Chromatographic characterization of vasoactive intestinal polypeptide in guinea pig and rhesus monkey eyes

Acid extracts of guinea pig and rhesus monkey anterior uvea, choroid and retina contain immunoreactive VIP. By reversed phase high performance liquid chromatography, the immunoreactive material from these ocular tissues elutes at a position similar to synthetic porcine VIP. Only in the guinea pig anterior uvea is a second smaller peak detected. By size exclusion high performance liquid chromatography, the peptide in the monkey anterior uvea, choroid and retina elutes in the identical position as the synthetic porcine VIP standard with an apparent molecular weight of 3450 daltons. We conclude that a single form of VIP, chromatographically similar to the porcine standard, is the predominant form of the peptide in the eye of guinea pig and rhesus monkey.     

Effects of oculomotor nerve stimulation on ocular blood flow in rabbits after sympathetic denervation.

Bilateral cervical sympathectomy was performed in rabbits to produce sympathetic denervation of both eyes. The effect was checked by testing the sensitivity of the dilator muscle of the iris. The effect of intracranial stimulation of the oculomotor nerve on ocular blood flow was determined 3–8 weeks after sympathectomy using radioactively labelled microspheres. Indomethacin was given to prevent the release of prostaglandins and biperiden was used to abolish muscarinic effects in the eye. Insertion of the electrode and/or test stimulation tended to cause a long-lasting vasodilation in the uvea. In the iris the blood flow on the side of the electrode was 142±13% of that in the control eyes before stimulation of the nerve. During stimulation it was 75±13% of that in the control eyes. In the ciliary processes the corresponding figures were 118±16% and 79±7% and in the choroid 105±5% and 106±5%, respectively.The results make it seem very unlikely that the vasoconstrictive response to oculomotor nerve stimulation is due to adrenergic nerves within the anterior uvea, and it is probably caused by some transmitter other than norepinephrine.     

Role for nitric oxide in the hyperpermeability and hemodynamic changes induced by intravenous VEGF

PURPOSE: To explore the effects of brief intravenous (IV) infusion of vascular endothelial growth factor (VEGF) on vascular albumin permeability, blood flow, and vascular conductance (blood flow normalized to arterial blood pressure) in ocular tissues and brain and to assess the role of nitric oxide in mediating these changes. METHODS: A quantitative, double-tracer, radiolabeled albumin permeation method was combined with radiolabeled microspheres for assessment of changes in vascular permeability and blood flow, respectively, induced in ocular tissues by IV infusion of recombinant human VEGF165 for 20 minutes (80-450 picomoles/kg body weight). An inhibitor of nitric oxide synthase (NOS), NG-monomethyl-L-arginine (L-NMMA; 50 micromoles/kg body weight infused simultaneously with VEGF), was used to explore the role of nitric oxide in mediating the vascular changes induced by VEGF. RESULTS: Infusion of VEGF165 in thiopental-anesthetized rats dose-dependently increased 125I albumin permeation in the retina, anterior uvea, and choroid/sclera and in brain, aorta, lung, kidney, small intestine, and peripheral nerve. Mean arterial blood pressure, cardiac output, and stroke volume were decreased only at the highest dose of VEGF, whereas heart rate remained unchanged. Blood flow was increased in the anterior uvea, and vascular conductance was increased in retina, anterior uvea, choroid/sclera, and brain at the highest dose of VEGF. The NOS inhibitor, L-NMMA, blocked VEGF-induced vascular hyperpermeability in all ocular and nonocular tissues, prevented the increase in vascular conductance in all ocular tissues, and blocked the decrease in mean arterial blood pressure, cardiac output, and stroke volume. Infusion of L-NMMA alone decreased vascular conductance in choroid/sclera and kidney, slightly increased mean arterial blood pressure, and in general, did not affect 125I-albumin permeation. (L-NMMA slightly decreased albumin permeation in the retina and increased it in the brain.) CONCLUSIONS: Intravenous infusion of VEGF can acutely impair endothelial cell barrier functional integrity and relax resistance arterioles in ocular tissues and brain through a mechanism involving activation of NOS.     

The influence of ions, ouabain, propranolol and amiloride on the transepithelial potential and resistance of rabbit cornea.

