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We have measured kinetic energy distributions (KEDs) of large clusters emitted from five different solid targets following a single impact of C 60 − ion at 14 keV kinetic energy. It was found that all the large clusters emitted from a given target move with nearly the same velocity and that their KEDs can be described by a thermal distribution riding on a common center-of-mass velocity (shifted Maxwellian) of some precursor. This behavior is in sharp contrast to that observed when the incoming projectile ion is monoatomic. Different trends were observed when comparing the behavior of the KED families of group 5 early transition metal elements (Ta and Nb) with those of group 11 late transition metals (Cu, Ag, and Au). We propose a model for the initial phase of formation of the precursor and show that the measured KEDs can serve as both pressure and temperature probes for the impact excited, highly energized subsurface nanovolume, driving the ejection of the clusters. It is also shown that under the proposed impact scenario, thermally equilibrated conditions (of the atomic subsystem) can be established at the subsurface nanovolume on the early subpicosecond time scale relevant for the emission process. This conclusion is demonstrated both experimentally by the KEDs of the emitted large clusters (very high temperatures and center-of-mass velocity) and by molecular dynamics simulation of the temporal evolution of the thermal characteristics of the impact energized subsurface nanovolume. [ABSTRACT FROM AUTHOR]

We report the experimental observation and computational analysis of the binary tin-carbon gas phase species. These novel ionic compounds are generated by impact of C- 60 anions on a clean tin target at some kiloelectronvolts kinetic energies. Positive SnmCn+ (m = 1-12, 1 ≤ n ≤ 8) ions were detected mass spectrometrically following ejection from the surface. Impact induced shattering of the C- 60 ion followed by sub-surface penetration of the resulting atomic carbon flux forces efficient mixing between target and projectile atoms even though the two elements (Sn/C) are completely immiscible in the bulk. This approach of C- 60 ion beam induced synthesis can be considered as an effective way for producing novel metal-carbon species of the so-called non-carbide forming elements, thus exploring the possible onset of molecular level miscibility in these systems. Sn2C2+ was found to be the most abundant carbide cluster ion. Its instantaneous formation kinetics and its measured kinetic energy distribution while exiting the surface demonstrate a single impact formation/emission event (on the sub-ps time scale). Optimal geometries were calculated for both neutral and positively charged species using Born-Oppenheimer molecular dynamics for identifying global minima, followed by density functional theory (DFT) structure optimization and energy calculations at the coupled cluster singles, doubles and perturbative triples [CCSD(T)] level. The calculated structures reflect two distinct binding tendencies. The carbon rich species exhibit polyynic/cummulenic nature (tin end capped carbon chains) while the more stoichiometrically balanced species have larger contributions of metal-metal bonding, sometimes resulting in distinct tin and carbon moieties attached to each other (segregated structures). The Sn2Cn (n = 3-8) and Sn2Cn+ (n = 2-8) are polyynic/cummulenic while all neutral SnmCn structures (m = 3-4) could be described as small tin clusters (dimer, trimer, and tetramer, correspondingly) attached to a nearly linear carbon chain. For example, the 1:1 (Sn:C) Sn3C3 and Sn4C4 clusters are composed of all-tin triangle and rhombus, correspondingly, with a short carbon chain (C3, C4) attached on top. The cationic Sn3Cn+ (n = 1-5) and Sn4Cn+ (n = 1-4) species exhibit various intermediate geometries. Structure calculations at the CCSD(T) level are essential since the segregation effect is not as easily evident based on the most stable structures calculated by DFT alone. Dependences of bond energies (per atom) reflect the evolution of the segregation effect. The mass spectral abundances could be reasonably rationalized in terms of calculated stabilities of the cluster ions with respect to various dissociation channels. [ABSTRACT FROM AUTHOR]

Acetylcholinesterase (AChE) expresses in non-cholinergic cells, but its role(s) there remain unknown. We have previously attributed a pro-apoptotic role for AChE in stressed retinal photoreceptors, though by unknown mechanism. Here, we examined its promoter only to find that it includes a binding sequence for the activating transcription factor 3 (ATF3); a prototypical mediator of apoptosis. This suggests that expression of AChE could be regulated by ATF3 in the retina. Indeed, ATF3 binds the AChE-promoter to down-regulate its expressions in vitro . Strikingly, retinas of "blinded" mice display hallmarks of apoptosis, almost exclusively in the outer nuclear layer (ONL); coinciding with elevated levels of AChE and absence of ATF3. A mirror image is observed in the inner nuclear layer (INL), namely prominent levels of ATF3 and lack of AChE as well as lack of apoptosis. We conclude that segregated patterns of expressions of ATF3 reflect its ability to repress apoptosis in different layers of the retina-a novel mechanism behind apoptosis.

