Effect of 4 ºC pretreatment on the membrane current and cell polarity in humanneutrophils
YUAN Chun-Hua, CAI Chun-Qing, ZOU Fei*
School of Public Health and Tropical Medicine, Southern Medical University, Guangzhou 510515, China
Abstract
To investigate the role of ion channels in the coupling responses of neutrophils to extracellular stimulus, it is necessary tostudy the membrane ion channel activities using patch-clamp technique. However, little has been known about the ion channel activitiesin neutrophils due to the difficulties in forming giga-seal with pipettes because of small diameter of neutrophils and the easily developedpolarization. Some studies indicated that favorable results could be achieved through pretreatment at low temperature before electrophysiologicalrecordings. But it remains unclear whether the pretreatment affects the membrane current and why the seal rate increasesafter low temperature pretreatment. The purpose of this study was to investigate the effects of 4 ºC pretreatment on the membranecurrent and cell polarity in human neutrophils. In the experiments, human neutrophils were isolated from fresh peripheral blood ofhealthy volunteers and divided into two groups (room temperature group and 4 ºC pretreatment group). Voltage-dependent K+ (Kv)currents were recorded in whole-cell voltage-clamp mode and large-conductance Ca2+-activated K+ (BKCa) currents were recorded usinginside-out patches. The results showed that 4 ºC pretreatment significantly inhibited cell polarity (P<0.05), and it took more time forneutrophils to form a polarity-cycle [(534±32) s, n=20] compared with those at room temperature [(257±24) s, n=20]. Meanwhile,seal rate significantly increased in 4 ºC pretreatment group (64%) compared with that in the room temperature group (27.5%). The sealrate and cell polarity rate during 0~1 min after 4 ºC pretreatment were significantly different from those at room temperature, while nosignificant difference was found during 9~10 min between the two groups. Our results suggest that 4 ºC pretreatment can inhibit cellpolarity and increase seal rate, but has no effects on membrane currents. It is also suggested that 0~1 min after 4 ºC pretreatment is amore suitable time for electrophysiological recording in neutrophils.
Key words: Neutrophils; polarity; patch-clamp technique
Received: Accepted:
Corresponding author: 邹飞 E-mail: zfei@fimmu.com
Citing This Article：
YUAN Chun-Hua, CAI Chun-Qing, ZOU Fei. Effect of 4 ºC pretreatment on the membrane current and cell polarity in humanneutrophils. Acta Physiol Sin 2006; 58 (5): 494-499 (in Chinese with English abstract).
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