By means of in vitro ion substitution experiments the potential difference and resistance dependency of rabbit corneal epithelium on Na⁺, K⁺ and Cl^? activities have been established at the tear side as well as at the aqueous side. In normal conditions only a K⁺ selectivity at the aqueous side of the epithelium exists and a Na⁺ and Cl^? selectivity at the tear side. The tear side was also K⁺ selective at K⁺ activities greater than 28 mm. In Cl^? Ringer bathing solutions a rectifying current-voltage relationship is obtained. Ouabain gives rise to a two phase decrease in potential from 29·9±2·4 to 22·9±2·1 to 14·4±1·7 mV (n = 9). In the first phase the resistance changes from 3·7±0·4 to 4·0±0·4 kΩcm² (n = 9) and in the second phase R decreased to 1·3±0·1 kΩcm² (n = 9). After ouabain the Na⁺ selectivity has disappeared and the Cl^? permeability has increased. The current-voltage relationship after ouabain has become linear. Amiloride induced a reversible change in resistance from 3·5±0·6 to 4·8±0·8 kΩcm² accompanied by a depolarization from 29·1±2·7 to 22·6±1·5 mV (n = 4), moreover the rectifying properties as exhibited in control cornea are not affected by amiloride. A voltage dependent Cl^?-conductivity of the tear side membranes of the corneal epithelium can be postulated as well as a voltage independent Cl^?-conductivity that can be induced by ouabain. The ouabain effects are inhibited by propranolol.     

Vasoactive intestinal polypeptide (VIP)-immunoreactive nerve fibers in the anterior uvea of the guinea pig

The presence of vasoactive intestinal polypeptide (VIP)-like immunoreactivity, especially in the anterior part of the guinea pig uvea, was studied using an indirect immunofluorescence technique. In the ciliary body, thin varicose VIP fibers displaying VIP immunoreactivity were observed in the ciliary muscle and in the ciliary processes. The iris was virtually free of VIP fibers. In the choroid, VIP-immunoreactive fibers were seen mainly in close association with the choroidal blood vessels. The histochemical localization of VIP-immunoreactive fibers in the anterior uvea suggests that the physiological role of neuronal VIP in the uvea is not only associated with the regulation of the blood flow of the choroid but also with the functions of ciliary muscle and ciliary epithelium.     

Ocular effects of clonidine in cats and monkeys (Macaca irus)

In monkeys administration of about 0·01 μg clonidine into the anterior chamber caused no significant change in the facility of outflow. At 10 μg/kg body weight given intravenously the facility decreased by 0·09±0·01 μl · min^?1 · mmHg^?1 from a level of 0·51±0·05 μl · min^?1 · mmHg^?1. The same dose had no significant effect on the rate of aqueous humor formation determined with an indicator dilution technique. Before the administration of the drug the formation of aqueous humor was 1·43±0·13 μl/min, and 30 min later it was 1·32±0·15 μl/min. The fall in eye pressure was 1·0±0·3 mmHg. In cats 2–10 μg/kg body weight clonidine given intravenously reduced the intrascleral venous pressure and the eye pressure in a way which indicated marked vasoconstriction in the eye. Unilateral carotid injection of the drug at a dose reducing the homolateral eye pressure by 4–5 mmHg caused a 19% reduction in the choroidal blood flow, a 34% reduction in the blood flow through the iris and a 42% reduction in the blood flow in the ciliary body when the injected side was compared to the other.     

Characterization of nitrergic neurons in the porcine and human ciliary nerves

PURPOSE: To further characterize a subpopulation of choroidal ganglion cells associated with the ciliary nerves. METHODS: Isolated long ciliary nerves of porcine and human eyes containing ciliary nerve-associated ganglion cells (CNGCs) were embedded in Epon for ultrastructural investigation, or wholemounts were stained with antibodies against nitric oxide synthase (NOS), vasoactive intestinal polypeptide (VIP), vesicular acetylcholine transporter, neuropeptide Y (NPY), tyrosine hydroxylase (TH), calcitonin gene-related peptide (CGRP), substance P (SP), and synaptophysin. In addition, wholemount preparations of the choroid and of the anterior segment were stained for reduced nicotinamide adenine dinucleotide phosphate-diaphorase (NADPH-D). Serial sections through choroid and anterior segment were stained with the prior antibodies listed. RESULTS: In the porcine choroid only CNGCs were present. They stained for brain (b)NOS and VIP and were surrounded by SP and VIP-immunoreactive (IR) nerve terminals. The axonal processes of the CNGCs followed the ciliary nerves to the anterior eye segment, where they formed a nerve fiber plexus that terminated in the trabecular meshwork. None of the axons passed into the sparse NOS-IR nerve fiber plexus surrounding the choroidal vasculature. The CNGCs in the human choroid morphologically resembled those seen in the pig. CONCLUSIONS: The CNGC proportion of choroidal ganglion cells is presumably involved in the intrinsic (peripheral) innervation of the aqueous outflow tissues and of the choroid.     

