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Effects of cholinergic drugs and adrenergic drugs on aqueous humor formation in the rabbit eye.

The aqueous humor formation rate was determined in the anesthetized rabbit with a tracer dilution technique. The anterior chamber was cannulated with two 25-gauge needles: one was inserted into the posterior chamber through the pupil and the other was placed in the anterior chamber near the iridocorneal angle. Fluorescein-dextran solution was infused into the posterior chamber at a constant rate and the aqueous humor was collected through the anterior chamber needle. The aqueous humor formation rate and the volume of dye distribution were calculated simultaneously from the dye dilution curve. Utilizing this technique, the effects of the following substances on aqueous humor formation were studied following topical administration to the conjunctival reservoir: cholinomimetic drugs, pilocarpine and physostigmine; the parasympatholytic drug, atropine; adrenoceptor agonists, norepinephrine, epinephrine, isoproterenol and salbutamol; and beta-adrenoceptor antagonists, timolol and propranolol. Cholinomimetic drugs, i.e., pilocarpine 0.1 mg ml-1 to 10 mg ml-1 and physostigmine 1 mg ml-1, increased the aqueous humor formation rate, while atropine 1 mg ml-1 decreased the rate. The increase in aqueous humor formation by the cholinomimetics was completely antagonized with simultaneous administration of atropine. Norepinephrine 5 mg ml-1 and epinephrine 5 mg ml-1 reduced significantly both the aqueous humor formation rate and intraocular pressure (IOP). Isoproterenol 5 mg ml-1 caused a significant lowering of the blood pressure and IOP and tended to lower aqueous humor formation, although the decrease was not statistically significant. No significant effect on the blood pressure, IOP or aqueous humor formation was observed with isoproterenol 2 mg ml-1. Salbutamol 5 mg ml-1 raised the IOP while timolol 1 mg ml-1 lowered the IOP without any significant change in aqueous humor formation or blood pressure. Timolol 0.5 mg ml-1 or propranolol 5 mg ml-1 did not cause a significant effect on the blood pressure, IOP or aqueous humor formation rate.

Albuterol↗

Does aqueous humor secretion decrease with age?

Aqueous humor flow was calculated during day-time in 148 healthy volunteers and 75 older patients using the Fluorotron Master II anterior chamber protocol (Coherent, Palo Alto, USA). Healthy volunteers as well as patients had no history of ocular pathology, surgery or laser treatment. Slitlamp examination revealed no ocular pathology. Hypertension, diabetes, local and systemic drug therapy, neoplasia, kidney or liver disease, contact lens and ocular trauma were excluded. Mean age of volunteers was 26.5 +/- 3.8 years; age of patients: 65.5 +/- 10.5 years. Aqueous humor flow during day-time in healthy volunteers in the OD: (mean +/- s.d.) 2.26 +/- 1.0 microliters/min, in the OS: 2.17 +/- 1.0 microliters/min, OS: 1.86 +/- 1.1 Ml/min. Correlation coefficient: r = 0.8. The mean aqueous humor flow in the older patients during day-time: OD: 1.91 +/- 1.1 microliters/min. Correlation coefficient: r = 0.54. The Mann-Whitney-U-test revealed a significant difference when comparing the right eyes of healthy volunteers with the right eyes of patients (p < 0.01). When comparing all left eyes the difference is also significant (p = 0.01). The results of the study underline, that the mean aqueous humor secretion does significantly decrease with age. However, the data show that there is only a slight decrease of flow of approximately 2.5% per decade. From the clinical point of view it should be concluded, that although the aqueous humor secretion does decrease with age, this is not of clinical importance, even in cases of glaucoma surgery.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗

Absence of angiogenesis-inhibitory activity in aqueous humor of diabetic rabbits.

