Agonists at the serotonin receptor (5-HT(1A)) protect the retina from severe photo-oxidative stress.

PURPOSE: 5-HT(1A) agonists are neuroprotective in CNS injury models. The authors evaluated the efficacy of 5-HT(1A) agonists to protect the retina from severe blue light-induced photo-oxidative damage. METHODS: Albino rats were dosed (subcutaneously) with AL-8309A, 8-OH DPAT, or buspirone once or three times before 6-hour exposure to blue light. Electroretinograms (ERGs) were measured to assess retinal function, and retinal damage was evaluated by light microscopy. Topical ocular dosing with 1.75% AL-8309B was also evaluated. Rats were dosed with WAY-100635, a 5-HT(1A) antagonist, to determine whether protection required activation of the 5-HT(1A) receptor. RESULTS: ERG response amplitudes were significantly (P < 0.05) depressed more than 66% in vehicle-dosed rats after light exposure. ERGs were significantly higher in rats treated with AL-8309A (0.1-30 mg/kg), 8-OH DPAT (0.1-1 mg/kg), buspirone (5-20 mg/kg) or topical ocular with 1.75% AL-8309B. Retinas from AL-8309A and 8-OH DPAT-treated rats were devoid of histologic lesions. Significant protection was measured in rats dosed once 0, 24, or 48 hours before light exposure. Protection provided by dosing with AL-8309B or 8-OH DPAT was inhibited in rats predosed with WAY-100635. CONCLUSIONS: 5-HT(1A) agonists provided potent and complete functional and structural protection. Protection was inhibited by treatment with WAY-100635, confirming the requirement for activating the 5-HT(1A) receptor in initiating this survival pathway. Single-dose experiments with AL-8309A suggest that the mechanism of protection is rapidly activated and protection persists for 48 hours. AL-8309B (1.75%) was effective after topical ocular dosing. AL-8309B is under evaluation in the clinic and may be useful in treating age-related macular degeneration.

Early manifestations of postnatal hyperoxia on the retinal structure and function of the neonatal rat.

PURPOSE: Postnatal hyperoxia in rats causes an arrest in growth of retinal blood vessels, along with severe changes in retinal ultrastructure and function. Previous studies focused on consequences of postnatal hyperoxia at time points substantially removed from the hyperoxic insult. In this study, the earliest manifestations of this retinopathy were examined. METHODS: Newborn rats were exposed to 80% O(2) from birth to postnatal day 14. The retinas were collected for vascular assessment at postnatal days 6, 9, 12, and 14, and electroretinograms were recorded at postnatal days 15, 16, 17, 19, 24, 30, and 60, after which retinal histology was performed. RESULTS: Hyperoxia significantly attenuated vascular development, especially after 6 and 9 days of exposure which resulted in 64% and 72% of normal coverage, respectively. Vascular growth continued despite hyperoxic exposure, reaching 87% of normal by postnatal day 14. Electroretinograms of hyperoxic rats retained very immature features throughout with nearly abolished b-waves and relatively preserved a-waves. Finally, while retinal structure was virtually complete in the control animals by postnatal day 15, hyperoxic rats always showed a significantly thinner outer plexiform layer (OPL) and lower horizontal cell count (P < 0.05), irrespective of the duration of exposure. CONCLUSION: The findings confirm previous reports of reduced retinal vascular coverage that accompanies the earliest manifestation of postnatal hyperoxia in rats and suggest increased retinal susceptibility to hyperoxia within the first week of life. However, despite the fact that vasculature appears to repair itself, irreversible cytoarchitectural and functional changes occur, the consequences of which are documented immediately after the cessation of hyperoxia.

Structural and functional consequences of trolox C treatment in the rat model of postnatal hyperoxia.