Neuroanatomy and neurochemistry of rat cornea: Changes with age

PurposeTo characterize the entire rat corneal nerve architecture, the changes that occur with aging, and its sensory, sympathetic, and parasympathetic fiber distribution.MethodsSprague-Dawley rats (aged 1 day to 2 years old) of both sexes were euthanized, and the whole corneas were immunostained with protein gene product 9.5 (PGP9.5). The specimens were double-labeled with antibodies against calcitonin gene-related peptide (CGRP) and substance P (SP) as sensory nerve markers, vasoactive intestinal peptide (VIP) as a parasympathetic nerve marker, and neuropeptide Y (NPY) and tyrosine hydroxylase (TH) as markers of sympathetic fibers. Relative nerve density positive for each antibody was assessed by computer-assisted image analysis.ResultsThick nerve trunks enter the cornea in the middle of the stroma and run towards the anterior stroma, subsequently dividing into smaller branches that penetrate upwards into the epithelium to form the subbasal nerve bundles. There was no significant difference in corneal innervation between sexes. CGRP and SP were the major sensory neuropeptides with 47.6% ± 3.5% and 34.9% ± 5.1%, respectively, of the total nerves. VIP was 18.4% ± 5.7%, and NPY and TH positive fibers took up 6.92% ± 2.66% and 2.92% ± 1.52%, respectively. Epithelial nerve density increased with age, reached full development at 5 weeks, and decreased at 120 weeks.ConclusionThis study provides a complete nerve architecture and content of components of sensory, parasympathetic, and sympathetic nerves in the rat cornea. The normal innervation pattern described here will provide an essential baseline for investigators who use the rat model for assessing corneal pathologies that involve nerve alterations.     

Effects of intracranial oculomotor nerve stimulation on ocular blood flow in rabbits: Modification by indomethacin

The oculomotor nerve was stimulated intracranially and regional blood flow in the eye was determined by using labelled microspheres. Stereotactic insertion of the electrode and/or electrical stimulation to test its position caused a long-lasting increase in choroidal blood flow. This effect could not be prevented by pretreatment with 20 mg/kg body weight indomethacin. Stimulation of the nerve 10–20 min after the insertion of the electrode caused miosis and a marked reduction in blood flow in the anterior uvea. In the ciliary processes that could be scraped off the iris the blood flow was 24±7% lower than on the control side. In the rest of the iris-ciliary body preparation the flow was reduced by 53±4%. In animals pretreated with indomethacin the corresponding figures were 51±8% and 75±4%, respectively. The results indicate that the stimulation applied caused a vasoconstriction in the anterior uvea which was potentiated by indomethacin. The anatomical basis for this effect and for the long-lasting vasodilation that was produced in the choroid remains unknown.     

delta 9-Tetrahydrocannabinol transport in rabbit eyes

The rabbit anterior uvea preparation in vitro accumulated Δ⁹-tetrahydrocannabinol (THC). The concentrating mechanism was temperature and oxygen dependent and required glucose in the media. Dinitrophenol, iodoacetate, and cyanide inhibited THC accumulation. Ouabain, iodide, perchlorate, probenecid, iodipamide, and prostaglandin E₁ did not alter the accumulation ratio. Para-aminohippurate enhanced THC uptake. Following intravitreal injection [³H]THC exited rapidly from the eye with a half-time of approximately 1·4 hr compared to a half-time of 15·8 hr for [¹⁴C]sucrose.     

The kinetics and energy dependence of prostaglandin transport processes. I. In vitro studies on the rate of PGF2α accumulation by the rabbit anterior uvea