We studied inhibitory activity of angiogenesis in the eye to explore the formation of new blood vessels in diabetic microvascular diseases. When we examined the effects of extracts from various ocular tissues of nondiabetic rabbits on the proliferation of bovine aortic endothelial cells, potent inhibitory activity was found in lens and aqueous humor. Aqueous humor inhibited both angiogenesis on chorioallantoic membrane in vivo and promotion of endothelial cell growth in vitro, which were induced by retinal extracts. However, in diabetic rabbits, aqueous humor lost the ability to inhibit the growth of endothelial cells. The loss of activity became evident 1 mo after the alloxan injection and lasted while the animals were hyperglycemic. In addition, diabetic aqueous humor failed to suppress retinal extract-induced angiogenesis promotion and endothelial cell growth. The absence of the inhibitory activity of angiogenesis in diabetic aqueous humor might be involved in promotion of neovascularization in diabetic ocular disease.

Allantois↗

Hyaluronate in human aqueous humor.

Samples of aqueous humor obtained at cataract operations were analyzed for sodium hyaluronate by a recently developed radioassay. The mean value +/- SD of 47 samples was 1.14 +/- 0.46 micrograms/g. The samples from patients with diabetes and/or glaucoma did not differ substantially from the general distribution.

Adolescent↗

Complement levels in normal and inflamed aqueous humor.

C2, C6, and C7 were measured by hemolytic assay and Factor B and IgG were measured by radial immunodiffusion in samples of normal and inflamed aqueous humor. Normal aqueous humor was found to contain functional C2, C6, and C7, but the small ratios of aqueous humor to serum measurements suggested that there was relatively little of these complement components in normal aqueous humor when compared to serum. The mean values of C2, C6, C7, and Factor B in aqueous humor and the median ratios of aqueous humor to serum measurements for each complement component were higher in patients with inflamed aqueous humor than in patients with normal aqueous humor. A comparison of the ratios of IgG to each complement component in normal and inflamed aqueous humor suggested that levels of IgG and complement increased proportionately in inflamed aqueous humor. Factor B, a component of the alternative pathway, was not detected in normal aqueous humor but measured in five of six samples of aqueous humor from eyes with anterior chamber inflammation.

Adult↗

Use of laser flaremetry to measure aqueous humor protein concentration in dogs.

The blood-aqueous barrier in dogs is compromised by uveitis, surgery, and limbal paracentesis. Breakdown of the blood-aqueous barrier allows protein into the aqueous humor and results in mild to severe inflammation. Diagnosis of protein in the aqueous humor is traditionally a subjective measurement. Laser flaremetry was used for noninvasive quantitation of aqueous humor protein concentration in dogs. Flaremetry data were compared with aqueous humor protein concentrations obtained from aqueous humor paracentesis and slit-lamp flare evaluations. Results from clinically normal eyes and those with uveitis and cataracts were compared. Subjective evaluations of flare were correlated with a range of flaremetry readings and aqueous humor protein concentrations. Clinically normal eyes had a range of flaremetry readings of 1.4 to 7.0 photon counts (PC)/ms, with a mean of 3.8 PC/ms. Corresponding aqueous humor protein concentrations ranged from 5 to 28 mg/dl, with a mean of 15.1 mg/dl. Eyes with uveitis or cataracts had a range of aqueous protein concentrations of 13 to 729 mg/dl. Flaremetry readings accurately and sensitively measured total protein concentrations in the aqueous humor of dogs.

Animals↗

Aqueous humor changes after experimental filtering surgery.

We studied aqueous humor of rhesus and owl monkeys for its effect on the growth of subconjunctival fibroblasts in tissue culture. Aqueous humor samples obtained before glaucoma surgery inhibited the initiation of growth of fibroblasts. However, postoperative aqueous humor samples supported growth of fibroblasts. The change in aqueous humor physiology lasted for up to two months after glaucoma surgery. Our study indicated that possibly material added to the postoperative aqueous humor inactivates an inhibitor normally present in primary aqueous humor. An alternative explanation would be that primary aqueous humor, in contrast to secondary aqueous humor, lacks sufficient nutrient material to support fibroblast growth in tissue culture.

Animals↗

Elevation of transforming growth factor alpha in cat aqueous humor after corneal endothelial injury.