Purpose: To assess the retinal toxicity of an intravitreal injection of infliximab, a monoclonal antibody to tumor necrosis factor α, in a rabbit model., Materials and Methods: Two groups of adult albino rabbits (n = 5) received intravitreal injections of infliximab (0.1 ml) in the study eye and balanced salt solution (BSS, 0.1 ml) in the control eye at baseline. Group 1 was administered with 1.5 mg/0.1 ml, and group 2 was injected with 7.5 mg/0.1 ml of infliximab solution. Electroretinography (ERG) was performed at baseline and at 1, 7, 30, and 45 days after the injection. Visual evoked potentials (VEPs) were recorded at 7 and 45 days after the injection. After the last electrophysiological assessment, the rabbits were euthanized and retinal histopathology and immunhistochemistry for glial fibrillary acidic protein (GFAP) were performed., Results: ERG responses demonstrated no significant deficit in retinal function in eyes injected with infliximab. Mean dark-adapted a-wave and b-wave maximal amplitude and semi-saturation constant values at baseline and throughout the 45 days of follow-up after the injection indicated no remarkable difference in outer retinal function between the control and experimental eyes. VEP responses were similar at each time point (7 and 45 days). No difference was seen in retinal histopathology and immunocytochemistry sections in eyes receiving the 1.5 mg/0.1 ml dose compared to the control eyes. However, increased GFAP labeling in retinal Müller cells was detected in rabbit eyes treated with the 7.5 mg/0.1 ml dose., Conclusions: Intravitreal injection of 1.5 mg/0.1 ml infliximab dose has no toxic effect on the integrity (functional or structural) of the retina in rabbits. A higher dose of 7.5 mg/0.1 ml may be slightly toxic as suggested by positive Müller cell GFAP expression. Additional studies of retinal toxicity at higher doses and after multiple injections are needed to establish the retinal safety of intravitreal infliximab therapy in humans.

Purpose: The purpose of the study was to assess potential interactions of light exposure and hyperglycemia upon ocular complications in diabetic rats., Methods: Streptozotocin-induced (STZ-induced) diabetic rats ( N = 39) and non-diabetic rats ( N = 9) were distributed into eight groups according to the irradiance and color of the light phase during the 12/12-hour light/dark regime. Follow-up lasted 90 days and included assessment of cataract development and electroretinogram (ERG) recordings. Stress to the retina was also assessed by glial fibrillary acidic protein immunocytochemistry., Results: Cataract development was fast in diabetic rats that were exposed to unattenuated white light or to bright colored lights during the light phase. Diabetic rats that were kept under attenuated brown or yellow light during the light phase exhibited slower rate of cataract development. Electroretinogram responses indicated very severe retinal damage in diabetic rats kept under bright colored lights in the blue-yellow range or bright white light during the light phase. Electroretinogram damage was milder in rats kept under bright red light or attenuated yellow or brown light during the light phase. Glial fibrillary acidic protein expression in retinal Müller cells was consistent with ERG assessment of retinal damage., Conclusions: Attenuating white light and filtering out short wavelengths have a protective effect on the eyes of diabetic rats as evident by slower rate of cataract formation and a smaller degree of retinal damage., Translational Relevance: Our findings suggest that special glasses attenuating light exposure and filtering out short wavelengths (400-530 nm) may be beneficial for diabetic patients.

We report the observation and structural analysis of novel indium carbide gas phase cluster ions generated by bombardment of a clean indium surface by keV C60(–) ions. Positive In(m)C(n)(+) (m = 1–21, 1 ≤ n ≤ 9) ions were ejected off the surface and analyzed mass spectrometrically. C60(–) ion beam irradiation is shown to be an efficient way of producing new kinds of gas phase carbide ions with relatively balanced stoichiometries. The rise kinetics of the ion signal (immediate jump within the beam opening time to a plateau value) indicates that the formation/ejection of the carbide ions constitute a single impact event. In3C2(+) was found to be the most abundant carbide cluster ion. Optimal geometries of the different clusters were derived via density functional theory calculations. The acetylenic/cumulenic nature of the impact emitted cluster ions is manifested by the high abundance of In2C2(+), In3C2(+), and the calculated structures for In(m)C(n)(+) (m = 3–4, n = 2–8). Odd/even intensity alternations in the In3C(n)(+) (n = 1–8) and In4C(n)(+) (n = 1–9) abundances are observed and rationalized by the calculations.

