Miller J L, Korenbrot J I
Department of Physiology, School of Medicine, University of California, San Francisco 94143.
J Gen Physiol. 1993 Jun;101(6):933-60. doi: 10.1085/jgp.101.6.933.
We measured outer segment currents under voltage clamp in solitary, single cone photoreceptors isolated from the retina of striped bass. In darkness, changes in membrane voltage to values more positive than 10 mV activate a time- and voltage-dependent outward current in the outer segment. This dark, voltage-activated current (DVAC) increases in amplitude with a sigmoidal time course up to a steady-state value, reached in 0.75-1.5 s. DVAC is entirely suppressed by light, and its current-voltage characteristics and reversal potential are the same as those of the light-sensitive currents. DVAC, therefore, arises from the activation by voltage in the dark of the light-sensitive, cGMP-gated channels of the cone outer segment. Since these channels are not directly gated by voltage, we explain DVAC as arising from a voltage-dependent decrease in cytoplasmic Ca concentration that, in turn, activates only guanylate cyclase and results in net synthesis of cGMP. This explanation is supported by the finding that the Ca buffer BAPTA, loaded into the cytoplasm of the cone outer segment, blocks DVAC. To link a decrease in cytoplasmic Ca concentration to the synthesis of cGMP and the characteristics of DVAC, we develop a quantitative model that assumes cytoplasmic Ca concentration can be continuously calculated from the balance between passive Ca influx via the cGMP-gated channel and its active efflux via a Na/Ca,K exchanger, and that further assumes that guanylate cyclase is activated by decreasing cytoplasmic Ca concentration with characteristics identical to those described for the enzyme in rods. The model successfully simulates experimental data by adjusting the Ca conductance of the cGMP-gated channels as a function of voltage and the Ca buffering power of the cytoplasm. This success suggests that the activity of guanylate cyclase in cone outer segments is indistinguishable from that in rods.
我们在电压钳制条件下，对从条纹鲈视网膜分离出的单个视锥光感受器的外段电流进行了测量。在黑暗中，当膜电压变化到比10 mV更正的值时，外段会激活一种时间和电压依赖性的外向电流。这种暗态电压激活电流（DVAC）的幅度会以S形时间进程增加，直至达到0.75 - 1.5秒时的稳态值。DVAC完全被光抑制，其电流 - 电压特性和反转电位与光敏电流相同。因此，DVAC是由视锥外段光敏的cGMP门控通道在黑暗中被电压激活而产生的。由于这些通道不是直接由电压门控的，我们将DVAC解释为细胞质Ca浓度的电压依赖性降低所导致的，而这种降低反过来仅激活鸟苷酸环化酶并导致cGMP的净合成。这一解释得到了以下发现的支持：加载到视锥外段细胞质中的Ca缓冲剂BAPTA会阻断DVAC。为了将细胞质Ca浓度的降低与cGMP的合成以及DVAC的特性联系起来，我们开发了一个定量模型，该模型假设细胞质Ca浓度可以通过cGMP门控通道的被动Ca内流与其通过Na/Ca，K交换器的主动外流之间的平衡来连续计算，并且进一步假设鸟苷酸环化酶通过降低细胞质Ca浓度而被激活，其特性与在视杆细胞中描述的该酶相同。该模型通过将cGMP门控通道的Ca电导作为电压的函数以及细胞质的Ca缓冲能力进行调整，成功地模拟了实验数据。这一成功表明，视锥外段中鸟苷酸环化酶的活性与视杆细胞中的无法区分。
