Intraretinal processing following photoreceptor rescue by non-retinal cells

Royal College of Surgeon (RCS) rats undergo retinal degeneration due to the inability of retinal pigment epithelial (RPE) cells to phagocytose shed outer segments. We explored the effect of introducing Schwann cells to the subretinal space of RCS rats (before the onset of retinal degeneration), by relying on electroretinogram (ERG) recordings and correlative retinal morphology. Scotopic ERGs recorded from cell-injected eyes showed preserved amplitudes of mixed a-wave b-wave, rod b-waves, and cone b-waves over controls (sham-injected eyes); photopic b-wave amplitudes and critical flicker fusion were also improved. Normal retinal morphology was found in areas of retinas that had received cell injections. Since Schwann cells have no phagocytic properties, their therapeutic effect is best explained through a paracrine mechanism (secretion of factors that ensure photoreceptor survival).     

正常SD大鼠视网膜电图随生长发育变化的特点

目的研究正常SD大鼠视网膜电图(ERG)随生长发育的变化特点。方法分别测量40只(40眼)SD大鼠在出生后第14、21、28、35和56d(P14、P21、P28、P35、P56)的ERC。结果正常SD大鼠Max-ERGa波、OPsO1波潜伏期在各个时间点无明显差异;Rod-ERGb波、Max-ERGb波、Cone-ERGb波、OPsO2波潜伏期和波幅值以及OPsO1波和Max-ERGa波幅值、Flick-ERG幅值在P14分别和其他时间点有差异。结论实验在一定程度上证实了正常SD大鼠ERG中各波形的起源。明确了各波形的成熟时间,P21时正常SD大鼠ERG基本发育成熟。     

Contribution of rod and cone pathways to the dark-adapted electroretinogram (ERG) b-wave following retinal degeneration in RCS rats

Although the RCS rat is widely used as a model of progressive photoreceptor loss, it is unclear how the relative rod and cone functions change with age. Rod and cone b-waves were isolated using a double flash ERG paradigm. In contrast to cones, rods never reached normal functional maturity levels, and the ERG b-wave changed from being predominantly rod-driven to being purely cone-driven by age 74 days, at which point, b-waves were progressively replaced by negative STR-like (scotopic threshold response) waves that persisted up to age 180 days. A double flash commonly abolished this wave and unveiled a b-wave.     

The Effect of Intravitreal <Emphasis Type="Italic">N</Emphasis>-Methyl-<Emphasis Type="SmallCaps">dl</Emphasis>-Aspartic Acid on the Electroretinogram in Royal College of Surgeons Rats

Purpose

To investigate how the third-order neuronal response contributes to shaping the electroretinogram (ERG) in the Royal College of Surgeons (RCS) rat.

Methods

Full-field ERGs were recorded from dystrophic RCS rats (n = 30) at 4, 6, 8, 10, 12, or 14 weeks of age in response to different stimulus intensities (maximum intensity, 0.84?log?cd-s/m²). N-methyl-dl-aspartic acid (NMDA, 5?mM) was injected into the vitreous cavity of the right eyes to eliminate the third-order neuronal response. The left eyes received the vehicle and served as controls. The third-order neuronal response was isolated by digitally subtracting waveforms of the NMDA-injected eyes from those of the control eyes.

Results

The ERG a- and b-waves deteriorated with the age of the rat. The third-order neuronal response was preserved to a greater degree than the b-wave despite progression of photoreceptor degeneration. Intravitreal injection of NMDA attenuated the a-wave and enhanced the b-wave across the stimulus range from low to middle intensities. This tendency became more pronounced with advancing rat age. In aged dystrophic RCS rats this phenomenon was seen even at maximum intensity. The difference between NMDA-injected and vehicle-injected eyes was larger for the threshold than for the maximum amplitude at each examined time point (P < 0.001). Intravitreal injection of NMDA decreased implicit times of the a- and b-waves after the rats reached 8 weeks of age (P < 0.005 for the a-wave).