The kinetics, energy dependence and specificity of prostaglandin (PG) accumulation by the anterior uvea of rabbits was studied in vitro by measuring the uptake of [³H]-PGF_2α under a variety of conditions. The initial accumulation process, 5-min uptake at 37°C, conformed to Michaelis-Menten kinetics, exhibiting an apparent K_t of 34 μm and a J_max of 13·6 nmol×g tissue^?1×min^?1. Accumulation was inhibited by bromcresol green (BrCG; 10 μm) or incubation at 0°C. PGE₂ (14 μm) or PAH (10 mm) increased the apparent PGF_2αK_t from 34 to 50 and 151 μm, respectively, without changing the J_max. The calculated K_i (2700 μm) of PAH on PGF_2α accumulation was approximately 20 times greater than the apparent K_t of PAH transport (127 μm). Probenecid (50 μm) increased the apparent K_t of PGF_2α transport to 104 μm and caused a 50% reduction in J_max. PGF_2α transport rate was reduced in the absence of glucose or oxygen, or in the presence of 0·1 mm iodoacetic acid. Ouabaiu (10^?5m) or substitution of Na with Li or mannitol significantly reduced the rate of PGF_2α accumulation by the anterior uvea. It was concluded that in vitro PGF_2α accumulation by the anterior uvea, and presumably the in vivo PG transport system of this tissue, is a saturable, energy and sodium dependent mechanism. PGE₂ and PGF_2α appear to share the same carrier while PAH interacts with, but is not primarily transported by, the PG system. The transport constants of this system indicate that anterior uveal PG transport would be fully effective even at the highest expected local PG concentration and would not be saturated even at pathological PG levels.     

Effects of calcitonin gene-related peptide and substance P on regional blood flow in the cat eye

The effects of calcitonin gene-related peptide (CGRP) and substance P (SP) on the regional blood flow of the eye were studied in cats. The animals were anaesthetized and the eyes were cannulated for intracameral administration of the test substances and intraocular pressure measurement. Regional blood flow was determined using the radioactively labelled microsphere method. Intracameral injection of 1.3 x 10(-9) mol of CGRP increased markedly the blood flow of the iris, the ciliary body, and the sclera. There was no clear-cut effect in the choroid or in the retina. Intracameral administration of 1.3 x 10(-9) mol of SP had no clear-cut effect on the blood flow of any of the ocular tissues studied. In addition, CGRP reduced the intraocular pressure statistically significantly, whereas SP had no effect. The results of the present study indicate that CGRP is a potent vasodilator in the anterior uvea of the cat eye when administered from the adventitial side, whereas SP seems to have little or no effect.     

A comparison of the miotic and inflammatory effects of biologically active polypeptides and prostaglandin E2 on the rabbit eye

The ocular effects of some biologically active peptides were studied and compared to those of prostaglandin E₂ (PGE₂) to determine whether the responses of the eye to trauma, characterized by increased intraocular pressure (IOP), the development of anterior chamber flare and partial miosis, might be mediated by such peptides. One to 2 hr after intravitreal injection of 10 μg of PGE₂ into rabbit eyes, ocular hypertension, flare and iridial hyperemia, but only minimal miosis were observed. Maximum miosis developed within 2–3 hr after intravitreal injection of 1·0–100 μg of substance P (SP), SP-octapeptide (SP-8), coherin or eledoisin-related peptide (EDR), while 10–100 μg of vasoactive intestinal polypeptide (VIP), somatostatin or bradykinin (BK) yielded only submaximal miosis and angiotensin II, α-MSH and poly-dl-alanine had little or no miotic effect. None of these peptides caused iridial hyperemia or a cellular invasion of the anterior chamber and only high doses (100 μg) of VIP or BK caused significant increases in the protein concentration of the aqueous humor. Miotic doses of SP, SP-8 or EDR caused a significant increase in IOP in some, but not all, experiments. Thus, PGE₂ can be regarded as a mediator of the ocular irritative response although it may not account for the miosis that is associated with chemical or surgical trauma. However, this autacoid should not be regarded as the mediator of ocular inflammation since it does not elicit a cellular response. In contrast, some polypeptides, particularly SP, SP-8 and EDR are strong miotics and, at least under some circumstances, can act as effective ocular hypertensives, but these peptides do not reproduce any other signs of ocular irritation or inflammation. It is therefore concluded that none of the peptides studied here could, by itself, be the sole mediator of the initial ocular irritative response although some of them may account for the miosis and contribute to the ocular hypertension associated with this response. A combination of some of these peptides together with PGE₂ and/or other PGs may account for all aspects of the ocular irritative response and for most aspects of the ocular inflammatory response.     