PURPOSE: Previous studies have shown that newborn rats exposed to hyperoxia within the first 2 weeks of life develop vasculopathy in addition to permanent changes in retinal structure and function. It has also been suggested that free radicals may be the source of these pathologic effects. Trolox C, a water-soluble analogue of vitamin E, was previously shown to limit the vascular consequences of exposure to postnatal hyperoxia. The aim of this study was to investigate whether trolox C could also help prevent the functional (electroretinography) and structural (retinal histology) consequences associated with oxygen-induced retinopathy (OIR). METHODS: Newborn albino Sprague-Dawley rats exposed or not exposed to hyperoxia received daily injections of trolox C in doses of 300, 600, and 900 microg/kg (total volume, 50 microL). The effect of treatment was evaluated through electroretinography and retinal histology. RESULTS: Although trolox C tended to have a retinoactive effect on the normal retina, normalization of the hyperoxia-treated group to hyperoxic control and of the normoxia-treated group to normoxic control revealed that the a-wave remained relatively unaffected by hyperoxia exposure and by treatment with trolox C, the efficacy of trolox C at doses of 600 and 900 microg/kg largely outweighed the retinoactive effect, and the oscillatory potentials (OPs) benefited to the greatest extent from trolox C treatment. Furthermore, trolox C was able to limit the reduction in outer plexiform layer thickness but not the concomitant reduction of the horizontal cell count, each of which is associated with OIR. CONCLUSIONS: These results show that, as had been previously demonstrated with retinal vasculature, trolox C limited the retinal functional and structural damages inherent in the rat model of OIR. However, despite treatment, there were still signs (albeit less severe) indicative of OIR. This suggests, as previously advanced, that the pathophysiology of OIR is not solely caused by the action of free radicals or that trolox C is inadequate in treating all aspects of OIR.

Ocular pharmacokinetics of moxifloxacin after topical treatment of animals and humans.

The ocular penetration and pharmacokinetics of moxifloxacin in comparison to other fluoroquinolones (ofloxacin, ciprofloxacin, gatifloxacin, norfloxacin, levofloxacin, and lomefloxacin) have been determined by in vitro and ex vivo techniques, as well as in animal and human studies. This article reviews the original pharmacokinetics work performed by Alcon and other studies reported in the ocular fluoroquinolone literature. The results consistently demonstrate higher maximum concentrations for moxifloxacin relative to the other fluoroquinolones in ocular tissues with levels well above its minimum inhibitory concentrations for relevant ocular pathogens. This superior performance is due to the unique structure of moxifloxacin that combines high lipophilicity for enhanced corneal penetration with high aqueous solubility at physiological pH. The latter property creates a high concentration gradient at the tear film/corneal epithelial interface providing a driving force for better ocular penetration for moxifloxacin. In addition, the higher concentration of moxifloxacin in VIGAMOX (i.e., 0.5% vs. 0.3%) allows more antibiotic to be available to ocular tissues. It is clear from the array of studies summarized in this report that moxifloxacin penetrates ocular tissues better (two- to three-fold) than gatifloxacin, ciprofloxacin, ofloxacin, or levofloxacin. This consistent, enhanced penetration of topical moxifloxacin offers powerful advantages for ophthalmic therapy.

Evaluation of triamcinolone acetonide following intravitreal injection in New Zealand white rabbits.

The safety of intravitreally injected triamcinolone acetonide suspension (TA) was evaluated in rabbits. Each animal received 0.1 ml (1) balanced salt solution (BSS) vehicle, (2) formulation vehicle, (3) 4% TA (4-mg dose), (4) 16% TrAc (16-mg dose) or (5) 25% TA (25-mg dose) as a single intravitreal injection into the right eye. The left eyes served as untreated controls. All animals were observed for 1 month following treatment. In-life evaluations included clinical signs, body weights, slit-lamp biomicroscopic and indirect ophthalmoscopic examinations, intraocular pressure and corneal thickness measurements, and electroretinograms (ERGs). Ocular tissues were harvested following a 1-month post-treatment observation period, fixed, processed, and evaluated by light microscopy. No significant or treatment-related clinical signs were observed for any animals during the study. The opaque white test article was clearly visible in the eye for all TrAc-treated groups, and remained so throughout the study. No statistically significant differences in mean body weights were present between the control and treatment groups, though changes in body weight varied. Corneal thickness was slightly reduced for some treated groups. Intraocular pressures were not statistically significantly different from controls for any treatment group. No significant changes in ERG were evident between treatment groups or from baseline readings. Microscopically, basophilic material (presumed to be drug) was seen in the vitreous of all or most treated eyes, with accumulations in the vitreous or in clumps adjacent to the retinal surface. No pathological changes were observed in the retina or other ocular structures. Triamcinolone acetonide suspension was safe and well tolerated following intravitreal injection in New Zealand white rabbits.