Conclusion

With advancing photoreceptor degeneration, the third-order neuronal response made a greater contribution to shaping the a- and b-waves in dystrophic RCS rats.?Jpn J Ophthalmol 2007;51:165–174 © Japanese Ophthalmological Society 2007

     

Color sensation of pseudophakic eye from a viewpoint of electrophysiological study]

Monochromatic ERG b-waves were recorded in normal eyes, cataractous eyes and pseudophakic eyes implanted with non-UV or UV IOLs. ERG b-waves were elicited by fourteen monochromatic stimulus lights ranging from 400 to 660 nm under white light adaptation, and spectral response curves were obtained from b-wave amplitudes. Compared with normal eyes or cataractous eyes, the relative b-wave amplitude of pseudophakic eyes was significantly larger in the short wavelength range from blue to green. Except for 400 nm, the spectral response curve of the eyes implanted with UV IOLs was similar to that of eyes implanted with non-UV IOLs. These results suggested that dyschromatopsia might not only occur in eyes implanted with non-UV IOLs but in eyes with UV IOLs.     

老年黄斑变性的闪烁光和明视视网膜电图

对45眼干性型和24眼湿性型老年黄斑变性（AMD）作30Hz刺激频率闪烁光和明视ERG检测，闪烁光ERG波形经傅里叶变换后取基波（30Hz）的相位和幅值作为特征性指标进行观察。结果干性型AMD闪烁光ERG基波相位、幅值，明视红、白光b波幅值的异常率分别为48.89%、8.89%、8.89%及10.11%，异常率间有显著差异（P＜0.01）。视力正常的早期AMD中，基波相位仍有较高的异常率。湿性型AMD的基波相位与幅值的异常分别为62.50%和45.84%.两型AMD基波幅值、明视ERGb波幅值间有显著差异（P＜0.01）.表明幅值异常与病变程度有关，基波相位是检测AMD早期视功能变化的较敏感指标。 （中华眼底病杂志，1996，12：41-43）     

Electroretinographic findings in panretinal photocoagulation for diabetic retinopathy

A total of 16 patients (16 eyes) with type II diabetes mellitus and proliferative retinopathy were studied electroretinographically before panretinal photocoagulation, during intervals between the treatment sittings, within 36 h of the final treatment and 4 months after its conclusion. Treatments were carried out with blue-green argon (eight eyes) and red krypton (eight eyes) lasers; the type of laser was randomly chosen. The analysis of the results obtained showed a significant and marked reduction in peak amplitudes of both a- and b-waves in photopic and dark-adapted conditions as early as between the multiple treatments. Electroretinograph (ERG) tracks did not change significantly at subsequent checks. Amplitude reductions were higher for scotopic b-waves and in eyes that had the less altered baseline tracks. Implicit times did not change significantly. The type of laser used did not significantly influence ERG evolution.     

腺病毒载体介导的色素上皮衍生因子对大鼠视网膜缺血再灌注损伤后视觉电生理的影响

目的:观察重组腺病毒介导的色素上皮衍生因子(Ad-PEDF)对大鼠视网膜缺血再灌注损伤后视网膜电流图的影响。方法:选用健康大鼠24只,随机分为正常组、缺血再灌注组、缺血再灌注+Ad-CMV组,缺血再灌注+Ad-PEDF组,以前房加压的方法制备大鼠视网膜缺血再灌注模型,缺血再灌注+Ad-CMV组,缺血再灌注+Ad-PEDF组分别玻璃体腔注射Ad-CMV或Ad-PEDF1μL(滴度3.8×109/PFU),每组按照时间点12,24,72,168h分为4亚组,以ERG分别测量双眼各时期ERGb波、Ops波波幅。结果:缺血再灌注后ERGb及Ops波幅明显降低,与正常组相比在各时间点有明显的统计学差异,缺血再灌注组ERGb波及Ops波幅24h降低最为明显,与缺血12,72,168h相比差异具有显著性。AD-PEDF能够明显促进ERGb波及Ops波幅恢复,与缺血组、缺血再灌注+AD-CMV在各时间点均有明显统计学差异。结论:腺病毒介导的色素上皮衍生因子玻璃体腔注射能够促进大鼠视网膜缺血再灌注损伤功能恢复。     

Contribution of N-methyl-dl-aspartic acid (NMDA)-sensitive neurons to generating oscillatory potentials in Royal College of Surgeons rats

Purpose

We investigated how the N-methyl-dl-aspartic acid (NMDA) receptor contributes to generating oscillatory potentials (OPs) of the electroretinogram (ERG) in the Royal College of Surgeons (RCS) rat.