Effects of delta9-tetrahydrocannabinol on ocular blood flow and aqueous humor formation

Blood flow, using a microsphere method, and aqueous humor formation, using an inulin dilution technique, have been measured in control and Δ⁹-tetrahydrocannabinol (Δ⁹-THC) treated animals. The dose level of drug employed was chosen to represent that used in many previous experiments. Blood flow through the iris, ciliary processes and choroid was increased by Δ⁹-THC and aqueous humor formation decreased: pseudofacility was unchanged. The increased blood flow occurs due to a dilation of vessels leading a way from the anterior uvea which consequently reduces the ultrafiltration pressure for aqueous humor formation. THC acts to reduce intraocular pressure by altering the hemodynamic relationships of the anterior uvea, and by increasing true outflow facility.     

Vasoactive intestinal polypeptide-like immunoreactive nerves to the human eye

Using immunohistochemical techniques, vasoactive intestinal polypeptide (VIP) is visualized in nerves distributed to the human eye. Immunoreactive nerve fibers occur about limbal blood vessels and within the trabecular meshwork. In the iris, free-running stromal nerves are the most common, but nerves to both dilator and sphincter muscles are present as well. Immunoreactive nerves are seen within the ciliary muscle and occasionally within a ciliary process. Innervation to choroidal blood vessels constitutes a prominent feature; innervation to more anterior uveal blood vessels is seen only irregularly. Immunoreactive to more anterior uveal blood vessels is seen only irregularly. Immunoreactive nerves are apposed to melanocytes throughout the uvea. The present findings extend prior reports in the human eye, indicating a potential role for VIP in ocular physiology. Additional neuroanatomical, biochemical and physiological studies are necessary to define fully the ocular function of VIP and to determine ultimely whether VIP has clinical and pharmacological implications.     

The effects of blepharorrhaphy induced depression of corneal cholinergic activity

Unilateral lid closure for 8 or more days had previously been shown to reduce rabbit corneal epithelial choline acetyltransferase (ChAc) activity without affecting lactic dehydrogenase activity or thymidine, uridine, leucine and alanine incorporation rates. In the present study, the reduction in ChAc activity was found to be associated with a similar reduction in acetylcholine (ACh) content, to a mean value of 16 and 17%, respectively, of control eye levels. However, on reopening the lids, ACh levels recovered much more rapidly, achieving a mean value 67% of control values (P > 0·05) in 48 hr while ChAc activity was only 23% of control eye levels at 48 hr and required more than 30 days to fully recover.Unilateral lid closure of 10 days was also associated with a small increase in corneal thickness, 0·46±0·09 mm vs. 0·41±0·04 mm (P < 0·05), and reductions in standing electrical potential, 5·5±7·8 mV vs. 13·2±15·2 mV (P < 0·05), and short circuit current, 2·7±2·6 μA vs. 4·9±3·5 μA (P < 0·05). On re-opening the lids, the short-circuit current recovered within 24 hr, the corneal thickness within 3 days and the resting potential within 4 days. Administration of ACh 10^?4m, pilocarpine 10^?4m, eserine 10^?6m or carbachol 10^?5m to the epithelial side of corneas mounted in chambers immediately after reopening the lids failed to elevate either the standing electrical potentials or short-circuit currents; a causal relationship between cholinergic activity and corneal electrical phenomena could not be demonstrated.     

Subbasal nerve morphology, corneal sensation, and tear film evaluation after refractive femtosecond laser lenticule extraction

Background

The purpose of this study was to compare corneal subbasal nerve morphology, corneal sensation, and tear film parameters after femtosecond lenticule extraction (FLEX) and small-incision lenticule extraction (SMILE).

Methods

A prospective, randomized, single-masked, paired-eye design clinical trial of 35 patients treated for moderate to high myopia with FLEX in one eye and SMILE in the other. In both techniques, an intrastromal lenticule was cut by a femtosecond laser and manually extracted. In FLEX, a LASIK-like flap allowed removal of the lenticule, whereas in SMILE, it was removed through a small incision. In-vivo confocal microscopy was used to acquire images of the central corneal subbasal nerve plexus, from which nerve density, total nerve number, and nerve tortuosity were analyzed. Corneal sensation was measured using Cochet–Bonnet esthesiometry. A visual analog scale, tear osmolarity, non-invasive tear film break-up time (keratograph) tear meniscus height (anterior segment OCT), Schirmer's test, and fluorescein tear film break-up time were used to evaluate tear film and ocular surface symptoms. Patients were examined before and 6 months after surgery.

