter ( n = 16) and ciliary muscle ( n = 16) strips were mounted in organ baths and tested for changes in isometric tension in response to formoterol and BRL 37344. Their relaxant effects on serotonin-induced contractions in the presence or absence of metoprolol, ICI 118.551 and SR 59230A (β 1-, β 2-, β 3-adrenoceptor antagonist, respectively) were investigated. Their effects on cAMP and cGMP levels in iris sphincter ( n = 12) and ciliary muscle ( n = 12) were evaluated. Formoterol (10 − 11 –10 − 5 M) and BRL 37344 (10 − 10 –10 − 5 M) decreased the serotonine-induced contractions in a concentration-dependent manner. E max values of formoterol were significantly higher than those of BRL 37344 in iris sphincter and ciliary muscle, with no significant change in p D 2 values. The relaxation responses by formoterol and BRL 37344 were antagonized with ICI 118.551 (10 − 6 M) and SR 59230A (10 − 6 M). cAMP levels of formoterol- and BRL 37344-treated tissues were significantly higher than those of the control tissues. cGMP levels of BRL 37344-treated tissues were significantly higher than those of control tissues, but this effect of BRL 37344 was less significant than its effect on cAMP levels. β-adrenoceptor relaxation responses in bovine iris sphincter and ciliary muscle are mediated by a mixed population of β 2- and β 3-adrenoceptor subtypes, with a predominant contribution of cAMP. Potency of formoterol and BRL 37344 was similar, but efficacy of formoterol was better than BRL 37344.

Pharmacological and Functional Characterization of Endothelin Receptors in Bovine Trabecular Meshwork and Ciliary Muscle

To clarify the potential role of endothelin-1 (ET-1) in the pathogenesis of glaucoma, the endothelin receptors expressed in bovine trabecular meshwork (TM) and ciliary muscle (CM) were identified. TM and CM strips were subjected to ET-1 as well as to specific endothelin receptor antagonists. In both tissues BQ123, a specific ET-A receptor antagonist, substantially inhibited ET-1-induced contraction. BQ788, a specific ET-B receptor antagonist, showed only moderate effects. Both ET receptor types were detected in bovine TM and CM using Western blot analysis. ET-1 produced an increase in intracellular calcium in cultured TM cells. This effect was inhibited by BQ123, but not by BQ788. Thus, although both receptors are present, the ET-A receptor appears to play the predominant role in mediating contraction in both the TM and CM, while the ET-B receptor seems to contribute little to the overall ET-1 effect.

Effects of protein kinase inhibitor, HA1077, on intraocular pressure and outflow facility in rabbit eyes.

To elucidate the roles of protein kinase in regulating the intraocular pressure (IOP) and outflow facility in rabbit eyes. A protein kinase inhibitor, 1-(5-isoquinolinesulfonyl)-homopiperazine (HA1077), was used. The IOP and the outflow facility were measured before and after topical, intracameral, or intravitreal administration of HA1077 in rabbits. Western blot analysis was performed to detect the 20-kd light chain of myosin in human trabecular meshwork (TM) cells and bovine ciliary muscle (CM) tissues. The cell morphologic condition and distribution of actin filaments and vinculin in TM cells were studied using cell biology techniques. Carbachol-induced contraction of isolated bovine CM strips following administration of HA1077 was examined in a perfusion chamber. In rabbit eyes, the administration of HA1077 resulted in a significant decrease in IOP in a dose-dependent manner. An increased outflow facility was also observed. Western blot analysis revealed the presence of 20-kd light chain of myosin in human TM cells and bovine CM tissues. In cultured human TM cells, exposure to HA1077 disrupted actin bundles and impaired focal adhesion formation. In addition HA1077 showed relaxation of bovine CM strips. Use of HA1077 caused a reduction in IOP and an increase in the outflow facility. The results of in vitro experiments suggest that the IOP-lowering effects of HA1077 may be related to the altered cellular behavior of TM cells and relaxation of CM contraction. The results of these studies suggested that protein kinase inhibitors have the potential to be developed into a treatment modality for glaucoma.

