Duszynski J, Elensky M, Cheung J Y, Tillotson D L, LaNoue K F
Department of Cellular and Molecular Physiology, Milton S. Hershey Medical Center, Pennsylvania State University Hershey, USA.
Cell Calcium. 1995 Jul;18(1):19-29. doi: 10.1016/0143-4160(95)90042-x.
An inward current responsible for hormone regulated Ca2+ entry has been identified in cultured rat hepatocytes using whole cell patch clamp. Addition of 20 nM vasopressin or of 100 microM ATP induced the inward current, which could be observed more clearly after blocking an outward K+ current. This large outward K+ current, which appeared after addition of vasopressin or ATP, could be blocked either by replacing K+ with Cs+ in the external medium and in the pipette solution, or by simply including 0.5 microM apamin in the K(+)-containing external medium. The outward current appears to be carried by a Ca2+ activated K+ channel. In the presence of apamin, hepatocytes pretreated with vasopressin in a Ca(2+)-free media reveal an inward current on addition of external Ca2+ (5 mM). The current could also be elicited by addition of vasopressin when cells are preincubated in the presence of 5 mM external Ca2+. No current is seen on addition of Ca2+ in the absence of vasopressin. Initially, the inward current was ca 200-300 pA at -60 mV, but it declined rapidly over 3 min to ca 20 pA. The current approached zero, as an asymptote at positive potential, and appeared to be somewhat inwardly rectifying. Additions of 5 mM Mn2+ or 5 mM Ba2+ in place of Ca2+ produced little or no current. An inhibitor of ER Ca(2+)-ATPase, thapsigargin, could also trigger the cascade of events leading to plasma membrane conductance of Ca2+. The data suggest that hormone-stimulated Ca2+ entry into hepatocytes is mediated by a Ca(2+)-release activated channel highly specific for Ca2+. This is the first demonstration of such a channel in hepatocytes, though similar ones have been described in mast cells, in vascular endothelial cells and T-lymphocytes.
利用全细胞膜片钳技术，在培养的大鼠肝细胞中已鉴定出一种负责激素调节钙离子内流的内向电流。添加20 nM血管加压素或100 μM ATP可诱导该内向电流，在阻断外向钾电流后能更清晰地观察到。添加血管加压素或ATP后出现的这种大的外向钾电流，可通过在外部培养基和移液管溶液中用Cs⁺替代K⁺，或简单地在含K⁺的外部培养基中加入0.5 μM蜂毒明肽来阻断。外向电流似乎由钙激活钾通道传导。在存在蜂毒明肽的情况下，在无钙培养基中用血管加压素预处理的肝细胞在加入外部钙离子（5 mM）时显示出内向电流。当细胞在5 mM外部钙离子存在下预孵育时，加入血管加压素也可引发该电流。在无血管加压素时加入钙离子则无电流出现。最初，在 -60 mV时内向电流约为200 - 300 pA，但在3分钟内迅速下降至约20 pA。该电流在正电位时趋近于零，呈渐近线，且似乎有点内向整流。用5 mM Mn²⁺或5 mM Ba²⁺代替Ca²⁺加入时产生的电流很小或无电流。内质网钙ATP酶抑制剂毒胡萝卜素也可触发导致质膜钙离子电导的一系列事件。数据表明，激素刺激的钙离子进入肝细胞是由对钙离子高度特异的钙释放激活通道介导的。这是首次在肝细胞中证明存在这样的通道，尽管在肥大细胞、血管内皮细胞和T淋巴细胞中已描述过类似的通道。
