Gap Junction Contributions to the Goldfish Electroretinogram at the Photopic Illumination Level 221 Fig. 2. Modulation of the b-wave. (A) The b-wave was recorded under the control condition, after injecting 1 mM SNP, and after injecting a mixture of 1 mM SNP and 200 μM MFA. (B) The b-wave was recorded under the control condition, after injecting 2 mM L-NAME, and after injecting a mixture of 2 mM L-NAME and 200 μM MFA. In bar graphs end of the each low line, the mean peak amplitudes of the b-wave by drugs are plotted as a percentage of the peak response under control condition. In this and all subsequent figures, each bar represents the mean±standard error and single (p＜0.05) and double stars (p＜0.01) above the data column indicate significant and very significant statistical differences, respectively. See text for details of the statistical comparisons. Statistical analysis All data are reported as mean±S.E.M. Student’s t-test was used with p＜0.05 and p＜0.01 as the criterion for stat- istical significance and very significance. Origin software (Microcal, Northampton, MA, USA) was used to calculate p values. RESULTS Fig. 1 shows the diagram of the light stimulation process 2 for ERG measurement. The white circle light (980 μW/cm ) emitted from the beam projector reached the eye of the goldfish and was reflected 45o onto a mirror (screen role gyeomham). As a lens is present inside the eye during the corneal ERG measurements, the circle light beam coming into the eye should have a path as shown in Fig. 1. First, we observed the effect of MFA on the ERG according to two different circle light beam diameters. The small and large sizes of the light beam imaged on the surface of the mirror were 6 mm and 60 mm in diameter corresponding to the values of 0.012o and 0.12o in the field of view. In control conditions, the amplitudes of a- and b-wave by large size stimulation were each 12±3.4% and 9.2±3.0% higher than those by the smaller size light beam (Fig. 1B. upper and down, left). After injecting of 200 μM MFA into the vitreous, ERG b-wave amplitudes increased 43±11% (mean ±standard error) by the small size light beam (Fig. 1B. up- per) and 8.1±1.8% by the large size light beam (Fig. 1B. down). The a- and d-waves remained unchanged. The en- hancement of the b-wave by the small and large sized stim- uli were statistically significant (n=14, p＜0.05 paired t-test). In the presence of 200 μM MFA, the enhancement by the small size beam was about 5 times larger than the enhancement by large size beam. Thus, we used the small sized beam as the light stimulation in this study. Intravitreal injection of 1 mM SNP enhanced b-wave am- plitude by 46±8% of the control (n=18, p＜0.01, paired t-test). This enhanced amplitude by SNP did not increase more after further injection of a mixture of 1 mM SNP and 200 μM MFA (Fig. 2A). Injection of NG-nitro-L-arginine methyl ester (L-NAME), a nitric oxide synthase (NOS) in- hibitor, resulted in a greatly increased a-wave by 103±18%, whereas the b-wave decreased slightly by －19±4.3% (n=6, Fig. 2B). The effects of 2 mM L-NAME on a- and b-waves were statistically significant (p＜0.01 and p＜0.05, respec- tively). The b-wave peak amplitude that decreased follow- ing administration of L-NAME was enhanced significantly by 51±12% (n=6, p＜0.01, paired t-test) compared to the peak of L-NAME by further injection of a mixture of 2 mM L-NAME and 200 μM MFA into the vitreous. To evaluate the contribution of DA receptors on ERG b-waves, we used the dopamine D1 and D2 receptors re- lated drugs alone first and then a mixture of these with MFA (Figs. 3, 4). DA is known to reduce gap-junction cou- pling through a protein kinase A metabolic cascade in reti- nal cells. Fig. 3 shows the relationship between dopamine D1 receptor-related drugs and MFA on b-waves at the pho- topic level. Injection of 100 μM SKF 38393, a D1 receptor agonist, greatly enhanced b-wave amplitude by 39±12% compared to that of the control (n=12, p＜0.01, paired t-test; Fig. 3A). This enhanced b-wave peak following the 100 μM SKF 38393 injection did not change much (2±0.4%) after a further injection of a mixture of 100 μM SKF 38393 and 200 μM MFA. The injection of 30 μM SCH 23390, a dop- amine D1 receptor antagonist, reduced b-wave amplitude by －15±9% compared to that in the control (Fig. 3B). This reduced peak was enhanced by up to 71±16% compared to that in the control. This reduction and increase were both statistically significant (n=7, p＜0.05, paired t-test). The b-wave decreased by －22±4.3% (n=6, p＜0.05, paired t-test; Fig. 4A) compared to that in the control following an injection of 100 μM quinpirole, a selective D2-dopamine receptor agonist. This decreased amplitude b-wave was sig- nificantly enhanced by 36±13% (p＜0.05, paired t-test) com- pared to that in the control after an additional injection of a mixture of 100 μM quinpirole and 200 μM MFA into the vitreous. In contrast, when the retina was treated with 200 μM sulpiride, a D2-dopamine receptor antagonist, the