Christensen O, Hoffmann E K
Institute of General Physiology and Biophysics, Panum Institute, University of Copenhagen, Denmark.
J Membr Biol. 1992 Jul;129(1):13-36. doi: 10.1007/BF00232052.
Cell-attached patch-clamp recordings from Ehrlich ascites tumor cells reveal nonselective cation channels which are activated by mechanical deformation of the membrane. These channels are seen when suction is applied to the patch pipette or after osmotic cell swelling. The channel activation does not occur instantaneously but within a time delay of 1/2 to 1 min. The channel is permeable to Ba2+ and hence presumably to Ca2+. It seems likely that the function of the nonselective, stretch-activated channels is correlated with their inferred Ca2+ permeability, as part of the volume-activated signal system. In isolated inside-out patches a Ca(2+)-dependent, inwardly rectifying K+ channel is demonstrated. The single-channel conductance recorded with symmetrical 150 mM K+ solutions is for inward current estimated at 40 pS and for outward current at 15 pS. Activation of the K+ channel takes place after an increase in Ca2+ from 10(-7) to 10(-6) M which is in the physiological range. Patch-clamp studies in cell-attached mode show K+ channels with spontaneous activity and with characteristics similar to those of the K+ channel seen in excised patches. The single-channel conductance for outward current at 5 mM external K+ is estimated at about 7 pS. A K+ channel with similar properties can be activated in the cell-attached mode by addition of Ca2+ plus ionophore A23187. The channel is also activated by cell swelling, within 1 min following hypotonic exposure. No evidence was found of channel activation by membrane stretch (suction). The time-averaged number of open K+ channels during regulatory volume decrease (RVD) can be estimated at 40 per cell. The number of open K+ channels following addition of Ca2+ plus ionophore A23187 was estimated at 250 per cell. Concurrent activation in cell-attached patches of stretch-activated, nonselective cation channels and K+ channels in the presence of 3 mM Ca2+ in the pipette suggests a close spatial relationship between the two channels. In excised inside-out patches (with NMDG chloride on both sides) a small 5-pS chloride channel with low spontaneous activity is observed. The channel activity was not dependent on Ca2+ and could not be activated by membrane stretch (suction). In cell-attached mode single-channel currents with characteristics similar to the channels seen in isolated patches are seen. In contrast to the channels seen in isolated patches, the channels in the cell-attached mode could be activated by addition of Ca2+ plus ionophore A23187.(ABSTRACT TRUNCATED AT 400 WORDS)
对艾氏腹水癌细胞进行的细胞贴附式膜片钳记录显示，存在非选择性阳离子通道，这些通道可被膜的机械变形激活。当向膜片吸管施加吸力或细胞渗透性肿胀后，可观察到这些通道。通道激活并非瞬间发生，而是有1/2到1分钟的延迟。该通道对Ba2+有通透性，因此推测对Ca2+也有通透性。非选择性的、牵张激活通道的功能似乎与其推测的Ca2+通透性相关，作为容积激活信号系统的一部分。在分离的内面向外膜片中，证明了一种Ca(2+)依赖性内向整流钾通道。在对称的150 mM K+溶液中记录的单通道电导，内向电流估计为40 pS，外向电流为15 pS。当Ca2+从10(-7) M增加到10(-6) M（处于生理范围内）时，钾通道被激活。细胞贴附模式下的膜片钳研究显示，存在具有自发活性且特性与切除膜片中所见钾通道相似的钾通道。在外部K+为5 mM时，外向电流的单通道电导估计约为7 pS。在细胞贴附模式下，通过添加Ca2+加离子载体A23187可激活具有类似特性的钾通道。该通道也可在低渗暴露后1分钟内被细胞肿胀激活。未发现膜牵张（吸力）激活通道的证据。在调节性容积减小（RVD）期间，开放钾通道的时间平均数量估计为每个细胞40个。添加Ca2+加离子载体A23187后，开放钾通道的数量估计为每个细胞250个。在吸管中存在3 mM Ca2+的情况下，细胞贴附膜片中牵张激活的非选择性阳离子通道和钾通道同时被激活，这表明这两种通道之间存在紧密的空间关系。在切除的内面向外膜片（两侧均有NMDG氯化物）中，观察到一个小的5-pS氯离子通道，其自发活性较低。通道活性不依赖于Ca2+，且不能被膜牵张（吸力）激活。在细胞贴附模式下，可观察到具有与分离膜片中所见通道相似特性的单通道电流。与分离膜片中所见通道不同，细胞贴附模式下的通道可通过添加Ca2+加离子载体A23187激活。（摘要截取自400字）
