Rebrik T I, Korenbrot J I
Department of Physiology, School of Medicine, University of California at San Francisco, San Francisco, California 94143, USA.
J Gen Physiol. 1998 Nov;112(5):537-48. doi: 10.1085/jgp.112.5.537.
We investigated the modulation of cGMP-gated ion channels in single cone photoreceptors isolated from a fish retina. A new method allowed us to record currents from an intact outer segment while controlling its cytoplasmic composition by superfusion of the electropermeabilized inner segment. The sensitivity of the channels to agonists in the intact outer segment differs from that measured in membrane patches detached from the same cell. This sensitivity, measured as the ligand concentration necessary to activate half-maximal currents, K1/2, also increases as Ca2+ concentration decreases. In electropermeabilized cones, K1/2 for cGMP is 335.5 +/- 64.4 microM in the presence of 20 microM Ca2+, and 84.3 +/- 12.6 microM in its absence. For 8Br-cGMP, K1/2 is 72.7 +/- 11.3 microM in the presence of 20 microM Ca2+ and 15.3 +/- 4.5 microM in its absence. The Ca2+-dependent change in agonist sensitivity is larger in extent than that measured in rods. In electropermeabilized tiger salamander rods, K1/2 for 8Br-cGMP is 17.9 +/- 3.8 microM in the presence of 20 microM Ca2+ and 7.2 +/- 1.2 microM in its absence. The Ca2+-dependent modulation is reversible in intact cone outer segments, but is progressively lost in the absence of divalent cations, suggesting that it is mediated by a diffusible factor. Comparison of data in intact cells and detached membrane fragments from cones indicates that this factor is not calmodulin. At 40 microM 8Br-cGMP, the Ca2+-dependent change in sensitivity in cones is half-maximal at KCa = 286 +/- 66 nM Ca2+. In rods, by contrast, KCa is approximately 50 nM Ca2+. The difference in magnitude and Ca2+ dependence of channel modulation between photoreceptor types suggests that this modulation may play a more significant role in the regulation of photocurrent gain in cones than in rods.
我们研究了从鱼视网膜分离出的单个视锥光感受器中cGMP门控离子通道的调节作用。一种新方法使我们能够在通过电通透化的内段进行超灌注来控制其细胞质成分的同时，记录完整外段的电流。通道对完整外段中激动剂的敏感性与从同一细胞分离的膜片上测得的敏感性不同。这种敏感性，以激活半数最大电流所需的配体浓度K1/2来衡量，也随着Ca2+浓度的降低而增加。在电通透化的视锥细胞中，在存在20μM Ca2+时，cGMP的K1/2为335.5±64.4μM，在不存在Ca2+时为84.3±12.6μM。对于8Br-cGMP，在存在20μM Ca2+时，K1/2为72.7±11.3μM，在不存在Ca2+时为15.3±4.5μM。激动剂敏感性的Ca2+依赖性变化程度比在视杆细胞中测得的更大。在电通透化的虎蝾螈视杆细胞中，对于8Br-cGMP，在存在20μM Ca2+时，K1/2为17.9±3.8μM，在不存在Ca2+时为7.2±1.2μM。Ca2+依赖性调节在完整的视锥外段中是可逆的，但在没有二价阳离子的情况下会逐渐丧失，这表明它是由一种可扩散因子介导的。对视锥细胞完整细胞和分离的膜片段中的数据进行比较表明，该因子不是钙调蛋白。在40μM 8Br-cGMP时，视锥细胞中敏感性的Ca2+依赖性变化在KCa = 286±66 nM Ca2+时达到半数最大。相比之下，在视杆细胞中，KCa约为50 nM Ca2+。光感受器类型之间通道调节的幅度和Ca2+依赖性差异表明，这种调节在视锥细胞光电流增益调节中可能比在视杆细胞中发挥更重要的作用。