Nitric Oxide Relaxes Bovine Ciliary Muscle contracted by Carbachol Through Elevation of Cyclic GMP

It is generally accepted that nitric oxide relaxes vascular smooth muscles by activating guanylyl cyclase, which in turn increases cyclic 3′:5′ guanosine monophosphate level. Despite the physiological significance of nitric oxide, very few studies have attempted to characterize the mode of action of this mediator in ciliary muscles. Therefore, the present experiments were designed to investigate whether or not the relaxation induced by sodium nitroprusside as a donor of nitric oxide is accompanied by the increase in cyclic 3′:5′ guanosine monophosphate level in the bovine ciliary muscle, and these responses are affected by methylene blue as an inhibitor of guanylyl cyclase and 3-isobutyl-1-methylxanthine as an inhibitor of phosphodiesterases. The relaxation activity of exogenous 8-bromo-cyclic 3′:5′ guanosine monophosphate was also determined. Sodium nitroprusside produced a concentration-dependent relaxation in the bovine ciliary muscle strips which had been contracted by carbachol as a cholinergic agonist. Relaxation in response to sodium nitroprusside was accompanied by a significant (P<0.05 andP<0.005) increase in cyclic 3′:5′ guanosine monophosphate level. The relaxation response and the increase in cyclic 3′:5′ guanosine monophosphate caused by sodium nitroprusside were significantly (P<0.01 andP<0.05) augmented by the pretreatment with 3-isobutyl-1-methylxanthine, and were significantly (P<0.005 andP<0.05) attenuated in the presence of methylene blue. The exogenously applied 8-bromo-cyclic 3′:5′ guanosine monophosphate relaxed the ciliary muscle strips in a concentration-dependent manner. These results suggest that nitric oxide causes relaxation of the bovine ciliary muscle through the activation of guanylyl cyclase and an increase in cyclic 3′:5′ guanosine monophosphate level.

A Possible Role of Endogenous Inhibitor for Nitric Oxide Synthesis in the Bovine Ciliary Muscle

The present experiments were designed to investigate the possible role of endogenous methylarginine derivatives such asNG-monomethyl-L-arginine, asymmetricalNG,NG-dimethyl-L-arginine and symmetricalNG,NG-dimethyl-L-arginine for the nitric oxide synthesis in the bovine ciliary muscle. The contents of asymmetricalNG,N″G-dimethyl-L-arginine and symmetricalNG,NG-dimethyl-L-arginine in the bovine ciliary muscle were determined to be 370.2±27.6 (n=5) and 182.4±22.9 (n=5) pmoles g−1wet weight, respectively by means of the automated high-performance liquid chromatography.NG-Monomethyl-L-arginine was below the assay limits. On the basis of the total tissue water content (0.792±0.006 ml g−1wet weight,n=14), the concentrations of asymmetricalNG,NG-dimethyl-L-arginine and symmetricalNG,NG-dimethyl-L-arginine were tentatively estimated to be (4.7±0.3)×10−7M(n=5) and (2.3±0.3)×10−7M(n=5), respectively. A23187 (10−7–3×10−6M) produced a concentration-dependent relaxation of the ciliary muscle strips which had been contracted with 10−5Mcarbachol. Authentic asymmetricalNG,NG-dimethyl-L-arginine (3×10−6–3×10−4M), but not symmetricalNG,N″G-dimethyl-L-arginine (3×10−4M), inhibited the 10−6MA23187-induced relaxation in a concentration-dependent manner. The inhibition with asymmetricalNG,NG-dimethyl-L-arginine (10−4M) was reversed by an addition of 3×10−3ML-arginine, but not by 3×10−3MD-arginine.>The A23187 (10−6M)-induced relaxation was enhanced by 3×10−3ML-arginine or superoxide dismutase (50 U ml−1), whereas it was inhibited by carboxy-PTIO (3×10−4M), a scavenger of nitric oxide, or methylene blue (10−5M), an inhibitor of guanylate cyclase. The carbachol-induced contraction was enhanced by asymmetrical,NG,N″G-dimethyl-L-arginine (10−5M) and inhibited by 3×10−3ML-arginine. Any effect of prostanoid formation during the A23187-induced relaxation was ruled out by using indomethacin (10−5M). Sodium nitroprusside (10−5M), a donor of nitric oxide, also produced a relaxation, which was inhibited by methylene blue (10−5M) or carboxy-PTIO (3×10−4M) and was augmented by superoxide dismutase (50 U ml−1), but unaffected by asymmetricalNG,NG-dimethyl-L-arginine (3×10−4M) orL-arginine (3×10−3M). These results lead us to speculate that the nitric oxide synthesized endogenously fromL-arginine may play a role for mediating relaxation of the bovine ciliary muscle and that the endogenous asymmetricalNG,N″G-dimethyl-L-arginine may be involved in inhibiting the biosynthesis of nitric oxide when there are increased intracellular concentrations of the methylarginine under certain circumstances.