Purpose: To evaluate retinal toxicity of intravitreal rituximab., Methods: Twelve albino rabbits were included in the study. 1 mg/0.1 mL of rituximab was injected to the right (experimental) eye of each rabbit, and 0.1 mL of saline was injected into the left (control) eye. Electroretinogram was recorded before injection and 3 hours, 4 days, 1 week, 2 weeks and 4 weeks after injection (12 rabbits), and visual evoked potential was recorded before injection and 4 weeks after injection (10 rabbits). Histology and glial fibrillary acidic protein immunocytochemistry (12 rabbits) were performed at 4 weeks after injection. Clinical examination was conducted at all time points in all rabbits., Results: The average dark-adapted electroretinogram b-wave Vmax ratios, and the average light-adapted b-wave amplitude ratios were approximately 1, and the average log σ difference was around zero throughout the follow-up period. The average visual evoked potential amplitude ratio and the average visual evoked potential implicit time difference were approximately 1 and 0, respectively. No histologic damage was seen, but glial fibrillary acidic protein was mildly expressed in 6 of 12 experimental eyes. Results of clinical examination were normal in all the eyes., Conclusion: Intravitreal injection of 1 mg rituximab does not cause functional or histologic signs of retinal toxicity in albino rabbits. Mild glial fibrillary acidic protein expression in Müller cells probably indicates a mild degree of retinal stress.

Ultrasound waves, widely used as a non-invasive diagnostic modality, were recently shown to stimulate neuronal activity. Functionally meaningful stimulation, as is required in order to form a unified percept, requires the dynamic generation of simultaneous stimulation patterns. In this paper, we examine the general feasibility and properties of an acoustic retinal prosthesis, a new vision restoration strategy that will combine ultrasonic neuro-stimulation and ultrasonic field sculpting technology towards non-invasive artificial stimulation of surviving neurons in a degenerating retina. We explain the conceptual framework for such a device, study its feasibility in an in vivo ultrasonic retinal stimulation study and discuss the associated design considerations and tradeoffs. Finally, we simulate and experimentally validate a new holographic method--the angular spectrum-GSW--for efficient generation of uniform and accurate continuous ultrasound patterns. This method provides a powerful, flexible solution to the problem of projecting complex acoustic images onto structures like the retina.

Purpose: Intracameral cefuroxime was found to lower the risk of endophthalmitis after cataract surgery. The purpose of this study was to evaluate the retinal toxicity of cefuroxime in a rabbit model., Methods: Twenty-two albino rabbits were divided into two cefuroxime groups: low-dose (1mg/0.1 mL, n = 9) and high dose (10 mg/0.1 mL, n = 13). The right eye of each rabbit was injected with 0.1 mL cefuroxime solution (experimental eye) and the left eye with 0.1 mL saline (control eye). Electroretinogram (ERG) responses were recorded at 3 hours, 4 days, and 1, 2, and 4 weeks after injection. After 4 weeks, the rabbits were euthanized, the eyes were enucleated, and the retinas were prepared for histologic evaluation and GFAP immunostaining., Results: No functional (ERG) or histologic damage was found in rabbits in the low-dose group. In the high-dose group, a significant decrease in the ERG amplitudes of the experimental eyes was seen 3 hours after injection, followed by partial recovery during 4 weeks of follow-up. Retinal histology of experimental eyes revealed marked damage. GFAP immunoreactivity in Müller cells was expressed in rabbits belonging to both groups, although it was more extensive in the high-dose group., Conclusions: ERG and histologic findings indicated that a dose of 1 mg cefuroxime, administered intravitreally, was not toxic to the rabbit retina. A dose of 10 mg, injected intravitreally, induced transient physiological effects, and was toxic to the rabbit retina, as was evident by the permanent reduction in the ERG responses and by the structural damage to the retina with signs of glial activation.

Purpose: The purpose of this was to study the long-term effects of a single intravitreal dose of bevacizumab injected at different postnatal age on retinal function and development of retinal blood vessels in the adult albino rabbit., Methods: Bevacizumab was injected into the right eye of newborn rabbits, aged 11 days to 25 days, whereas the left eye of each rabbit was injected with identical volume of saline and served as control. The electroretinogram was recorded 1 week and 10 weeks after injection. Visual evoked potentials were recorded 10 weeks after injection. At termination of the follow-up period, the rabbits were killed and the retinas were prepared for histopathologic studies at the light microscopic level, for glial fibrillary acidic protein immunoreactivity, and for reduced form of nicotinamide adenine dinucleotide phosphate diaphorase histochemistry to assess the integrity of the retinal vascular system., Results: The electroretinogram responses demonstrated normal retinal function in adult rabbits injected at postnatal age of 11 days to 25 days. Mean Vmax and σ values calculated for each group of rabbits, 10 weeks after bevacizumab injection, indicated similar retinal function of the experimental and control eyes. Visual evoked potentials recorded by stimulating each eye separately were also similar. The histopathologic studies yielded similar results; no structural retinal damage was observed in the experimental eyes compared with the control eyes of rabbits from all age groups, and no increased glial fibrillary acidic protein immunoreactivity in Müller cells was observed in the experimental eyes. Staining of blood vessels with reduced form of nicotinamide adenine dinucleotide phosphate diaphorase revealed decreased branching of the capillary network in the experimental eyes compared with the control eyes in all age groups., Conclusion: The electroretinographic and morphologic data showed no deleterious effects of a single intravitreal dose of bevacizumab, injected during the first 30 days postnatally, on the structural and functional integrity of the sensory retina in the adult rabbit. Even partial blockage of vascular endothelial growth factor with bevacizumab applied during retinal development seems to interfere with the development of the retinal vascular system in the albino rabbit. However, extrapolation from rabbits to humans should be made with caution.