PURPOSE: To determine if a scrape injury to cat corneal endothelial cells increases the level of mitogenic proteins such as transforming growth factor alpha (TGF alpha) in aqueous humor. METHODS: Aqueous humor of cats was collected at 0, 2, 6, and 24 hours after wounding the endothelium by contact with a cannula tip. Aqueous humor samples collected from sham-wounded cats served as controls. Aqueous humor samples were analyzed for levels of protein, for mitogenic activity using incorporation of tritiated thymidine by cultures of bovine corneal endothelial cells, and for immunoreactive TGF alpha protein using a specific radioimmunoassay. RESULTS: The average protein level in aqueous humor obtained before wounding was low (0.5 mg/ml), increased 26-fold at 2 hours after injury (13 mg/ml), then progressively decreased at 6 hours (8 mg/ml) and 24 hours (2 mg/ml). Levels of mitogenic activity of aqueous humor samples collected 2, 6, and 24 hours after wounding were 2-fold, 2.5-fold, and 0.6-fold higher, respectively, compared to the level of mitogenic activity measured in aqueous humor collected before wounding (0 hours) or in aqueous humor collected from sham-wounded eyes. TGF alpha concentration in aqueous humor collected before endothelial wounding was low (6.8 ng/ml), increased 14-fold 2 hours after wounding (97.4 ng/ml), then progressively decreased at 6 hours (63.3 ng/ml) and 24 hours (35.5 ng/ml) after wounding. TGF alpha concentrations in aqueous humor collected from sham-wounded eyes at 2 hours (9.5 ng/ml) and 6 hours (5.3 ng/ml) were not significantly different from prewound levels. Detergent extracts of bovine corneal endothelial cells contained substantial levels of TGF alpha immunoreactive protein (20 ng/mg protein). CONCLUSIONS: Wounding of cat endothelium causes a rapid increase in mitogenic proteins in aqueous humor including TGF alpha, which may act by an autocrine mechanism to stimulate endothelial wound healing.

Animals↗

A direct correlation between the levels of ascorbic acid and H2O2 in aqueous humor.

There is evidence that H2O2 present in aqueous humor arises from ascorbic acid which is also present in this fluid, but the extent to which peroxide is derived from ascorbic acid is not known. We have measured the concentrations of H2O2 and ascorbic acid normally present in the aqueous humor of various species and also under conditions in which the level of ascorbic acid in the fluid was experimentally altered. In aqueous humor of rabbit and guinea pig the concentration of ascorbic acid was 10 times higher than that present in aqueous of rat and frog. Similarly, the concentration of H2O2 was four to 10 times higher in rabbit and guinea pig aqueous compared to that in rat and frog. Consistent with the higher concentration of ascorbic acid in posterior compared to anterior aqueous humor in the rabbit, the concentration of H2O2 was also significantly higher in the posterior aqueous. When ascorbic acid in rabbit aqueous humor was elevated by intraperitoneal administration of the compound, there was a significant increase in the level of H2O2 in both anterior and posterior aqueous humor. Moreover, when the level of ascorbic acid was lowered experimentally by placing guinea pigs on an ascorbic acid deficient diet, a 10-fold decrease in the level of both ascorbic acid and H2O2 was observed in the aqueous humor. Upon returning the animals to a normal diet, the concentrations of both compounds returned to control values. The direct correlation between the concentrations of ascorbic acid and H2O2 in aqueous humor suggests that ascorbic acid is the primary source of H2O2 in this fluid.

Animals↗

Pharmacodynamics of beta-blocker modulation of aqueous humor production.

A conscious rabbit model with microdialysis sampling of endogenous aqueous humor ascorbate was developed in order to assess the pharmacodynamics of beta-blocker modulation of aqueous humor production. CMA/20 microdialysis probes were implanted in the anterior chamber of each eye of rabbits (n = 6). After a 2 week recovery period, an i.v. bolus of 14C-ascorbate (20 microCi) was administered. Blood samples and aqueous humor microdialysis probe effluent were collected and analysed for endogenous and 14C-ascorbate to estimate the basal rate of ascorbate blood to aqueous humor secretion (Ro). After a 1 hr washout, each rabbit received a series of three doses of 3H-propranolol (750-3000 microg, 16.5 microCi mg(-1)) every 60 min into the lower cul-de-sac of each eye. Probe effluent was analysed for endogenous ascorbate and 3H-propranolol; ascorbate and propranolol in the iris/ciliary body, vitreous and aqueous was determined at the end of the experiment. Nonlinear least-squares regression analysis of the concentration-time profiles for aqueous humor ascorbate was performed to estimate the change in aqueous humor flow. The average basal aqueous humor ascorbate secretion rate was approximately 48/microg hr(-1). Propranolol (1500 microg) produced significant increases in aqueous humor ascorbate, this observation is consistent with a reduction in aqueous humor production (approximately 47%). Analysis of intraocular tissue ascorbate indicated that propranolol inhibited ascorbate secretion at the 3000 microg dose, the highest dose examined in this study; this inhibition was not observed at the 750 microg or 1500 microg doses. Changes in aqueous humor production precipitated by the administration of beta-adrenergic antagonists can be estimated by measuring changes in aqueous humor ascorbate concentrations in the conscious rabbit. Microdialysis sampling of aqueous humor for endogenous ascorbate provides a relevant analytic tool to estimate modulatory effects of anti-glaucoma drugs on aqueous humor production.