Nitric oxide-induced ciliary muscle relaxation during contraction with endothelin-1 is mediated through elevation of cyclic GMP

PURPOSE. Nitric oxide (NO) relaxes ciliary smooth muscle, and endothelin-1 (ET-1) is reported to regulate ciliary muscle tone. Despite the physiological significance of nitric oxide and ET-1, very few studies have attempted to characterize the mutual modes of action of these mediators in this tissue. Thus, the present experiments were designed to investigate a possible relaxation mechanism of nitric oxide in bovine ciliary muscle that has been contracted by ET-1. METHODS. The effects of sodium nitroprusside (SNP), as a nitric oxide donor, methylene blue, as an inhibitor of guanylate cyclase, and 8-bromo-cyclic GMP on the bovine ciliary muscle contracted with ET-1 were examined. The changes in cyclic GMP level and relaxation, in response to SNP alone or in combination with 3-isobutyl-1-methylxanthine (IBMX) as a nonselective inhibitor of phosphodiesterases, were also determined. RESULTS Results. Sodium nitroprusside (SNP) produced a concentration-dependent relaxation, which was significantly (p < 0.005) augmented by 10 -5 M 3-isobutyl-1-methylxanthine (IBMX) and significantly (p < 0.005) attenuated by 3 × 10 -5 M methylene blue as an inhibitor of guanylate cyclase. The relaxation in response to SNP was accompanied by an increase in the cyclic 3':5' guanosine monophosphate (cyclic GMP) level, which was again significantly (p < 0.05) augmented by 10 -5 M IBMX and significantly (p < 0.005) attenuated by 3 × 10 -5 M methylene blue. The exogenously applied 8-bromo-cyclic GMP relaxed the ciliary muscle strips during the contraction caused by ET-1. CONCLUSIONS. These results lead us to assume that NO generated from SNP is closely related to cyclic GMP production via the activation of guanylate cyclase and, in turn, causes a relaxation response in the bovine ciliary muscle contracted with ET-1.

The effects of endothelin-1 on isolated bovine ciliary muscles

The effects of endothelin-1 on bovine ciliary muscle were investigated in vitro using muscle strips prepared in two different directions (longitudinal and circular). Fifty-two bovine ciliary muscle strips (4 x 6 mm) were prepared. Chemicals were added to both types of strips suspended in an organ chamber, and their changes in isometric tension were recorded. The concentration-response relationship of endothelin-1 (n = 20), the magnitudes of contractions caused by endothelin-1 and carbachol (n = 12), and the effects of an endothelin receptor subtype A antagonist BQ123 (n = 4) and an endothelin receptor subtype B agonist IRL1620 (n = 4) were studied. The cumulative addition of endothelin-1 caused relaxation at low concentrations (10(-11) and 10(-10) M), while it caused contraction of the ciliary muscles at high concentrations (10(-9), 10(-8) and 3 x 10(-8) M). The magnitude of the contraction caused by 10(-8) M endothelin-1 was about 30% in the longitudinal muscle strips and 29% in the circular muscle strips, relative to the contraction caused by 10(-5) M carbachol, a muscarinic agonist. The contractile response caused by 10(-8) M endothelin-1 was abolished and converted to relaxation by pretreatment with BQ123, a selective endothelin receptor subtype A antagonist. Single addition of IRL1620, a selective endothelin receptor subtype B agonist, caused only relaxation. These results suggest that, endothelin-1 causes not only contraction at high concentrations, but also relaxation at low concentrations in bovine ciliary muscle. Also, it suggested that the relaxation is mediated by endothelin receptor subtype B, whereas the contraction is mediated by endothelin receptor subtype A.

