Objective To study the response of the retinal neuronal adaptive system to changes of background illumination (BG) by measuring the oscillatory potentials (OPs) and the a- and b-waves of the electroretinogram (ERG) in different BG illuminations. Methods The a- and b-wave and the digitally filtered OPs were simultaneously recorded from Wistar Fu rats aged from 25 to 29 days during dark adaptation (DA) and during 6~8 minutes of BG illuminations at four levels increased successively by steps of two log units, i.e., ldquo;low scotopicrdquo; level of 1.43times;10-6cd/m2, ldquo;high scotopicrdquo; of 1.43times;10-4cd/m2 , ldquo;low mesopicrdquo; of 1.43times;10-2cd/m2 and ldquo;high mesopicrdquo; of 1.43times;10-2cd/m2. Full field stimulus flashes of 75 msec duration and 1.43times;10-2cd/m2intensity was delivered at an interval of 1 minute. Results Five OP wavelets were recorded in DA and during scotopic BG illuminations. The number of wavelets was reduced to three as the eyes were exposed to mesopic BG levels. However, the sum of OPs amplitudes (SOPs) increased as the BG was intensified, except at ldquo;high mesopicrdquo; level, by which a significant decrease of SOPs occurred. The amplitudes of the a-and b-waves remained unchanged at the two scotopic BG and decreased as the BG intensity increased to mesopic levels. Conclusion The response of retinal neural adaptive system of the Albino rat to changes of BG light is more sensitive and robust than the slow components of the ERG. The enhancement of the oscillatory responses at ldquo;low mesopicrdquo; illumination level suggests that using proper BG light may be conducive to reducing the variation of OPs. (Chin J Ocul Fundus Dis, 2001,17:286-288)
PURPOSE:The changes of expression level of rhodopsin mRNA and its relationship with the morphology in light damaged rat retinas were studied. METHODS:The changes of expresson level of rhodopsin mRNA in light damaged rat retinas and the changes on retinal morphology were observed through the technique of in situ hybridization and electron microscopy. RESULTS:The hybridization signals of rhodopsin mRNA mainly distributed in the photoreceptor layer of retina,relatively b in the inner and outer segments. As the increase of light exposure time,the expression level of rhodopsin mRNA in retinas greatly decreased before the changes on morphological injury of retina. For the same eye globe of the same rat at the same time,the hybridization signals at the upper and posterior region of the retina decreased more obviously than the lower and peripheral region of the retina. CONCLUSIONS:It was demonstrated for the first time that the expression of rhodopsin mRNA was located at the photoreceptor layer of the retina. Continuous exposure to light could greatly decrease the expression of rhodopsin mRNA and the decreases differ regionally. It might be the early signals of retinal photic injury.It is a good method to study the expression level of retina mRNA through the in situ hybridization. （Chin J Ocul Fundus Dis,1997,13: 228-210）
Objective To observe the effect of visible light on apoptosis of cultured human retinal pigment epithelium (RPE) cells. Methods Being the light source,500lx,(2 000±500)lx and (3 400±200)lx cold white light were used. The duration of exposure was 0,6,12 and 24 hours respectively. Apoptosis was detected by terminal deoxynucleotidyl transferase mediated dUTP nick end labelling, Annexin V-flunorescein isothiocyanate/Propidium iodium labelling and flow cytometry. Results Apoptosis and necrosis were found in cultured human RPE cells which were exposed to visible light.(1)A significant increase in apoptotic and necrotic percentages was consistent with a higher light intensity.(2)Apoptosis was the main response to shorter (6 h and 12 h) exposure duration,while necrosis was more pronounced correlated to the prolongation of post-exposure culture (P＜0.05),and the longer the post-exposure period was, the more apoptotic necrosis were seen.Thirty-six hours after exposure the necrotic percentages were more pronounced (P＜0.01). Conclusions Visible light (＞500 lx) increases the proportion of apoptosis and necrosis of human RPE cells in vitro.The extent is related to exposure intensity and duration. It demonstrates that the lower intensity and the shorter duration of exposure to light are, the more pronounced apoptotic percentages are observed,otherwise necrosis. (Chin J Ocul Fundus Dis, 2002, 18: 227-230)