Acetazolamide↗

Enzyme-linked immunosorbent assays for the detection of Toxoplasma gondii-specific antibodies and antigens in the aqueous humor of cats.

Serum and aqueous humor samples, collected from 14 clinically normal cats and 96 cats with clinical evidence of intraocular inflammation, were assayed with ELISA for Toxoplasma gondii-specific immunoglobulin M (IgM), T gondii-specific IgG, T gondii-specific antigens, total IgG, and total IgM. Additionally, serum was assayed with ELISA for feline leukemia virus p27 antigen and antibodies against the feline immunodeficiency virus as well as with an immunofluorescent antibody assay for antibodies against feline coronaviruses. Calculation of the Goldmann-Witmer coefficient (C-value) for the T gondii-specific antibodies detected in aqueous humor established the likelihood of local antibody production. Serologic evidence of present or prior infection by an infectious agent was found in 81.9% of the clinically affected cats from which serologic results were available (77/94 cats). Seropositive results for toxoplasmosis were found in 74.0% of the clinically affected cats. Anterior segment inflammation was found in 93.1% (81/87 cats from which information was available) of the clinically affected cats, most of which were older males. Toxoplasma gondii-specific antibodies were not detected in the aqueous humor of 6 seropositive, clinically normal cats. The C-values for aqueous T gondii antibodies were greater than 1 in 44.8% of the cats and greater than 8 in 24.0% of the cats. Response to treatment with clindamycin HCl was positive in 15/20 (75%) of the T gondii-seropositive, clinically affected cats treated with this drug. In 13/15 (86.7%) T gondii-seropositive, clinically affected cats having a C-value greater than 1, response to treatment with clindamycin HCl was positive.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

Effect of desmopressin on aqueous humor flow in humans.

The rate of aqueous humor flow was studied in 17 human subjects with neurogenic diabetes insipidus. Flow was measured by fluorophotometry on two consecutive afternoons. The first day, subjects used desmopressin to control their diuresis; on the second day, subjects did not use desmopressin and had uncontrolled diuresis. On both days, one eye was treated with the beta-adrenergic antagonist, timolol. With desmopressin, the rate of aqueous humor flow in the untreated eye was 2.53 +/- 0.79 microliters/min (mean +/- standard deviation) and in the timolol-treated eye was 1.69 +/- 0.40 microliters/min. Without desmopressin, the rate of aqueous humor flow in the untreated eye was 2.34 +/- 0.69 microliters/min and in the timolol-treated eye was 1.53 +/- 0.43 microliters/min. Thus, the use of desmopressin was associated with a slightly higher rate of aqueous humor flow in both the normal and the beta-adrenergically inhibited eye (P = .05), and the suppression of aqueous humor flow associated with beta-adrenergic inhibition occurred with and without antidiuretic hormone (P < .001). The observed differences in aqueous humor flow on the two days could have been caused by a direct effect on the eye or to indirect effects, such as the change in plasma osmolality, which changed from 291 +/- 6 mOsm on the desmopressin day to 299 +/- 8 mOsm on the desmopressin-abstention day (P < .001).

Administration, Intranasal↗

Effects of pilocarpine, salbutamol, and timolol on aqueous humor formation in cynomolgus monkeys.