Regulation of outflow rate and resistance in the perfused anterior segment of the bovine eye

Contractile properties of isolated trabecular meshwork strips have recently been described. In the present paper we characterize the regulation of the outflow pathway in the isolated perfused anterior segment of the bovine eye. Anterior segments of bovine eyes with detached iris, ciliary body and ciliary muscle were perfused at constant pressure of 8.8 mmHg. A constant outflow of approximately 6-8 microliters min-1 could be obtained for at least 3 hr. The calculated outflow resistance was in the range 1.1-1.4 mmHg min microliter-1. The relative outflow was significantly reduced after application of carbachol, reaching a maximal inhibition of 30%. EC50 for carbachol was 3 x 10(-8) mol l-1. Atropin completely blocked the effect of carbachol on outflow. Morphological examination of perfused anterior segments which were perfused with carbachol revealed an intact fine structure of the meshwork cells. Pilocarpine at 10(-5) mol l-1 reduced outflow by 15%. Epinephrine at 10(-5) mol l-1 reduced outflow, while epinephrine at 10(-6) mol l-1 slightly increased the outflow rate. This effect could be blocked by metipranolol. Endothelin-1 in concentrations of 2 x 10(-9) and 2 x 10(-8) mol l-1 inhibited relative outflow by > 30%. Carbachol, pilocarpine, endothelin and a high dose of epinephrine, which have been shown to induce contractions in isolated bovine trabecular meshwork and ciliary muscle strips, induced a reduction of outflow rate and an increase of outflow resistance of the anterior segment. Thus, at least in the bovine eye, the trabecular meshwork per se is directly involved in the regulation of aqueous humor outflow.

Age does not affect contractile responses of the isolated rhesus monkey ciliary muscle to muscarinic agonists.

In primates, ciliary muscle contraction causes accommodation and facilitates aqueous outflow. In living rhesus monkeys, accommodative, outflow facility, and ciliary muscle movement responses to cholinergic agonists all decline with age. We developed an apparatus to determine in vitro whether the latter is related to intra- or extra-ciliary muscle factors, and whether ciliary muscle contraction in the coronal (putatively more accommodation-relevant) and longitudinal (putatively more facility-relevant) vectors can be dissociated pharmacologically. In fresh ciliary muscle strips, carbachol and aceclidine each induced dose-dependent contraction in the longitudinal and coronal vectors. With neither drug was there any apparent dissociation of the responses in the two vectors. Atropine pretreatment completely prevented a supramaximal dose of carbachol from inducing ciliary muscle contraction in either vector. Ciliary muscle strips responded to several cholinergic agonists as well on day 2 (24-32 hours post-enucleation) as on day 1 (1-9 hours post-enucleation) when kept in a cell culture medium at 4 degrees C. By light microscopy, the general architecture of the ciliary muscle, the muscle bundles, and the single muscle cells appeared normal; however, cellular and nuclear swelling were apparent following the 32-hour culturing period. Contractile responses to near-maximal doses of carbachol and aceclidine did not vary markedly with age in either vector, suggesting that the age-related decrease in ciliary muscle mobility in vivo is due to extra-muscular restrictive factors rather than diminished muscular contractility.

Measurements of intracellular calcium and contractility in human ciliary muscle

Electromechanical and pharmacomechanical coupling was investigated in human ciliary muscle by measuring the intracellular free calcium in single cultured ciliary muscle cells and the contractility in meridional ciliary muscle strips. The basal resting calcium concentration was 75 +/- 8.7 nmol/l, n = 23. Application of acetylcholine (0.1 mmol/l) and carbachol (0.1 mmol/l) resulted in an initial [Ca2+]i peak followed by a recovery phase and a [Ca2+]i plateau. The initial [Ca2+]i peak was still observed in the absence of extracellular calcium and in the presence of verapamil (0.1 mmol/l). During its plateau [Ca2+]i was decreased by withdrawal of extracellular calcium or application of verapamil (0.1 mmol/l). Depolarization induced by a high level of extracellular potassium yielded only a small transient [Ca2+]i peak without a [Ca2+]i plateau. In isolated ciliary muscle strips, muscarinic stimulation (carbachol 0.1 mmol/l) resulted in an initial phasic and a subsequent tonic contraction. Removal of external calcium reduced the phasic contraction to 30.6 +/- 4.4% (n = 8) and completely abolished the tonic one. Verapamil (0.1 mmol/l) had only a slight relaxing effect when applied during the tonic contraction. We conclude that human ciliary muscle contraction is mediated by calcium release from intracellular stores and calcium entry through calcium channels, which are most probably receptor-operated. Depolarization of the muscle cell membrane and calcium entry through voltage-operated calcium channels do not contribute significantly to human ciliary muscle contraction.