Novel RPGR mutations with distinct retinitis pigmentosa phenotypes in French-Canadian families.

PURPOSE: To characterize the molecular defects in two x-linked retinitis pigmentosa (RP) families. We hypothesized that different RPGR mutations result in distinct RP phenotypes. DESIGN: Observational case series. METHODS: Fifteen members in family I and three members in family II were evaluated. Full ophthalmic evaluations were done. Linkage analyses were performed and likelihood of odds scores (LOD score) were calculated. For mutation analyses, we used dHPLC and automated sequencing. RESULTS: Two novel RPGR mutations were identified in the two families; a Glu 414 (2-bp del) frameshift mutation in family I and an IVS 2-1 (g to a) splice site mutation in family II. All male family members in family I were severely affected by RP but maintained central visual acuities until their 50s and did not develop a bull's eye maculopathy. The female phenotype was highly variable. Some of the carriers exhibited a severe phenotype, one female displayed an asymmetric phenotype, and other carriers were asymptomatic. All members with the RPGR frameshift mutation exhibited rod-cone electroretinograms abnormalities, whereas five members had hearing loss. Male members of family II were severely affected, with early visual acuity loss, central scotomas, and bull's eye maculopathy. The female family members were asymptomatic but displayed cone-rod electroretinograms changes. There was no hearing loss. CONCLUSIONS: Different RPGR mutations lead to distinct RP phenotypes, with a highly variable inter- and intrafamilial phenotypic spectrum of disease that is associated with the type of mutation in RPGR and nonrandom X chromosome inactivation, respectively.

The photopic ERG luminance-response function (photopic hill): method of analysis and clinical application.

With progressively brighter stimuli, the amplitude of the photopic b-wave first increases, briefly saturates and then decreases gradually to reach a plateau, where the amplitude of the b-wave equals that of the a-wave; a phenomenon previously presented as the photopic hill. The unique presentation of this luminance-response function seriously complicates its analysis with curve fitting equations such as that of Naka-Rushton used for scotopic electroretinogram. We report a method of analysis of the photopic hill based on easily identifiable and reproducible features of the ascending and descending limbs of this function. The clinical usefulness of these parameters is illustrated with selected cases of retinal disorders.

Evidence for a brief period of enhanced oxygen susceptibility in the rat model of oxygen-induced retinopathy.