The rate of aqueous humor formation was determined in the cynomolgus monkey eyes by a tracer dilution technique. One 25-gauge needle was inserted into the posterior chamber and a solution of fluorescein labeled dextran with a molecular weight of 40,000 was infused at a constant rate. The aqueous humor was collected through a needle inserted into the anterior chamber, while the intraocular pressure (IOP) was maintained at a constant level. The aqueous humor formation rate and the dye distribution volume were calculated from the time profile of the dye concentration in the effluent aqueous humor. By means of this technique, the effects of pilocarpine, salbutamol, and timolol on the aqueous humor formation rate were studied. The test drug solution was administered into the conjunctival reservoir during a 90-min period before measurements and also during the measurement so that a steady-state drug concentration was maintained in the anterior chamber during the measurement. Pilocarpine 0.1% reduced the aqueous humor formation rate to approximately 50% of the control without significantly changing the IOP or the distribution volume. Salbutamol 0.5%, a beta-adrenergic agonist, increased the rate by about 38%, but timolol 0.1%, a beta-adrenergic antagonist, reduced the rate by an average of 36%. Timolol caused a statistically significant lowering of the IOP by about 2 mmHg. Simultaneous administration of salbutamol 0.5% and timolol 0.2% caused no change in the aqueous humor formation rate or the IOP. The barrier function of the blood aqueous barrier was not altered by these drugs as revealed by aqueous protein determinations.

Albuterol↗

Physiological factors in the circadian rhythm of protein concentration in aqueous humor.

PURPOSE: The authors addressed three questions concerning the circadian rhythm of aqueous humor protein concentration in rabbits. First, is there an endogenous oscillator for this circadian rhythm? Second, does a circadian rhythm occur for individual aqueous humor protein components? Third, what is the role of ocular sympathetic nerves, which are more active in the dark phase, in this circadian rhythm? METHODS: Adult New Zealand albino rabbits were entrained to a daily 12-hour light/12-hour dark cycle. Under a constant dark environment for 24 hours, rabbits were killed at 4-hour intervals, beginning at 2 hours before the onset of the subjective light phase. Eight rabbits were used for each of the six time points. Aqueous humor and vitreous humor were collected, and their protein concentrations were determined. Major aqueous humor protein components were resolved by polyacrylamide gel electrophoresis (PAGE), stained with silver reagent, and analyzed using densitometry. Another group of eight light-dark-entrained rabbits underwent unilateral transection of the cervical sympathetic trunk. Three weeks after the operation; the circadian elevation of intraocular pressure (IOP) at 2 hours into the dark phase was determined for both eyes. Rabbits were later killed at this time point, and total protein concentrations in aqueous humor and vitreous humor were determined in both eyes. Major aqueous humor protein components in both eyes were resolved by PAGE and were compared. RESULTS: In light-dark-entrained rabbits, a circadian rhythm of protein concentration appeared in the aqueous humor under a constant dark environment. Total protein concentration in aqueous humor increased sharply in the early subjective light phase, remained relatively high during the remainder of the subjective light phase, and decreased in the subjective dark phase. Analyses of albumin and other abundant proteins in the aqueous humor showed that all of them varied similarly in a circadian pattern. In contrast, total protein concentration in the vitreous humor remained unchanged. In rabbits with unilaterally decentralized ocular sympathetic nerves, total protein concentrations in the aqueous humor and the vitreous humor in the early dark phase showed no difference between the two eyes. In addition, there was no difference in individual aqueous humor protein concentration between the two eyes. However, the nocturnal IOP elevation in the decentralized eye was less than that in the contralateral, intact eye. CONCLUSIONS: The circadian rhythm of aqueous humor protein concentration in rabbits can continue without an external signal of dark-light change, indicating the existence of an endogenous oscillator. A similar circadian rhythm occurs for various major aqueous humor protein components. The nocturnal increase in ocular sympathetic activities plays a limited role in the circadian rhythm of aqueous humor protein concentration.

Animals↗

Identification of unknown quaternary ammonium compounds in corneal epithelium and aqueous humor.