Methods

Scotopic ERGs were recorded from dystrophic and wild-type congenic (WT) RCS rats (n = 20 of each) at 25, 30, 35, and 40 days of age. The stimulus intensity was increased from ?2.82 to 0.71 log cd-s/m² to obtain intensity-response function. NMDA was injected into the vitreous cavity of the right eyes. The left eyes were injected with saline as controls. The P3 obtained by a-wave fitting was digitally subtracted from the scotopic ERG to isolate the P2. For the OPs, the P2 was digitally filtered between 65 and 500 Hz. The amplitudes of OP1, OP2, OP3, and OP4 were then measured and summed and designated as ΣOPs. The implicit times of OP1, OP2, and OP3 were also measured. The frequency spectra of the OPs were analyzed using fast Fourier transform (FFT).

Results

The maximum ERG a- and b-waves as well as ΣOPs amplitudes reduced with age in dystrophic rats. Compared with intravitreal saline injection, administration of NMDA decreased ΣOPs amplitudes from 30 days of age in dystrophic rats, while it did not attenuate ΣOPs amplitudes in WT rats. The implicit times of the OPs of the maximum ERG were prolonged by NMDA injections in WT and dystrophic rats. NMDA/saline ratios of ΣOPs amplitudes area under the FFT curves were significantly lower in dystrophic rats from 30 days of age than that in WT rats.

Conclusion

In the early stage of photoreceptor degeneration, intravitreal NMDA injection attenuated OPs amplitudes in dystrophic rats. This indicates that NMDA receptors play a significant role in generating OPs amplitudes with advancing photoreceptor degeneration.     

Quantitative evaluation of electroretinogram before cataract surgery

The ERGs (electroretinogram) of dense mature cataractous eyes were compared with those of the fellow eyes with good visual acuity in 22 senile patients who showed normal preoperative ERG. The mean amplitudes of the a- and b-waves were slightly reduced in the cataractous eyes as compared to those of the fellow eyes, but the difference was not significant. The light-absorbing effect of the cataract based on the a- and b-wave amplitudes was 0.5 log units. Another group of 13 eyes with senile mature cataract showed abnormal preoperative ERG. Chorioretinal atrophy was the most common cause of the subnormal ERG. Postoperative visual acuity was lower in cataractous eyes with subnormal ERGs than in eyes with normal ERGs. Despite the fact that it does not always reflect the macular function and the visual pathway, the ERG still remains a reliable guide in evaluating the visual prognosis before cataract surgery.     

mGluR5对大鼠视网膜对光反应的调控

目的探索代谢型谷氨酸受体5（mGluR5）对大鼠暗适应视网膜电图（ERG）不同成分的贡献。方法实验研究。出生后30日龄大鼠（RCS-rdy+-p+）12只,暗适应大于12 h后,分别行视网膜下腔注射mGluR5激动剂CHPG[（R,S）-2-chloro-5-hydroxyphenylglycine] 200 µmol（6只,CHPG组）和抑制剂MPEP[2-methyl-6-（phenylethynyl）-pyridine] 200 µmol（6只,MPEP组）,对侧眼注射同等体积PBS作为对照。注射后2 d采用RETI-scan系统记录其系列刺激光强度下（-4.5、-2.5、-0.5、-0.02、0.5 1 logcd×m×s-2）的暗适应ERG。导出数据,采用配对t检验进行双眼间数据比较。结果CHPG组,a、b波幅值在不同光强下均显著降低（除最低光强）（-4.5＜t＜-2.3,P＜0.05）,进一步分析得出a、b波最大幅值（Rmax和Vmax）均明显降低（-4.5＜t＜-2.3,P＜0.05）,但其敏感度均未出现明显变化,且b/a未见明显变化。MPEP组,a、b波幅值在不同光强下均显著升高（除最低光强）（-3＜t＜-2,P＜0.05）,进一步分析得出a、b波Rmax和Vmax均明显升高（-3＜t＜-2,P＜0.05）,但其敏感度均未出现明显变化,且b/a未见明显变化。2组的振荡电位（OPs）与相应的对照眼比较未见明显差别。结论mGluR5激动后,ERG a、b波幅值明显降低,mGluR5主要调控外层视网膜的对光反应。     

Neuroprotective effect of hepatocyte growth factor against photoreceptor degeneration in rats