Differential smooth muscle-like contractile properties of trabecular meshwork and ciliary muscle

The contractile properties of bovine trabecular meshwork and ciliary muscle strips were investigated using an electromagnetic force-length transducer for isometric force measurements. Acetylcholine, pilocarpine and aceclidine administration resulted in dose-dependent contractions of trabecular meshwork and ciliary muscle. Absolute forces were approximately 10 times larger in ciliary muscle than in trabecular meshwork. Maximal force evoked by aceclidine (5 × 10 −5 m), when compared to the pilocarpine (5 × 10 −5 m) response, was significantly higher in trabecular meshwork than in ciliary muscle. The results were 172·5 ± 12·6% ( n = 7) and 138·9±4·0% ( n = 8, P < 0·05), respectively. Depolarization induced by raised external potassium (120 m m), when compared to the acetylcholine response (10 −3 m), resulted in a small contraction of 19·3±4·2% in trabecular meshwork ( n = 5), and of 59·0±13·7% in ciliary muscle ( n = 4, P < 0·01). Both responses were inhibited by atropine (10 −5 m). The differential potassium effect may be explained by the large number of cholinergic nerve endings in ciliary muscle as compared to trabecular meshwork tissue. Recently, a dissociation between the effects of aceclidine on outflow resistance and accommodation has been described. Our data are consistent with these observations and provide evidence for a direct role of trabecular meshwork contractility in aqueous outflow regulation.

Endothelin-evoked contractions in bovine ciliary muscle and trabecular meshwork: Interaction with calcium, nifedipine and nickel

In the present study, we compared contractile responses of isolated bovine ciliary muscle and trabecular meshwork strips to endothelin-1 and carbachol. 1. Endothelin-1 is a potent contracting agent for ciliary muscle and trabecular meshwork. The EC50 was 5 x 10(-9) mol/l for both tissues. The maximal force evoked by endothelin was 73% of the maximal carbachol response in trabecular meshwork and 52% in ciliary muscle. 2. Carbachol contracted both tissues with an EC50 of 2 x 10(-7) mol/l. 3. In ciliary muscle, the tension was completely dependent on extracellular calcium. 4. In trabecular meshwork, 23 +/- 4% of the endothelin- and 42 +/- 10% of the carbachol-induced force response remained after removal of extracellular calcium. 5. Nifedipine (10(-5) mol/l) had only a slight relaxing effect in both tissues. 6. Nickel (10(3) mol/l) inhibited the development of force in both tissues. The relaxation induced by nickel was more pronounced in endothelin- than in carbachol-induced contractions. Different intracellular mechanisms mediating the action of endothelin and carbachol can be described: i) a calcium- and a nickel-sensitive pathway in both tissues and ii) an additional mechanism independent of external calcium in trabecular meshwork. These results indicate functional differences between the contractile region of trabecular meshwork and ciliary muscle. Endothelin may participate in accommodation and regulation of the intraocular pressure.

Activity of prostaglandin E, F, A and B on sphincter, dilator and ciliary muscle preparations of the cat eye

Both sphincter and dilator muscle preparations of the cat iris contract to prostaglandins; F 2α and E 2 are the most potent and A 1 and B 1 the least. Ciliary muscle strips relax to PG's provided that the strips are precontracted. E 1, E 2 and often F 2α are more potent relaxants than the remaining PG's. The effects of PG's are not altered by α or β blockade nor by atropine; however, propranolol blocks the PG induced relaxation of the ciliary muscle. The effects of PG's on the sphincter are antagonized by catecholamines; but the latter act synergistically in contracting the dilator and in relaxing the ciliary muscle. Indomethacin markedly potentiates the effects of PG's on all three muscle preparations.

Steroid eye drops and their components. Effect on isolated intraocular muscles of monkeys.

The effects of steroid eye drops were tested on isolated preparations of the sphincter, dilator, and ciliary muscles of the monkey. The resting tension of sphincter and ciliary muscle strips decreased in the presence of the topical steroid preparations while that of the dilator muscle increased, and muscle contractions evoked by acetylcholine, epinephrine, serotonin, or potassium chloride were markedly inhibited. In all cases the steroids when tested without vehicles were ineffective while responses to the vehicles alone were identical to those of the steroid-containing eye drops. Some ingredients of the vehicles are capable of altering smooth muscle function in vitro possibly by disturbance of the permeability of the cell membrane.