Purpose: To determine whether intravitreal injection of trypan blue is toxic to the retina of the albino rabbit., Methods: Sixteen albino rabbits were studied for the effects of intravitreal trypan blue (eight with 0.06% solution and eight with 0.15% solution). Saline was injected into the fellow control eye of all rabbits. The electroretinogram and visual evoked potentials were recorded from each rabbit at different time intervals after injection. The rabbits were killed at the termination of the follow-up period, and their retinas were prepared for histologic examination under light microscopy., Results: In all rabbits, short-term follow-up showed significant reduction of ERG responses in the experimental eye, with the b-wave more affected than the a-wave. Partial to complete recovery was observed during follow-up. After 4 weeks, negligible ERG deficit was observed in the rabbits treated with 0.06% trypan blue, whereas significant ERG deficit was measured in rabbits tested by the 0.15% trypan blue. No differences in flash VEP responses between experimental and control rabbit eyes were found. Light microscopy showed no significant histologic effects in the retinas exposed to the 0.06% solution. Marked disorganization of all retinal layers was observed in areas close to the site of injection in the rabbits injected with the 0.15% solution., Conclusions: Trypan blue exerts transient physiological effects on the distal retina of the rabbit, but in concentrations of 0.15% it can induce permanent toxic effects. Therefore, caution should be used when using this dye in vitreoretinal surgery.

Purpose: Repeated intravitreal injections of ranibizumab or bevacizumab are a common treatment for several retinal diseases. The aim of the present study was to evaluate the long-term retinal toxicity of repeated injections of ranibizumab and bevacizumab in rabbits., Methods: Albino rabbits were injected intravitreally with ranibizumab (1 mg/0.1 mL) or bevacizumab (2.5 mg/0.1 mL) into the right eye, whereas the left eye of each rabbit was injected with saline. Nine consecutive injections were administered at 14-day intervals. Electroretinographic responses and flash visual-evoked potentials were recorded periodically. After 18 weeks of follow-up, the rabbits were killed, and the retinas were prepared for morphologic examination and for immunocytochemistry for glial fibrillary acidic protein., Results: Electroretinographic and visual-evoked potential responses of the experimental and control eyes were similar in amplitude and pattern throughout the follow-up period. The histopathologic studies yielded similar results. No retinal damage was observed in the experimental and control eyes of all rabbits. Glial fibrillary acidic protein immunoreactivity showed staining only in retinal astrocytes but not in Müller cells in all rabbits., Conclusion: The electrophysiological tests and the morphologic data indicate that the repeated intravitreal injections of ranibizumab or bevacizumab have no cumulative long-term toxic effect on the retina in rabbits.

Gentamicin retinal toxicity was tested in numerous studies on adult animal models, but not in the developing retina. Since differentiating cells and developing tissues may exhibit different degrees of sensitivity to toxic drugs, we aimed here to test the susceptibility of the developing mammalian retina to the toxic action of gentamicin. Gentamicin was injected in the right eye of newborn rabbits, aged 11-38 days. The left eye of each rabbit was injected with saline. Eight weeks after injection, electroretinographic (ERG) responses were recorded in the dark- and light-adapted states. The rabbits were then sacrificed, and the retinas were prepared for morphological examination and immunostaining for Glial Fibrillary Acidic Protein (GFAP) and Protein Kinase C (PKC). The ERG responses demonstrated practically no functional damage to the retina of rabbits injected at postnatal age of 11 and 13 days, and a gradually increasing severity of functional damage with increasing postnatal age of intravitreal gentamicin injection. The histopathological studies yielded similar results. GFAP immunoreactivity showed staining in Müller cells only in retinas that exhibited functional and structural damage. PKC immunoreactivity indicated lesser damage to the rod-bipolar cells compared to the photoreceptors. The ERG data and morphological observations suggest that processes involved in development of the rabbit retina, such as outer segment elongation may provide partial or even complete protection to the retina from toxic insults such as a single dose of gentamicin.

