粒细胞巨噬细胞集落刺激因子诱导的巨噬细胞外向K^+电流的特性

PROPERTIES OF THE GM-CSF-INDUCED OUTWARD K^+ CURRENT IN MURINE PERITONEAL EXUDATE MACROPHAGES

以粒细胞巨噬细胞集落刺激因子（GM－CSF，16ng／ml）长期（0.5～6d）刺激小鼠腹腔渗出的巨噬细胞，采用全细胞膜片箝技术研究在GM－CSF刺激过程中细胞膜电流的变化，观察到一种CM－CSF诱导的瞬间失活的外向K+电流（记作IA），该电流在生理电压范围内可发生稳态失活，且当施加0.5Hz的去极化脉冲刺激时其失活具有频率依赖性。该电流对胞外4AP（3mmol／L）高度敏感，胞内Ca2+浓度[Ca2+]；升高可抑制其幅度，环己酮亚胺（0.3μg／ml）作用细胞12h可抑制其表达。值得注意的是这种K+电流的表达和相应电导的激活行为及动力学均随GM－CSF剜激时间长短而异。GM－CSF作用2d,表达该电流的细胞数增至最大（大约55％），通道蛋白的密度亦达高峰，该电流激活的半值电导电位V1／2最低，为-27.55mV，电流激活和失活迅速。随着GM－CSF作用时间的继续延长；表达该电流的细胞数下降，通道蛋白不断下调，电流激活曲线向正电位方向移动，电流激活和失活的时间过程减慢。这些结果表明，CM-CSF可诱导一种瞬间失活的外向K+电流（IA）的表达，提示该电流可能与GM－CSF激活的巨噬细胞的功能活性状态密切相关。

Whole-cell patch-clamp technique was used to study the changes of ionic currents in murine peritoneal exudate macrophages (PEMs ) prestimulated with granulocytemacrophage colony-stimulating factor (GM-CSF, 16 ng/ml) for periods from 0. 5 up to 6 d. The GM-CSF-treated PEMs developed a GM-CSF-induced transient inactivating outwwh K+ current (IA ). IA showed steady-state inactivation over the physiological voltage range and possessed frequency dependence of inactivation When depolarizing pulses were applied at a frequency of 0.5 Hz.IA was selectively inhibited by extracellulax 4-AP (3 mmol/L). When [Ca2+], was increased (from pCa 8 to 6), the amplitudes of IA were depressed significantly. When the PEMs were exposed to cycloheximide (0.3μg/ml), a protein synthesis inhibitor, for 12 h, IA expression was completely suppressed. It was notable that the changes of the current expression, activation hehavior and kinetic properties occurred during GM-CSF treatment. When PEMs were pretreated for a 2-d period, the frequency of IA expression reached a peak value (55% in a total of 27 cells),PEMs exhibited the highest density of the corresponding channel proteins, halfmaximal activation of IA was most easily achieved with a value of-27.55 mV, and the time course of activation and inactivation during depoldrization proceeded rapidly.However, along with continuous incubation with GM-CSF, Ihe nuTnber of PEMs expressing IA decreased, the channel proteins were down regulated constanily, the activation curve for IA shifted to positive potentials, and the activation time and inactivation time of IA slowed down. These results indicated that GM-CSF could induce a transient inactivating outwwh K+ current in PEMs,which may have a close relation to the state of functional activation of macrophages primed with GM-CSF.