Objective To observe the effect of blue light on apoptosis of cultured human retinal pigment epithelial (RPE) cells in vitro. Methods Human RPE cells were exposed to blue light, and the cells were divided into 3 groups: group A, with various intensity of illumination; group B: with same intensity but different time of illumination; group C: with same intensity and time of illumination but different finish time of the culture. The apoptosis of RPE cells was observed by TdT-dUTP terminal nick-end labeling (TUNEL) and annexin V-fluoresein isothiocyanate (FITC)/propidium iodide (PI) flow cytometry, and transmission electron microscopy. Results The positive cells stained by TUNEL shrinked and turned round, whose nuclei concentrated and congregated like the crescent or hat. Cracked nuclei and membrane bleb were found. Swollen mitochondrial, disappeared inner limiting membrane of mitochondria, and dilation of the rough endoplasmic reticulum with metabolite were observed by transmission electronmicroscopy. In group A, mild damage of RPE cells was found when the threshold value of the intensity of illumination was less than（500±100）lx, and the apoptosis and necrosis of RPE cells aggravated as the intensity of illumination increased; in group B, as the time of illumination extended, the number of apoptotic RPE cells didn′t increase while the necrosis increased; in group C, 6 and 12 hours after illumination, apoptosis of cells was the main injury, while apoptosis with necrosis was found and necrotic cells increased as the time of illumination was prolonged. Conclusions Illumination with blue light may cause damages of human RPE cells in vitro, with the modalities of apoptosis, apoptotic necrosis and necrosis. The extent of injury is dependent on intensity and duration of the illumination. (Chin J Ocul Fundus Dis, 2005, 21: 384-387)
Objective To observe the effect of persistent flickering stimulus on the structure and function of retina in guinea pigs during a developmentally sensitive period.Methods Twenty-four two- week-old guinea pigs were randomly divided into flicker light (FL) group and control group, with 12 guinea pigs in each group. Animals in FL group were raised under 500 Lux illumination with a duty diurnal cycle of 50% at a flash rate of 0.5 Hz. Animals in control group were reared under steady 500 Lux illumination. Light emitting diode (LED) lamps were used for lighting under a 12-hour light/12-hour dark cycle. After the collection of fundus photographs and electroretinograms recorded at week 12, eyeballs were taken out, three dimensions were measured, and histopathological changes were examined.Results Compared to control group, tessellated fundus in FL group appeared more prevalent; implicit time of ldquo;ardquo; waves were prolonged in electroretinogram; the eyeballs were increased in horizontal, vertical, axial dimensions by (0.89plusmn;0.30), (0.69plusmn;0.20) and (0.96plusmn;0.30) mm respectively, the differences between two groups were statistically significant (t=12.7,11.9,15.8;P＜0.05). The gap of sclera collagen fiber was slightly widened.The photoreceptor layer was more likely to develop a disordered outer segment, which contained deciduous disc membranes.Conclusion Persistent flickering stimulus is attended by development of excessive ocular enlargement,which could affect the retinal structure and function of photoreceptors.
Objective To investigate the relationship between exposure intensity and illumination time of blue light and replicative senescence of rat retinal pigment epithelial （RPE） cells.Methods Thirtysix 12-14 weeks Wistar rats were kept in the cage with a bluelight bulb ［(450plusmn;10) nm］, and were randomly divided into four groups (no light，nature light，500 lx light and 1000 lx light illumination), each has nine rats. The rats in each group were further divided into three subgroups according to illumination time (one month，two months or three months). Eyeballs were collected after intraperitoneal injection of 10% chloral hydrate. The right eye of each rat was embedded in paraffin and sectioned for hematoxylineosin (HE) staining, while frozen sections of the left eye were stained for the senescence-associated beta;-galactosidase (SA-beta;-Gal). The data were analyzed by SPSS11.5 statistical software.Results The amounts of SA-beta;-Gal positive RPE cells were significantly different between all groups under the same illumination time 17 (P＝0.000）, and between all subgroups of different illumination time with same exposure intensity （P＜0.01）except for the control group (no light). Conclusion Bluelight can induce replicative senescence in rat RPE cells in an intensity and timedependent manner.