Purpose: To evaluate possible toxicity of intravitreal Kenalog (commercial triamcinolone acetonide) to the retina of albino rabbits., Methods: Forty-three albino rabbits were injected intravitreally with 0.1 mL of experimental solution to the right eye and 0.1 mL of saline to the left eye (control). Rabbits in Group A (n=28) were injected with 4 mg/0.1 mL of Kenalog suspension; rabbits in Group B (n=8) were injected with 0.1 mL of Kenalog vehicle; and rabbits in Group C (n=7) were injected with 4 mg/0.1 mL of triamcinolone acetonide. Rabbits were examined ophthalmoscopically and by electroretinogram (ERG) recordings before and at different time intervals after injection. At the end of follow-up, animals were killed and the retinas were prepared for light microscopy., Results: Thirty-eight rabbits completed 4 weeks of follow-up. Follow-up for 8 and 17 weeks was completed by 29 and 3 rabbits, respectively. Intravitreal commercial Kenalog or its vehicle alone caused approximately 50% reduction in the ERG b-wave amplitude at the end of follow-up. Pure triamcinolone acetonide caused only mild (up to 14%) reduction of the ERG b-wave amplitude. Histologic examination of retinas exposed to Kenalog or its vehicle showed severe damage to all retinal layers in areas close to the site of Kenalog injection., Conclusions: Intravitreal injection of 4 mg Kenalog suspension is retinotoxic to albino rabbit eyes. The vehicle of Kenalog is probably the main cause of this toxicity.

Purpose: To study the involvement of stress-induced acetylcholinesterase (AChE) expression in light-induced retinal damage in albino rats., Methods: Adult albino rats were exposed for 24 hours to bright, damaging light. AChE expression was monitored by in situ hybridization, by histochemistry for AChE activity, and by immunocytochemistry. An orphan antisense agent (Monarsen; Ester Neurosciences, Ltd., Herzlia Pituach, Israel) was administered intraperitoneally to minimize light-induced AChE expression. The electroretinogram (ERG) was recorded to assess retinal function., Results: Twenty-four-hour exposure to bright light caused severe reduction in the ERG responses and augmented expression of mRNA for the "read-through" variant of AChE (AChE-R) in photoreceptor inner segments (IS), bipolar cells, and ganglion cells. AChE activity increased in IS. The expressed AChE protein was a novel variant, characterized by an extended N terminus (N-AChE). Systemic administration of the orphan antisense agent, Monarsen, reduced the photic induction of mRNA for AChE-R, and of the N-AChE protein. Rats exposed to bright, damaging light and treated daily with Monarsen exhibited larger ERG responses, relatively thicker outer nuclear layer (ONL), and more ONL nuclei than did rats exposed to the same damaging light but treated daily with saline., Conclusions: The findings indicate that the photic-induced novel variant of AChE (N-AChE-R) may be causally involved with retinal light damage and suggest the use of RNA targeting for limiting such damage.

Purpose: Intravitreal indocyanine green (ICG) is commonly used in vitreoretinal surgery. The purpose of this study was to evaluate possible toxicity of ICG in the retina of albino rabbits., Methods: Twenty-two albino rabbits were injected intravitreally with 0.1 mL ICG solution in one eye, and three rabbits were studied for the effects of 0.1 mL distilled water. All rabbits were injected intravitreally with 0.1 mL saline into the fellow eye, which served as the control. The electroretinogram (ERG) and visual evoked potential (VEP) were recorded from each rabbit at different time intervals after injection. The rabbits were killed at the termination of the follow-up periods and their retinas prepared for histologic examination at the light microscopic level., Results: Three hours after injection, the ERG responses were reduced in amplitude in all ICG-injected eyes, and the VEPs were of abnormal pattern (reduced amplitude and delayed). Partial dose-dependent recovery was observed during 4 weeks of follow-up. Light microscopy of the retinas of the experimental eyes exhibited considerable damage to all retinal layers in all eyes studied that received the highest ICG dose., Conclusions: ICG is potentially toxic to all retinal layers of the albino rabbit. Although it is difficult to extrapolate these findings directly to human eyes, caution should be exercised when using ICG intravitreally.

Purpose: Intravitreal bevacizumab (Avastin; Genentech Inc., San Francisco, CA) is a new treatment for age-related macular degeneration. The aim of this study was to evaluate retinal penetration and toxicity of bevacizumab., Methods: Ten albino rabbits were injected intravitreally with 0.1 mL (2.5 mg) of Avastin into one eye and 0.1 mL saline into the fellow eye. The electroretinogram (ERG) was recorded after 3 hours, 3 days, and 1, 2, and 4 weeks. The visual evoked potential (VEP) was recorded after 4 weeks. Confocal immunohistochemistry was used to assess retinal penetration., Results: The ERG responses of the control and experimental eyes were similar in amplitude and pattern throughout the follow-up period. The flash VEP responses of the experimental eyes were of normal pattern and amplitude and did not differ from those recorded by stimulation of the control eye alone. Full thickness retinal penetration was present at 24 hours and was essentially absent at 4 weeks., Conclusions: Bevacizumab was found to be nontoxic to the retina of rabbits based on electrophysiologic studies. Full thickness retinal penetration may explain observed clinical effects of intravitreal bevacizumab. Although it is difficult to directly extrapolate to humans, our study supports the safe use of intravitreal bevacizumab injection.