PURPOSE: To determine whether hepatocyte growth factor (HGF) has a neuroprotective effect against photoreceptor degeneration in rats. METHODS: Eight-week-old Sprague-Dawley (SD) and 24-day-old Royal College of Surgeons (RCS) rats received an intravitreal injection of HGF in the right eyes. The left eyes were injected with vehicle and served as the control. Two days after the injections, the SD rats were exposed to fluorescent light of 3000 lux for 72 hours. Scotopic and photopic electroretinograms (ERGs) were recorded 2 weeks after the light damage and at 70 days of age in RCS rats. After the ERG recordings, the animals were killed for histologic analysis. Some RCS rats were killed at 2 weeks after HGF-treatment for TdT-dUTP terminal nick-end labeling (TUNEL) studies. RESULTS: In both light-damaged and RCS rats, the thresholds for the scotopic and photopic b-wave were significantly lower in the HGF-treated eyes than in the control eyes (P <0.02). The maximum b-wave amplitudes (Vbmax) of the scotopic and photopic ERGs were significantly larger in the HGF-treated eyes (P <0.0005) with a significantly greater number of photoreceptor nuclei than in the control eyes in both animal models (P <0.005). The vehicle-injected eyes of RCS rats had significantly larger numbers of TUNEL-positive photoreceptor nuclei than the HGF-treated eyes (P=0.005). CONCLUSIONS: Intravitreal HGF led to the morphologic and physiological preservation of photoreceptors in rats with photoreceptor degeneration induced by phototoxicity or a gene mutation. The antiapoptotic effect may be the mechanism for the neuroprotective action of HGF.     

S-cone electroretinograms in multiple evanescent white dot syndrome

Cone electroretinograms (ERGs), elicited by different color flashes under Ganzfeld conditions, were recorded from 6 patients with multiple evanescent white dot syndrome (MEWDS). All of the patients had normal color vision as determined by the Farnsworth Panel D-15 except for one who showed non-specific errors. The b-waves elicited from short wavelength sensitive (S-) cones were reduced more than the mixed long (L-) and middle (M-) wavelength sensitive cones in the affected eyes. The ratio of the S-cone b-wave amplitude of the affected eyes to that of the normal fellow eyes was significantly lower than the comparable ratio for the L- and M-cone ERG b-waves (p= 0.012). The S-cone ERGs recorded from 2 patients recovered to normal levels after their symptoms abated. These ERG results indicate that the S-cone system is more impaired than the L- and M-cone systems in the acute stage of MEWDS, and the changes in the S-cones may be reversible.     

Changes in ERG amplitude following laser induced damage to the primate retina

The flash evoked ERG is used extensively as a measure of retinal function. To understand better the relationship between extent of retinal damage and the ERG we created multistage laser lesions in one eye each of three Macaca fascicularis monkeys. An argon-dye laser was used to deliver 630 nm energy to cumulative quarter sectors of the temporal hemiretina and the foveal region at two week intervals. Latencies of the a- and b-waves and amplitude from the trough of the a-wave to peak of the b-wave were measured preoperatively and one week after each new lesion. Following the last recording session the eyes were removed and prepared for histological examination. In general, but depending on stimulus condition, the maximum decrease in ERG amplitude (50-70% of normal) occurred following the third or fourth sector lesions. These lesions accounted for a loss of the outer nuclear layer in 18-25% of the entire retinal surface area extending from the edge of the fovea into the anterior equatorial region. Surprisingly, subsequent lesions of the remaining sector and fovea did not result in further reduction of ERG amplitudes. Significant increases in amplitudes were even observed for some stimulus conditions following these final lesions. Latencies of the a- and b-waves were not affected. The fact that in the present study ERG amplitudes recovered, suggest that such values may be tenuous evidence of amount of functional retina in laser-lesioned eyes.     

Clinical and electrophysiologic results after intracameral lidocaine 1% anesthesia: a prospective randomized study.

