Lollike K, Borregaard N, Lindau M
Department of Internal Medicine and Haematology L, National University Hospital, Copenhagen, Denmark.
J Cell Biol. 1995 Apr;129(1):99-104. doi: 10.1083/jcb.129.1.99.
We measured capacitance changes in cell attached patches of human neutrophils using a high frequency lock-in method. With this technique the noise level is reduced to 0.025 fF such that capacitance steps of 0.1 fF are clearly detected corresponding to exo- and endocytosis of single 60 nm vesicles. It is thus possible to detect almost all known exocytotic and endocytotic processes including exocytosis of small neurotransmitter containing vesicles in most cell types as well as endocytosis of coated and uncoated pits. In neutrophils we demonstrate a stepwise capacitance decrease generated by 60-165 nm vesicles as expected for endocytosis of coated and non-coated pits. Following ionomycin stimulation a stepwise capacitance increase is observed consisting of 0.1-5 fF steps corresponding to the different granule types of human neutrophils from secretory vesicles to azurophil granules. The opening of individual fusion pores is resolved during exocytosis of 200 nm vesicles. The initial conductance has a mean value of 150 pS and can be as low as 35 pS which is similar to the conductance of many ion channels suggesting that the initial fusion pore is formed by a protein complex.
我们使用高频锁相法测量了人中性粒细胞贴壁膜片的电容变化。通过这种技术，噪声水平降低到0.025 fF，从而能够清晰检测到0.1 fF的电容阶跃，这对应于单个60 nm囊泡的胞吐和胞吞作用。因此，几乎可以检测到所有已知的胞吐和胞吞过程，包括大多数细胞类型中含小神经递质囊泡的胞吐作用以及有被小窝和无被小窝的胞吞作用。在中性粒细胞中，我们证明了60 - 165 nm囊泡产生的电容逐步下降，这与有被小窝和无被小窝的胞吞作用预期相符。离子霉素刺激后，观察到电容逐步增加，由0.1 - 5 fF的阶跃组成，对应于人中性粒细胞从分泌囊泡到嗜天青颗粒的不同颗粒类型。在200 nm囊泡的胞吐过程中分辨出了单个融合孔的开放。初始电导的平均值为150 pS，最低可达35 pS，这与许多离子通道的电导相似，表明初始融合孔是由蛋白质复合体形成的。