Background: Nitric oxide (NO) plays an important physiological role in inter-cellular communication, but when produced in excess it can become toxic. Our goal was to evaluate possible involvement of NO in the development of retinopathy in diabetic rats., Methods: Diabetes was induced in male albino rats by intravenous injection of streptozotocin. Some of the normal and diabetic rats were raised with added L-arginine to increase in vivo NO synthesis, or with added L-NAME to inhibit the rate of in vivo NO synthesis. NADPH diaphorase histochemistry was conducted on retinal whole mounts and transverse sections at different time intervals after induction of diabetes. The electroretinogram (ERG) was recorded to assess retinal function., Results: After 6 weeks of diabetes, NADPH diaphorase amacrine cells in the diabetic retinas appeared abnormal in their morphology and the degree of staining was decreased in their processes. In contrast, NADPH diaphorase activity was augmented in Müller cells. Supplementing the rats' diet with L-arginine for 10 weeks slightly reduced NADPH diaphorase activity in amacrine cell in normal rats but had no effect on the diabetic rats. Adding L-NAME for 10 weeks did not alter NADPH diaphorase histochemistry in either normal or diabetic rats. The ERG responses were reduced by L-arginine supplementation in normal and diabetic rats, and were unaffected by adding L-NAME to the drinking water., Conclusions: Our findings are consistent with the hypothesis that high glucose levels are deleterious to the rat retina and that excessive synthesis of NO may contribute to the development of diabetic retinopathy.

NADPH diaphorase activity in the rabbit retina is modulated by the state of visual adaptation. In this study, we tested possible glutamatergic control of this phenomenon. Rabbits were injected intravitreally with agonists and antagonists of glutamate. After adaptation (3 hours) to either room light or darkness, the rabbits were killed and the retinae were prepared for NADPH diaphorase histochemistry. Kainic acid significantly reduced the number of NADPH diaphorase amacrine cells but augmented NADPH diaphorase activity in horizontal cells in both light- and dark-adapted animals. 6,7-Dinitroquinoxaline-2,3(1H,4H)-dione exerted no effect on amacrine cells but eliminated NADPH diaphorase activity in horizontal cells. 2-Amino-4-phosphono butyric acid did not affect NADPH diaphorase activity in horizontal cells but reduced the degree of staining in the neuronal processes of amacrine cells. MK-801 and N-methyl-D-aspartic acid (NMDA) had no effect on NADPH diaphorase activity in horizontal cells. However, MK-801 reduced staining in the neuronal processes of amacrine cells but not in their cell bodies. NMDA effects were expressed in a significant reduction in the number and size of amacrine cells that were NADPH diaphorase positive. These results indicate that activation of NADPH diaphorase in horizontal cells by darkness is mediated by the activation of alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA)/kainic acid (KA)-type glutamate receptors. The ON pathway in the retina is probably involved in modulation of NADPH diaphorase in the neuronal processes of amacrine cells. Amacrine cells that are NADPH diaphorase positive contain NMDA-type and AMPA/KA-type receptors and are highly susceptible to NMDA and kainic acid toxicity., (Copyright 2001 Wiley-Liss, Inc.)

Background: Laser lesions may induce retinal damage that is larger than expected from the size of the coagulated area. This study was designed to follow the development of laser-induced reduction in retinal function and to correlate it with structural changes., Methods: Pigmented rabbits were treated in one eye with 225 argon laser lesions. The ERG responses were recorded at different times after treatment. The effect of the laser treatment upon the functional integrity of the retina was assessed from the ERG responses. Structural damage was examined by light microscopy., Results: Shortly (1-2 h) after laser treatment, the ERG responses were reduced by about 50%. ERG deficit continued to develop and reached a maximal level about 24 h after treatment. Thereafter, slow recovery was observed but permanent deficit, relative to the initial laser effect, was seen even 30 days after treatment. Histological observations indicated extensive serous retinal detachment between laser lesions that developed within 24 h after treatment. At 30 days post-treatment, lesioned areas were completely destroyed and heavily pigmented. The retina between lesions was attached to the pigment epithelium but exhibited different degrees of structural damage., Conclusions: The immediate laser damage is confined to the coagulated areas while secondary functional damage develops within 24 h and probably reflects serous retinal detachment between lesions. The serous retinal detachment completely resolves with time but may induce permanent structural abnormalities in non-coagulated retinal areas that is reflected in a functional deficit larger than the initial laser effect.

