Figure 3

Cell-attached single-channel recordings from identified alveolar epithelial type I cells in a rat lung slice. A(i) A 200 μm rat lung slice immunostained stained with the mVIIIB2 antibody (Alexa Fluor 488 secondary antibody, green emission). (ii) Patch-clamp recording pipette in close proximity to an alveolar epithelial cells whose tip has been filled with TRITC (1:400). (iii) Transmitted light image. (iv) Overlay of images (i) and (ii) plus blue transmitted image to show that the patch recording pipette has formed a gigaohm seal on an VIIIB2-immunopositive alveolar type I cell. B) Exemplar current recording without 10 mM TEA in the pipette to demonstrate the complex nature of the current. Currents were recorded at 0 mV (Vm) and filtered at 2 kHz C) Representative family of cell-attached currents recorded from an alveolar epithelial type I cell, identified by live immunohistochemistry (as in (A)) in an acutely isolated rat lung slice. Single channel activity was recorded in the cell-attached configuration of the patch-clamp technique using a low chloride pipette solution with 10 μM amiloride, niflumic acid and 10 mM TEA. Currents were recorded at the potentials indicated to the left of each trace and were filtered at 200 Hz. L1, L2 and L3 indicate channel open levels 1, 2 and 3, respectively, whilst C indicates the channel closed level. D) Current voltage relationship of the single-channel activity recorded in 3 identified type I cells from 3 separate lung slice preparations. E) NPo (product of open state probability and number of channels) versus voltage plot of the single-channel activity shown in (E).