Department of Ophthalmology and Visual Science, University of Nebraska Medical Center, Omaha, NE 68198, USA.
Curr Eye Res. 2012 Mar;37(3):218-27. doi: 10.3109/02713683.2011.652756.
PURPOSE/AIM: To compare the efficacy of optical techniques with electrophysiological recordings for mapping retinal activity in response to electrical stimulation.
Whole cell patch clamp, Ca(2+) imaging (Fluo-4-AM), and Na(+) imaging (CoroNa Green-AM) techniques were used to detect responses of neurons from mouse and salamander retina to electrical stimulation.
Synaptic currents were observed in ≥23% of retinal ganglion cells (RGCs), indicating presynaptic Ca(2+) increases in the inner plexiform layer (IPL). Modest depolarization with 20-30 mM K(+) consistently evoked Ca(2+) responses measured with Fluo4, but Ca(2+) responses were almost never evoked by epiretinal stimulation. In salamander retina, responses were seen in the inner nuclear layer (INL) and IPL. In mouse retina, responses were also sometimes seen in the outer pexiform layer (OPL). OPL responses showed a longer latency than IPL responses, suggesting that outer retinal circuits do not trigger synaptic responses of RGCs. Simultaneous Ca(2+) imaging and electrophysiological recording of synaptic currents confirmed that Fluo4-loaded retinas remained responsive to stimulation. Epiretinal stimulation evoked action potentials in ≥67% of RGCs. CoroNa Green detected Na(+) changes stimulated by 20 mM K(+), but epiretinal stimulation did not evoke detectable Na(+) responses. Simultaneous imaging and electrophysiological recording confirmed the health of CoroNa Green-loaded retinas. We confirmed stimulation efficacy by simultaneously recording Na(+) changes and electrophysiological responses.
These data demonstrate that electrophysiological recordings show greater sensitivity than Na(+) or Ca(2+) imaging in response to electrical stimulation. The paucity of Ca(2+) responses is consistent with limited risk for Ca(2+)-mediated cell damage during electrical stimulation.
比较光学技术与电生理记录在探测电刺激视网膜反应中的效果。
应用全细胞膜片钳、钙成像（Fluo-4-AM）和钠成像（CoroNa Green-AM）技术检测来自小鼠和蝾螈视网膜神经元对电刺激的反应。
≥23%的视网膜神经节细胞（RGCs）中观察到突触电流，表明内丛状层（IPL）存在前突触 Ca(2+)增加。用 Fluo4 测量时，20-30mM K(+)引起的适度去极化可诱发 Ca(2+)反应，但视网膜外刺激几乎从未诱发 Ca(2+)反应。在蝾螈视网膜中，在内核层（INL）和 IPL 中观察到反应。在小鼠视网膜中，在外侧膝状体层（OPL）中也观察到反应。OPL 反应的潜伏期比 IPL 反应长，这表明外视网膜回路不会触发 RGC 的突触反应。同时进行 Ca(2+)成像和突触电流电生理记录证实，负载 Fluo4 的视网膜仍能对刺激产生反应。视网膜外刺激可诱发≥67%的 RGC 产生动作电位。CoroNa Green 可检测 20mM K(+)刺激引起的 Na(+)变化，但视网膜外刺激未引起可检测的 Na(+)反应。同时进行成像和电生理记录证实负载 CoroNa Green 的视网膜状态良好。我们通过同时记录 Na(+)变化和电生理反应来证实刺激效果。
这些数据表明，电生理记录在探测电刺激反应中的敏感性高于 Na(+)或 Ca(2+)成像。Ca(2+)反应的缺乏与电刺激期间 Ca(2+)介导的细胞损伤风险有限一致。