NADPH diaphorase histochemistry is commonly used to identify cells containing nitric oxide synthase (NOS), the enzyme catalyzing the production of nitric oxide from L-arginine. NADPH diaphorase activity and NOS immunostaining was demonstrated in different cells of the vertebrate retina; photoreceptors, horizontal cells, amacrine cells, ganglion cells, and Müller cells. However, the physiological role of nitric oxide (NO) in the retina has yet to be elucidated. In this study, we tested the assumption that NADPH diaphorase activity in the retinas of rabbits and rats depended on the state of visual adaptation. In the rabbit, light adaptation enhanced NADPH diaphorase activity in amacrine cells and practically eliminated it in horizontal cells. Dark adaptation induced the opposite effects; the NADPH diaphorase activity was reduced in amacrine cells and enhanced in horizontal cells. Retinas from eyes that were injected intravitreally with L-glutamate exhibited a pattern of NADPH diaphorase activity that was similar to that seen in dark-adapted retinas. In rats, the NADPH diaphorase activity of amacrine and horizontal cells exhibited adaptation dependency similar to that of the rabbit retina. But, the most pronounced effect of dark adaptation in the rat's retina was an enhancement of NADPH diaphorase activity in Müller cells, especially of the endfoot region. Assuming that NADPH diaphorase activity is a marker for NOS, these findings suggest that NO production in the mammalian retina is modulated by the level of ambient illumination and support the notion that NO plays a physiological role in the retina.

Purpose: This study was designed to investigate the possibility that gentamicin-induced retinal toxicity is dependent on ocular pigmentation by comparing the effects of the drug on the functional and morphologic integrity of the retina in albino and pigmented rabbits., Methods: In each rabbit, a solution of gentamicin sulfate was injected into the vitreous of one eye, and saline was injected into the other eye. Retinal function was assessed by electroretinogram (ERG) at different time intervals after injection. Retinal structure was examined at the light microscopic level., Results: In albino and pigmented rabbits, functional retinal damage developed to a maximal level within the first week after gentamicin injection. Thereafter, gradual recovery could be seen in eyes that suffered less than 80% maximal reduction in the ERG b-wave. For each dose >0.1 mg studied, retinal damage was more severe in the albino rabbits than in the pigmented ones. The degree of damage was not affected by the level of ambient illumination, nor was it reduced by the administration of N-acetylcystein, a free radical scavenger, together with gentamicin., Conclusions: Ocular pigmentation partially protects the rabbit retina from the toxic action of gentamicin. This protection probably reflects binding of the drug by the melanin, which thereby reduces the concentration of the free gentamicin. When the initial gentamicin-induced retinal damage is expressed in < 80% reduction in the ERG, substantial recovery may occur in both strains of rabbits.

The toxic action of two commercial anesthetics, lidocaine and bupivacaine, on the functional and morphologic integrity of the retina was investigated in albino and pigmented rabbits. The experimental drug was injected into the vitreous of one eye, while saline solution was injected into the fellow eye. Retinal function was assessed from the electroretinogram and the visual evoked potential. Retinal structure was examined at the light microscopic level. Ten milligrams of lidocaine did not affect the electroretinogram and the visual evoked potential responses, though structural damage could be detected close to the site of injection. A lower dose of 5 mg did not produce any detectable physiologic or morphologic damage. The only dose of bupivacaine used, 0.5 mg, was not toxic to the albino and pigmented rabbit retinas, as assessed by the electroretinogram, visual evoked potential, and light microscopy. The results of this study demonstrate that lidocaine and bupivacaine are nontoxic to the rabbit retina at concentrations that are effective for retrobulbar anesthesia.

Purpose: To test the toxic action of two antibiotics, imipenem and aztreonam, on the functional and morphologic integrity of the albino rabbit retina., Methods: Two commercial drugs were used--Tienam, which contains imipenem, and Azactam, which contains aztreonam. Different doses of these drugs were injected intravitreally. Retinal function was assessed from the electroretinogram (ERG) and the visual evoked potential (VEP). Retinal structure was examined at the light microscopic level., Results: Imipenem did not affect the ERG and the VEP responses or the morphology of the retina up to a total injected dose of 0.98 mg (2 mg Tienam). Aztreonam was not toxic to the albino rabbit retina up to a total injected dose of 2.8 mg (5 mg of Azactam). Severe functional and morphologic retinal damage was seen when 10 mg of Azactam was injected. A similar degree of damage was seen when a dose of 5 mg L-arginine, an ingredient of Azactam, was injected into the vitreous., Conclusions: Imipenem and aztreonam are nontoxic to the albino rabbit retina at concentrations that are 500-fold higher than their effective dose against bacterial infection. Azactam is highly toxic at high levels (more than 10 mg injected into the vitreous). Most of the toxicity could be explained by the L-arginine content of the drug.