Objective To observe the expression of vascular endothelial growth factor A (VEGFA) and its receptors sFlt-1, kinase insert domain receptor (KDR) in lightinjured human retinal pigment epithelial (RPE) cells. Methods Cultured human RPE cells (8th - 12th generations) were divided into normal control group and light damage group. The cells of two groups were exposed to the 18 W cold white light (2200±300) Lux for 12 hours to induce light damage responses, but the cells of normal control group were packed by tinfoil with doubledeck high pressure disinfection. The VEGF-A, sFlt-1 and KDR mRNA and protein expressions were detected by reverse transcriptionpolymerase chain reaction (RT-PCR) and Western blot at 0, 6, 12, 24 hours after light damage. Results The VEGF-A mRNA and protein expressions in light damage group were significantly increased at 6 hours, and reached its peak at 12 hours after light damage which obviously higher than that in normal group (t=2.74, 2.93; P＜0.05), and then went down gradually. The sFlt-1 mRNA and protein expressions in light damage group reached its peak at 12 hours after light damage which obviously higher than that in normal group (t=4.32, P＜0.01), but obviously lower than that in normal group at 24 hours after light damage (t=2.41, P＜0.05). The KDR mRNA and protein expressions in light damage group were obviously higher than that in normal group at 24 hours after light damage （t=2.89, P＜0.05）,but there was no changes at 6, 12 hours after light damage (t=1.84, P＞0.05). Conclusions At 6, 12 hours after light damage, the expressions of VEGF-A and sFlt-1 increases significantly and KDR expression is stable in lightinjured RPE cells. At 24 hours after light damage, the expression of VEGF-A and sFlt-1 decreases, but KDR expression increases in light-injured RPE cells.
Objective To investigate the effect of blue light on mRNA expression of L-type calcium channel subtypes of human retinal pigment epithelial (RPE) cells in vitro. Methods The fourth-generation of human RPE cells were randomly divided into four groups including control group (no light group), light group, light + nifedipine group, and light + (-) BayK8644 group. The cells were exposed to blue light (2000plusmn;500) lux for 6 hours, and then cultured for another 24 hours. Reverse transcription polymerase chain reaction real time (RT-PCR) and fluorescence quantitative PCR technologies were used to analyze mRNA expression of L-type calcium channel subunit of cardiac subtype ( 1C or CaV1.2), neuroendocrine subtype ( 1D or CaV1.3) and retinal subtypes ( 1F or CaV1.4) in each group. Results The length of PCR product of 1C, 1D, 1F subunit and actin was 68, 157, 125 and 186 base pairs respectively. (1) 1C mRNA expression in light, light + nifedipine and light + (-) BayK8644 group was higher than that in control group, the difference was statistically significant (P＜0.05). 1C mRNA expression in light +nifedipine group and light + (-) BayK8644 group was higher than in light group (P＜0.05). 1C mRNA expression in light + (-) BayK8644 group was higher than that in light + nifedipine group (P＜0.05). (2) Comparing with control group, 1D mRNA expression was higher in light, light +nifedipine and light + (-) BayK8644 group, the difference was statistically significant (P＜0.05). Light + (-) BayK8644 group was higher than light group and light + nifedipine group (P＜0.05), light group and the light + nifedipine group was not statistically significant (P＞0.05). (3) 1F mRNA expression in light, light + nifedipine and light + (-) BayK8644 group was higher than those in control group, there was statistically significant (P＜0.05), light +nifedipine group and light + (-) BayK8644 group was higher than light group (P＜0.05), light + nifedipine group and the light + (-) BayK8644 group was not statistically significant (P＞0.05). Conclusions The human RPE cells mRNA expression of L-type calcium channel 1C, 1D and 1F subunit was increased after exposing to blue light. Application of the 1times;10-5 mmol/L (-) BayK8644 can increase mRNA expression of 1C, 1D and 1F subunit.
Objective To investigate the degenerative changes in the inner rat retina after photic injury.Methods After 24 hour-dark adaptation, sixty Lewis rats were exposed in a ventilated green plexiglass chamber that transmitted continuous green light between 480-520 nm with an intensity of 900～1 000 lx. After 24 hour exposure, the rats stayed in darkness and were sacrificed after 1 day, 3,7 or 14 days. The neurons in the inner retina were marked by immunohisto chemical technique and observed by light and electronic microscope.Results The apoptotic photoreceptor cells were noted after photic injury. The degeneration and decreasing number of rod bipolar cells were found after 3 days; the edema of horizontal cells occurred after 1 day but ameliorated gradually; decreasing number of amacrine cells was found after 1 day; sustained edema of ganglion cells and prolifeeration of the Müller cells were found after photic injury. Pyknotic and edematous neruronal degenerations of inner retina were found in ultrastructural study.Conclusion The neurons in the inner retina as well as Müller cells are involved in the degeneration after photic injury. Different neurons manifest different patterns of degeneration.(Chin J Ocul Fundus Dis，2003，19：201-268)