Amiloride-sensitive Na+ channels in fetal type II pneumocytes are regulated by G proteins

We have used the patch-clamp technique to record single channels in excised membrane patches from type II pneumocytes freshly isolated from fetal guinea pig lung by elastase digestion and differential filtration. The 10/56 patches exhibited spontaneous channel activity with a mean open-state probability (NPo) of 0.5 +/- 0.1. In symmetrical Na(+)-rich solutions, the channels had a unitary conductance of 11.1 +/- 0.5 pS and showed current reversal at approximately 0 mV. Superfusing the inner membrane leaflet of the patch with a K(+)-rich solution resulted in single-channel current activity with a conductance of 5.6 +/- 0.2 pS being resolved. Current reversed at +22.1 +/- 1.9 mV, which is compatible with a PNa+/PK+ of 1.8 +/- 0.1. The addition of 0.1 mM guanosine 5'-O-(3-thiotriphosphate) to the cytoplasmic face of the patch elicited channel activity in 12/31 previously quiescent patches, whereas, in spontaneously active patches, channel NPo was increased. Amiloride, in the concentration range 0.4-4 microM, reduced the frequency of observed spontaneous (or activatable) channel activity, reduced NPo, and induced flickery channel behavior. No activity was seen in the presence of 10 microM amiloride in the pipette. This is the first direct observation of a G protein regulated Na(+)-conductive pathway in alveolar epithelium, and it may represent one route by which the alveolar epithelium of the fetus can regulate the Na(+)-driven fluid reabsorption necessary for the adaptation of the newborn lung to air breathing at birth.