Department of Integrative Biology and Physiology, University of California, Los Angeles, CA.
Department of Ophthalmology and Jules Stein Eye Institute, University of California, Los Angeles, CA.
J Gen Physiol. 2020 Mar 2;152(3). doi: 10.1085/jgp.201912520.
Vertebrate photoreceptor cells respond to light through a closure of CNG channels located in the outer segment. Multiple voltage-sensitive channels in the photoreceptor inner segment serve to transform and transmit the light-induced outer-segment current response. Despite extensive studies in lower vertebrates, we do not know how these channels produce the photoresponse of mammalian photoreceptors. Here we examined these ionic conductances recorded from single mouse cones in unlabeled, dark-adapted retinal slices. First, we show measurements of the voltage dependence of the light response. After block of voltage-gated Ca2+ channels, the light-dependent current was nearly linear within the physiological range of voltages with constant chord conductance and a reversal potential similar to that previously determined in lower vertebrate photoreceptors. At a dark resting membrane potential of -45 mV, cones maintain a standing Ca2+ current (iCa) between 15 and 20 pA. We characterized the time and voltage dependence of iCa and a calcium-activated anion channel. After constitutive closure of the CNG channels by the nonhydrolysable analogue GTP-γ-S, we observed a light-dependent increase in iCa followed by a Ca2+-activated K+ current, both probably the result of feedback from horizontal cells. We also recorded the hyperpolarization-activated cyclic nucleotide-gated (HCN) conductance (ih) and measured its current-voltage relationship and reversal potential. With small hyperpolarizations, ih activated with a time constant of 25 ms; activation was speeded with larger hyperpolarizations. Finally, we characterized two voltage-gated K+-conductances (iK). Depolarizing steps beginning at -10 mV activated a transient, outwardly rectifying iK blocked by 4-AP and insensitive to TEA. A sustained iK isolated through subtraction was blocked by TEA but was insensitive to 4-AP. The sustained iK had a nearly linear voltage dependence throughout the physiological voltage range of the cone. Together these data constitute the first comprehensive study of the channel conductances of mouse photoreceptors.
脊椎动物光感受器细胞通过位于外节的 CNG 通道关闭来对光作出反应。光感受器内节中的多个电压敏感通道用于转换和传递光诱导的外节电流反应。尽管在低等脊椎动物中进行了广泛的研究，但我们不知道这些通道如何产生哺乳动物光感受器的光反应。在这里，我们在未经标记的暗适应视网膜切片中检查了从单个小鼠锥体记录的这些离子电导。首先，我们展示了光反应的电压依赖性测量。在阻断电压门控 Ca2+通道后，光依赖性电流在生理电压范围内几乎呈线性，具有恒定的弦电导和与先前在低等脊椎动物光感受器中确定的反转电位相似的反转电位。在暗静息膜电位为-45 mV 时，锥体维持 15 到 20 pA 的静息 Ca2+电流 (iCa)。我们描述了 iCa 和钙激活阴离子通道的时间和电压依赖性。在非水解类似物 GTP-γ-S 构成性关闭 CNG 通道后，我们观察到 iCa 的光依赖性增加，随后是钙激活的 K+电流，这两者可能是水平细胞反馈的结果。我们还记录了超极化激活环核苷酸门控 (HCN) 电导 (ih)，并测量了其电流-电压关系和反转电位。用小超极化，ih 以 25 ms 的时间常数激活; 随着更大的超极化，激活速度加快。最后，我们描述了两种电压门控 K+电导 (iK)。从-10 mV 开始的去极化步骤激活了被 4-AP 阻断且对 TEA 不敏感的瞬态、外向整流 iK。通过减法分离的持续 iK 被 TEA 阻断，但对 4-AP 不敏感。持续的 iK 在锥体的整个生理电压范围内具有近乎线性的电压依赖性。这些数据共同构成了对小鼠光感受器通道电导的首次全面研究。