OBJECTIVE: To evaluate the efficacy and safety of intracameral lidocaine in cataract surgery compared to peribulbar anesthesia. DESIGN: A prospective, randomized, controlled study. PARTICIPANTS: A total of 200 consecutive cataract patients (200 eyes) participated. INTERVENTION: Eyes were randomly assigned to two groups: one group received 0.15 ml intracameral 1% unpreserved lidocaine combined with topical anesthesia (oxybuprocaine); the other group received 6 ml prilocaine peribulbar before phacoemulsification with sclerocorneal tunnel incision. MAIN OUTCOME MEASURES: Duration of surgery was measured; implicit time and amplitudes of the b-waves of the photopic electroretinogram (ERG) potentials (single-flash ERG and the 30-Hz flicker ERG) were recorded; frequencies of intraoperative problems, complications, intraoperative, and postoperative pain were evaluated. RESULTS: After lidocaine anesthesia combined with topical anesthesia, similar complications were found, longer operation time (P < 0.001), and significantly better visual acuity immediately after surgery (P < 0.001). The ERG amplitudes were not significantly reduced after 0.15-ml intracameral lidocaine half an hour after surgery (P > 0.05). CONCLUSION: Intracameral lidocaine 1% combined with topical anesthesia can be recommended as an alternative procedure to peribulbar anesthesia in cataract surgery with corneoscleral tunnel incision.     

Neuroprotective effect of subretinal implants in the RCS rat

PURPOSE: Retinal prosthetics have been designed to interface with the neural retina by electrically stimulating the remaining retinal circuits after photoreceptor degeneration. However, the electrical stimulation provided by the subretinal implant may also stimulate neurotrophic factors that provide neuroprotection to the retina. This study was undertaken to determine whether electrical stimulation from a subretinal photodiode-based implant has a neuroprotective effect on photoreceptors in the RCS rat, a model of photoreceptor degeneration. METHODS: Eyes of RCS rats were implanted with an active or inactive device or underwent sham surgery before photoreceptor degeneration. Outer retinal function was assessed with electroretinogram (ERG) recordings weekly until 8 weeks after surgery, at which time retinal tissue was collected and processed for morphologic assessment, including photoreceptor cell counts and retinal layer thickness. RESULTS: At 4 to 6 weeks after surgery, the ERG responses in the active-implant eyes were 30% to 70% greater in b-wave amplitude than the responses from eyes implanted with inactive devices, those undergoing sham surgery, or the nonsurgical control eyes. At 8 weeks after surgery the ERG responses from active-implant eyes were not significantly different from the control groups. However, the number of photoreceptors in eyes implanted with the active or inactive device was significantly greater in the regions over and around the implant versus sham-surgical and nonsurgical control eyes. CONCLUSIONS: These results suggest that subretinal electrical stimulation provides temporary preservation of retinal function in the RCS rat. In addition, implantation of an active or inactive device into the subretinal space causes morphologic preservation of photoreceptors in the RCS rat until 8 weeks after surgery. Further studies are needed to determine whether the correlation of neuropreservation with subretinal implantation is due to electrical stimulation and/or a mechanical presence of the implant in the subretinal space.     

Lens epithelium-derived growth factor promotes photoreceptor survival in light-damaged and RCS rats

PURPOSE: To investigate possible protective effects of lens epithelium-derived growth factor (LEDGF) against photoreceptor death in light-damaged, Royal College of Surgeons (RCS) and P23H rhodopsin transgenic rats. METHODS: Twelve-week-old Sprague-Dawley (SD), 6-week-old RCS, and 10-day-old P23H (line 1, heterozygote) rats received an intravitreal injection of LEDGF fused with glutathione-S-transferase (GST-LEDGF). Fellow eyes received vehicle and served as control specimens. Two days after the injections, the SD rats were exposed to light of 2000 lux for 48 hours. Corneal Ganzfeld ERGs were recorded 10 days after light damage, at 10 weeks of age in RCS rats, and at 4 weeks of age in P23H rats. The eyes were then processed for histologic analysis. Heat shock protein (hsp) content in the sensory retina was analyzed quantitatively by protein immunoblot. RESULTS: In light-damaged rats, the ERG indicated retinal protection in GST-LEDGF-injected eyes, with b-wave and STR thresholds being 1.14 +/- 0.50 (mean +/- SD) and 0.60 +/- 0.26 log candela (cd)/m2 lower, respectively, than in vehicle-injected eyes (P < 0.01). The GST-LEDGF-treated eyes had maximum b-wave amplitudes that were significantly larger (P < 0.0005), had more than twice as many remaining photoreceptors, and had better organized outer segments than the control eyes. In RCS rats, the treated eyes had 2.76 +/- 0.73 and 0.83 +/- 0.09 log cd/m(2) lower thresholds for the b-wave and STR, respectively (P < 0.005), and had significantly larger maximum b-wave amplitude (P < 0.0005). GST-LEDGF-treated eyes of RCS rats also had more photoreceptors remaining (P < 0.005) and a thinner debris layer than control eyes. In P23H rats, GST-LEDGF treatment did not protect either retinal function or structure. The retinas from GST-LEDGF-treated eyes of SD and RCS rats had higher levels of hsp25 and alphaB-crystallin than vehicle-injected eyes. CONCLUSIONS: GST-LEDGF protects photoreceptor structure and function in both light-damaged and RCS rats. The increased expression of hsp25 and alphaB-crystallin may play a role in this protection. The absence of rescue in P23H raises the possibility that some forms of inherited retinal degeneration may not be amenable to treatment by intraocular injection of LEDGF.     