Bovine corneal epithelium and bovine aqueous humor are investigated for their content of quaternary ammonium compounds. In total, four compounds are found. Three of these are identified. For the fourth compound, a proposal for its structure is made on the basis of tandem mass spectrometry fragmentation spectra. The compounds investigated have m/z values of 146, 160, and 174. The compounds with m/z 146 are confirmed as acetylcholine (in corneal epithelium) and (3-carboxypropyl)-trimethylammonium (in both corneal epithelium and aqueous humor). The compound with m/z 174 is identified as butyrylcholine (in corneal epithelium). The compound with m/z 160 is probably acetyl-g-homocholine (in both corneal epithelium and aqueous humor). For both butyrylcholine and acetyl-g-homocholine, it is the first time the presence of these compounds in corneal epithelium or aqueous humor (or both) is described. Both acetylcholine and butyrylcholine are unstable compounds, which are probably susceptible to enzymatic degradation by acetylcholine-esterase and butytrylcholine- esterase, respectively.

Animals↗

Fluorophotometric determination of aqueous humor flow rate in clinically normal dogs.

OBJECTIVE: To determine aqueous humor flow rate in clinically normal dogs, using fluorophotometry. ANIMALS: 20 clinically normal Beagles. PROCEDURE: A study was performed on 5 dogs to establish an optimal protocol for fluorophotometric determination of aqueous humor flow rate. This protocol then was used to measure aqueous humor flow rate in 15 dogs. Corneas were loaded with fluorescein by topical application, and corneal and aqueous humor fluorescein concentrations were measured 5, 6.5, and 8 hours after application. Concentration-versus-time plots were generated, and slopes and ratios of the fluorescein concentration in the cornea and aqueous humor from these graphs were used to calculate flow rates. Calculations were performed by use of automated software provided with the fluorophotometer and by manual computation, and the 2 calculation methods were compared. RESULTS: The protocol established for the 5 dogs resulted in semilogarithmic and parallel decay of corneal and aqueous humor concentrations. Manually calculated mean +/- SD aqueous humor flow rates for left, right, and both eyes were 5.58 +/- 2.42, 4.86 +/- 2.49, and 5.22 +/- 1.87 microl/min, respectively, whereas corresponding flow rates calculated by use of the automated software were 4.54 +/- 3.08, 4.54 +/- 3.10, and 4.54 +/- 2.57 microl/min, respectively. Values for the left eye were significantly different between the 2 computation methods. CONCLUSIONS AND CLINICAL RELEVANCE: Aqueous humor flow rates can be determined in dogs, using fluorophotometry. This technique can be used to assess pathologic states and medical and surgical treatments that alter aqueous humor dynamics.

Animals↗

Polymerase chain reaction for the detection of Toxoplasma gondii in aqueous humor of cats.

OBJECTIVES: To develop Toxoplasma gondii B1 gene polymerase chain reaction (PCR) for use with aqueous humor of cats, and to report PCR and antibody detection results in naturally exposed cats with and without uveitis. SAMPLE POPULATION: Serum and aqueous humor samples from client-owned, healthy cats (n = 23) and client-owned cats with uveitis (n = 43). PROCEDURE: T gondii-specific IgM and IgG were measured in serum and aqueous humor from all cats. The Goldman-Witmer coefficient for ocular antibody production was calculated for cats positive for T gondii-specific IgM or IgG in aqueous humor. Aqueous humor from all cats was assessed by the B1 gene PCR. RESULTS: T gondii was detected in aqueous humor by PCR from 2 of 23 (8.7%) healthy cats and 8 of 43 (18.6%) cats with uveitis. T gondii-specific IgM in either serum or aqueous humor was detected in 5 of 8 (62.5%) cats with uveitis and T gondii in aqueous humor. All cats with uveitis and T gondii in aqueous humor had anterior segment disease. In 5 of 8 (62.5%) cats with uveitis and T gondii in aqueous humor, ocular production of T gondii antibodies was not detected. T gondii was not detected in aqueous humor from 14 of 17 (82.4%) cats with ocular production of T gondii-specific antibody. CONCLUSIONS: The presence of T gondii in aqueous humor may correlate to clinical disease in some, but not all, cats. CLINICAL RELEVANCE: T gondii-specific aqueous humor antibody tests and PCR should be used together to aid in the diagnosis of ocular toxoplasmosis in cats.

Animals↗