Purpose: This study was designed to localize the site of action of myristyl gamma-picolinium chloride (MGP) in the rabbit retina and to evaluate the extent of the structural damage induced by the drug., Methods: The structural damage was assessed at the light microscopic level in eyes treated with various concentrations of MGP at different time intervals after intravitreal injection of the drug. Glial fibrillary acidic protein (GFAP) immunoreactivity was tested in the same eyes and served as an index of retinal damage., Results: The rabbit retinas, examined about 1 mo after MGP injection, exhibited loss of photoreceptors and thinning of the retina in the regions close to the site of injection; remote retinal areas appeared morphologically intact or only slightly affected. Immunocytochemical analysis demonstrated the presence of GFAP in Müller (glial) cells throughout the entire retina. When the effects of MGP were examined at short time intervals (24 and 72 hr) after injection, severe morphologic damage in areas adjacent to the site of drug injection developed in parallel with the electroretinographic findings. However, GFAP could not be demonstrated., Conclusions: MGP, the preservative used in Depo-Medrol (Upjohn, Kalamazoo, MI), is highly toxic to the rabbit retina.

Periocular injections of corticosteroids play an important role in the management of various ophthalmologic diseases. The Depo-Medrol vehicle, injected into the vitreous, was shown to be toxic to the lens and to the retina when applied at double strength. The authors examined the effects of Depo-Medrol and one of the components of its vehicle, myristyl-gamma-picolinium chloride (MGP), on the functional integrity of the rabbit visual system. Visual function was assessed objectively from the electroretinogram (ERG) and the visual evoked potential (VEP). The experimental drugs were injected into the vitreous humor of one eye while saline was injected into the fellow eye for control. Depo-Medrol did not produce any measurable effects on the ERG or the VEP. When MGP solutions were injected in concentrations at least twice as large as that in the Depo-Medrol, significant reductions in the light- and dark-adapted ERG responses were seen. The effects of the drug on the ERG responses was seen as early as 3 days postinjection and developed to its maximal level within 1-2 weeks. No ERG recovery was seen over a period of more than 2 months. The VEP, elicited by applying light stimuli to the experimental eye, was characterized by low amplitude and delayed implicit time compared with the response obtained from the control eye.

C-Reactive protein (CRP) is a trace component of normal human serum which has a mol. wt of 105,000 and is composed of five apparently identical subunits arranged in cyclic symmetry. The serum concentration of this protein is known to increase dramatically in response to acute inflammatory or necrotic processes. We report here that in the presence of high concentrations of urea significant antigenic, electrophoretic and binding site modifications of CRP occur selectively in the absence of calcium. CRP treated in this way (designated F-CRP) had a pI of 5.4 and alpha-electrophoretic mobility in contrast to the native molecule which had a pI of 6.4 and gamma-mobility. F-CRP retained a partial antigenic identity to native CRP, displayed decreased intrinsic tryptophan fluorescence, and expressed a new antigenic reactivity. A similar neoantigen was expressed by heating CRP selectively in the absence if calcium (63 degrees C, 5 min). Treatment with guanidinium-HCl or deliberate carbamylation did not produce F-CRP or the expression of the F-antigen. The formation of F-CRP in urea or by heat was prevented by the presence of 0.7 mM or more calcium. CRP treated in this way retained identical characteristics to native CRP. F-CRP chromatographed through Sephadex G-150 in the presence or absence of 6M urea as a protein of apparent mol. wt 75, 000 with no evidence for free CRP subunits. The formation of F-CRP from native CRP resulted in a loss of capacity for calcium-dependent binding to the C-polysaccharide despite the persistence of calcium-independent binding reactivity for polycations. These data suggest that in the presence of sufficient calcium CRP can resist urea- or heat-induced structural denaturation, maintaining antigenic, electrophoretic and binding identity to the native molecule. In the absence of calcium, exposure to urea led to increased electrophoretic mobility and exposure of a new antigenic reactivity, and to alterations in the phosphocholine- but not the polycation-binding sites of the native CRP molecule; this new antigenic reactivity may be of value in further studies on the CRP molecule.

The activity of uridine diphosphoglucose dehydrogenase (UDPGD), a key enzyme in the synthesis of proteoglycans was measured by a quantitative cytochemical method in normal and in osteoarthritic (OA) human cartilage. Normal adult chondrocytes showed low UDPGD activity, which was about half the activity of glucose-6-phosphate dehydrogenase of the same specimens. No significant increase in UDPGD activity was noted in OA chondrocytes. The lack of significantly enhanced UDPGD activity in OA indirectly agrees with studies showing normal 35S uptake in this disease and might explain in part the inability of chondrocytes to cope with continuous proteoglycan loss.