PURPOSE: Findings in a previous study have shown that the retina of newborn rats exposed to hyperoxia during the first days of life sustain permanent functional (as determined with the rod ERG) and structural (as determined with histology) damage that appears to be determined by the level of retinal maturity reached at the time of oxygen exposure-the retinas of rat pups being more susceptible to hyperoxic shock during the second week of life than during the first week. Given that the cone ERG has been shown to mature later than the rod ERG, the purpose of the present study was to examine whether cone responses also demonstrates a similar maturational susceptibility to postnatal hyperoxia. Also examined was whether the oscillatory potentials (OPs) were affected by postnatal hyperoxia. METHODS: Newborn rats were exposed to hyperoxia during selected postnatal day intervals either initiated at birth (early-onset exposure) or at a later postnatal age (late-onset exposure). Photopic and scotopic (mixed cone-rod) electroretinograms were recorded at 30 days. RESULTS: Data analysis reveals that photopic and scotopic responses (b-wave and OPs) demonstrated a similar maturational susceptibility to postnatal hyperoxia, in which exposure regimens initiated during the second week of life were most detrimental to retinal function. The results also revealed a temporal window of enhanced oxygen susceptibility at approximately postnatal day 10. The duration of this window was longer when estimated with the scotopic responses, but the extent of the functional damage was more pronounced when estimated with the photopic signals. Finally, compared with the b-wave, the OPs, especially the short-latency OPs, were proportionally more affected. CONCLUSIONS: The results suggest that cone function is significantly more susceptible to postnatal hyperoxia than rod function, and the OPs appear to be the most susceptible ERG components, thus suggesting a differential susceptibility to oxygen toxicity of the different retinal components. However, despite a clear demonstration of its existence, the exact nature of the temporal window of enhanced oxygen susceptibility as well as a possible equivalence in other animal models of oxygen induced retinopathy, including the human form (retinopathy of prematurity), remains to be determined.

Cone-dominated ERG luminance-response function: the Photopic Hill revisited.

PURPOSE: In response to progressively brighter stimuli, the b-wave of the photopic ERG gradually augments in amplitude, reaches a plateau for a narrow range of intensities and then rapidly decreases with further increments in the luminance of the flash. This unique luminance-response function was originally introduced as the Photopic Hill. The purpose of this study was to further characterize this unique feature of the cone ERG, investigate if it was only limited to b-wave measurements and if it could be obtained under different photopic background luminances. METHODS: Photopic ERGs and oscillatory potentials were generated in response to flashes of light ranging from 0.5 to 16 cd m(-2) s in intensity and presented against photopic backgrounds varying from 18 to 525 cd m(-2) in luminance. RESULTS: All but the brightest background yielded a clear Photopic Hill like luminance-response function which could only be evidenced with the b-wave, the i-wave and OP4 amplitude measurements. Interestingly, the maximal amplitude reached remained almost identical irrespective of the background luminance. CONCLUSIONS: Our results suggest that the retinal mechanisms at the origin of the Photopic Hill effect could represent a voltage limitation mechanism, intimately tied to the OFF pathway. The latter would however be intrinsic to the cone system only and not to the entire retinal network since significantly higher peak amplitudes are reached with dark adaptation.

Visual improvement in Leber congenital amaurosis and the CRX genotype.

PURPOSE: In order to determine genotype-phenotype correlations in Leber congenital amaurosis (LCA), we analyzed the phenotype and genotype of 250 LCA children. We identified a heterozygous CRX mutation in an affected mother and son, and describe the ocular phenotype of the proband from birth through infancy to age 11 years. METHODS: Best-corrected Snellen visual acuities, electroretinograms (ERGs), and Goldmann visual fields were measured, while SSCP and direct sequencing were done for genotyping. RESULTS: The proband had congenital nystagmus, amaurotic, paradoxical pupils, and arteriolar narrowing, without a pigmentary retinopathy. The child had very poor fixation and wandering nystagmus at age 5 months, but had measurable vision at age 6 years. Snellen visual acuities were 20/900 at that time, and slowly improved to 20/150 by age 11 years. Perimetry revealed 60 degrees fields with the V4e target at ages 9 and 10 years, with a new 20 degrees inferior island to the III4e target. ERGs at 5 and 8 months were non-detectable, while the photopic ERGs at age 10 years and again at 11 years showed measurable cone a- and b-waves. At age 47, the phenotype of the affected mother consisted of hand motion vision, a pigmentary retinopathy, and non-detectable visual fields and ERGs. We identified a heterozygous CRX mutation, A177Delta1bp (529delG), in both affected individuals, which is predicted to cause a frameshift and introduces a premature termination codon at position 186. CONCLUSIONS: We report a CRX genotype with an ocular phenotype that consists of spontaneous, marked visual improvement in the proband from birth to age 11 years, which is unlike the previous six reports of LCA patients with the CRX genotype.