Results

There were no statistically significant differences in baseline parameters between FLEX and SMILE (p?>?0.050). With regard to changes from before to 6 months after surgery, mean reduction in subbasal nerve density was 14.22?±?6.24 mm/mm² in FLEX eyes, and 9.21?±?7.80 mm/mm² in SMILE eyes (p?<?0.05). The total number of nerves decreased more in FLEX eyes than in SMILE eyes (p?<?0.05). No change was found when comparing tortuosity (p?>?0.05). Corneal sensation was reduced with 0.38?±?0.49 cm in FLEX eyes, and 0.10?±?0.34 cm in SMILE eyes (p?<?0.01). No differences were found between FLEX and SMILE in tear film evaluation tests (p?>?0.05). Significantly more patients felt postoperative foreign body sensation in the FLEX eye within the first days after surgery, as compared to the SMILE eye.

Conclusions

Six months after surgery, the less invasive SMILE technique seemed better at sparing the central corneal nerves as compared to FLEX. Corneal sensation was only significantly reduced in FLEX eyes. There were no differences between FLEX and SMILE when comparing tear film evaluation tests 6 months after surgery.     

Glucose transport in isolated mammalian pigment epithelium

The isolated retinal pigment epithelium-choroid of sheep was studied in vitro as a single membrane mounted in a Ussing chamber. The average potential of 40 tissue preparations was 6·3 mV. In all preparations the retina (apical) side was positive with respect to the choroid (basal) side. Changes in Na⁺, K⁺, or Ca²⁺ concentration on the choroid side did not affect the tissue potential, whereas a ten-fold increase in K⁺ concentration or Ca²⁺ chelation by EGTA on the retina side decreased the potential. The potential was also decreased by 0·75 mm-phloridzin and 1·0 mm-dinitrophenol. Utilizing a glucose concentration gradient of 4 mm-choroid to 1 mm-retina side, the unidirectional glucose fluxes were found to be 883 nmol/cm² · hr choroid to retina and 222 nmol/cm² · hr retina to choroid. Both unidirectional fluxes were decreased by 65% Na⁺ replacement with choline Cl. The retina to choroid flux was also decreased by 0·75 mm-phloridzin and 1·0 mm-dinitrophenol. The sodium dependence and inhibitor effects suggest that mechanisms of glucose transport other than simple diffusion are operative in sheep retinal pigment epithelium.     

Enzymes metabolizing ornithine-proline pathway in the rovine eye

We studied bovine ocular tissues biochemically for the following enzyme activities related to ornithine metabolism: arginase, Δ¹-pyrroline-5-carboxylate reductase, Δ¹-pyrroline-5-carboxylate dehydrogenase and proline oxidase. Arginase activity was high in the retina and uvea; Δ¹-pyrroline-5-carboxylate reductase activity was high in the lens, cornea and retina; Δ¹-pyrroline-5-carboxylate dehydrogenase activity was high in the uvea but lower in the retina; and proline oxidase activity was negligible in all ocular tissues. It is possible that ornithine in the retina may be supplied from arginine, or from the uptake of ornithine itself, and converted into proline by the cooperative action of ornithine-ketoacid-transaminase and Δ¹-pyrroline-5-carboxylate reductase.     

The effect of colchicine on the rate of formation of aqueous humour and the gross outflow facility in the albino rabbit

Colchicine, a naturally occurring plant alkaloid which prevents the polymerisation of cytoplasmic microtubules, lowers the intraocular pressure after topical administration or intravitreal injection. In this study wer have examined the effect of topically administered colchicine on the rate of formation of aqueous humour and the gross outflow facility in the albino rabbit eye. The disappearance of [¹⁴C]inulin from the anterior chamber was measured to calculate the rate of aqueous humour formation and in a separate group of animals the gross outflow facility determined using a constant pressure perfusion technique.Topically administered colchicine (10, 20 and 40 μg/eye) inhibited the rate of aqueous humour formation dose-dependently. The mean rate of formation in control eyes was 3·68 ± 0·09 μl/min (n = 16) decreasing to 1·8 μl/min (n = 9) in eyes treated with 40 μg colchicine. Furthermore, the gross outflow facility of colchicine treated (10 μg/eye) eyes (0·49 ± 0·03 μl/min/mmHg) was significantly greater (P < 0·05) than that of contralateral control eyes (0·39 ± 0·03 μl/min/mmHg).The pharmacological evidence available indicates that colchicine is acting via mechanisms of microtubule disruption to produce an ocular hypotensive response, suggesting that microtubules may be involved in the formation of aqueous humour and possibly the maintainance of cell shape and form in the outflow vessels of the anterior chamber.