Milesi V, Aiello E A, Rebolledo A, Gomez Alvis A, Grassi de Gende A O
Anatomía y Fisiología, Departamento de Ciencias Biológicas, Facultad de Ciencias Exactas, Universidad Nacional de La Plata. Calles 47 y 115, La Plata (1900), Argentina.
Pflugers Arch. 1999 Feb;437(3):455-61. doi: 10.1007/s004240050801.
Calcium-activated potassium currents were studied in dissociated smooth muscle cells from human saphenous vein (HSV) using the patch-clamp technique in the whole-cell configuration. The average measured resting membrane potential (Vm) was -41+/-2 mV (n=39), when the cells were dialysed with an intracellular pipette solution (IPS) containing 0.1 mM ethyleneglycol-bis(beta-aminoethylether)-N,N,N', N'-tetraacetic acid (EGTA) (IPS-0.1 mM EGTA). When the EGTA concentration was increased to 10 mM (IPS-10 mM EGTA) Vm became significantly less negative: -13+/-2 mV (n=23, P<0.05). These results suggest that 10 mM EGTA reduces a calcium-dependent current involved in the maintenance of Vm. Depolarizing voltage steps up to +60 mV from holding potentials of -60 mV resulted in large (1-10 nA) time- and voltage-dependent outward currents. The amplitudes of total whole-cell current densities measured at voltages above -20 mV were significantly greater in the cells dialysed with IPS-0.1 mM EGTA than in those dialysed with IPS-10 mM EGTA. In the cells dialysed with IPS-0.1 mM EGTA, 0.1 mM tetraethylammonium chloride (TEA) and 50 nM iberiotoxin (IBTX), which selectively block large conductance Ca2+-activated potassium channels (BKCa), diminished the total current recorded at +60 mV by 45+/-14% (P<0.05, n=5) and 50+/-6% (n=8, P<0.05), respectively. These blockers at the same concentrations did not affect the total current in cells dialysed with IPS-10 mM EGTA. When tested on intact HSV rings, both 0.1 mM TEA and 50 nM IBTX elicited vessel contraction. We conclude that BKCa channels present in HSV smooth muscle cells contribute to the maintenance of the Vm and sustain a significant portion of the total voltage-activated, outward current. Finally, BKCa channels appear to play a significant role in the regulation of HSV smooth muscle contractile activity.
采用全细胞膜片钳技术，对人隐静脉（HSV）分离的平滑肌细胞中的钙激活钾电流进行了研究。当用含有0.1 mM乙二醇双（β-氨基乙基醚）-N，N，N'，N'-四乙酸（EGTA）的细胞内微电极溶液（IPS）（IPS-0.1 mM EGTA）透析细胞时，测得的平均静息膜电位（Vm）为-41±2 mV（n = 39）。当EGTA浓度增加到10 mM（IPS-10 mM EGTA）时，Vm的负值显著减小：-13±2 mV（n = 23，P<0.05）。这些结果表明，10 mM EGTA降低了参与维持Vm的钙依赖性电流。从-60 mV的钳制电位去极化至+60 mV的电压阶跃会导致大的（1 - 10 nA）时间和电压依赖性外向电流。在-20 mV以上电压下测得的全细胞总电流密度幅值，在用IPS-0.1 mM EGTA透析的细胞中显著大于用IPS-10 mM EGTA透析的细胞。在用IPS-0.1 mM EGTA透析的细胞中，0.1 mM氯化四乙铵（TEA）和50 nM埃博毒素（IBTX），它们选择性阻断大电导钙激活钾通道（BKCa），使在+60 mV记录的总电流分别减少45±14%（P<0.05，n = 5）和50±6%（n = 8，P<0.05）。相同浓度的这些阻断剂对用IPS-10 mM EGTA透析的细胞中的总电流没有影响。在用完整的HSV环进行测试时，0.1 mM TEA和50 nM IBTX均引起血管收缩。我们得出结论，HSV平滑肌细胞中存在的BKCa通道有助于维持Vm，并维持总电压激活外向电流的很大一部分。最后，BKCa通道似乎在HSV平滑肌收缩活动的调节中起重要作用。