Study of blue and red flash in dark-adapted electroretinogram

PURPOSE: To investigate the characteristics of the waveform generated by blue and red light stimulations in a dark-adapted electroretinogram (ERG) and those of cone responses in the dark-adapted condition. METHODS: The study subjects were 52 persons (88 eyes) with no previous medical history. The author recorded b-waves (rod response) with red light stimulation and the x-waves (dark-adapted cone response) that appeared before the b-waves. The author also recorded b-waves with blue light stimulation, which had the same amplitude as the b-waves from the red light stimulation. The differences with respect to age and gender were studied. Waveforms of the dark-adapted cone ERGs were recorded by using a digital subtraction technique. RESULTS: The x-wave always appeared before the b-wave with 0 dB (2.4 cd.s/m2) red stimulation. With blue stimulation, a b-wave equivalent to the b-wave stimulated with the red light of 0 dB intensity appeared at an average of -14.57 dB. The implicit time for the b-wave was delayed significantly for the male group. There were no significant differences between different age groups. The dark-adapted cone ERG demonstrated the waveform of a negative response followed by a series of oscillatory potentials (OPs) and a positive response. CONCLUSIONS: The cone responses were followed by the rod responses with red light stimulation of 0 dB in the dark-adapted ERG. The waveforms of the cone ERGs were obtained in dark adaptation with red and blue light stimulation.     

Intraocular CNTF reduces vision in normal rats in a dose-dependent manner

PURPOSE: CNTF is a neuroprotective agent for retinal degenerations that can cause reduced electroretinogram (ERG) amplitudes. The goal of the present study was to determine the effects of intraocular delivery of CNTF on normal rat visual function. METHODS: Full-field scotopic and photopic ERG amplitudes and spatial frequency thresholds of the optokinetic response (OKR) of adult Long-Evans rats were measured before and after intravitreous injection of CNTF or subretinal delivery of adenoassociated virus-vectored CNTF (AAV-CNTF) into one eye. Visual acuity was also measured by using the Visual Water Task in AAV-CNTF-injected animals. Multiunit luminance thresholds were recorded in the superior colliculus after CNTF injection, and the eyes were examined histologically. RESULTS: In eyes injected with a high dose of CNTF, ERG amplitudes and OKR thresholds measured through CNTF-injected eyes were decreased by 45% to 70% within 6 days after injection. ERG amplitudes had begun to recover by 21 days, whereas OKR thresholds only began to recover after 56 days. Neither OKR thresholds nor ERG amplitudes fully recovered until 90 to 100 days. When measured in the superior colliculus at 2 weeks after CNTF injection, luminance thresholds were elevated by 0.35 log units. In AAV-CNTF-injected eyes, OKR thresholds, and visual acuity were reduced by approximately 50% for at least 6 months, and scotopic and photopic ERG b-waves were reduced by 30% to 50%. Photoreceptor loss occurred in the injected regions in some of the eyes. By contrast, comparison of dose-response analysis with a dose-response study of light damage strongly suggests that therapeutic doses of CNTF exist that do not suppress ERG responses. CONCLUSIONS: Intraocular delivery of CNTF, which preserves photoreceptors in animal models of retinal degeneration, impairs visual function in normal rats at very high doses, but not at lower doses that still provide protection from constant light damage.