Characterization of a delayed rectifier K+ channel in NG108-15 neuroblastoma X glioma cells: gating kinetics and the effects of enrichment of membrane phospholipids with arachidonic acid

Abstract

A voltage-sensitive K+ channel with characteristics of the delayed rectifier was studied in NG108-15 cells using the cell-attached patch-clamp technique. The primary conductance of the channel was 18 pS, but occasional openings to a subconductance state were observed. The average latency to first opening of the channel was about 4 msec. Based on about 20,000 channel openings, the open time probability density function (pdf) required at least three exponentials (time constants of about 0.2, 3 and 9 msec) to achieve an adequate fit to the data. The closed time pdf required at least six exponentials to describe the data (time constants ranging from 0.093 to 440 msec). Thus, the channel exists in at least three open and six closed states. The ensemble average describing the inactivation of the channel was well fit by two exponentials with time constants of 170 msec and 4.2 sec. To examine the effect of changes in membrane lipid composition on the properties of the channel, the phospholipids of the cells were enriched with polyunsaturated fatty acids. In patches from 20:4-enriched cells the conductance, mean first latency, and open-time pdf were similar to control cells. However, the open state probability was increased from 0.25 to 0.44 and the mean closed time was decreased from 20 to 9 msec. The closed time pdf exhibited a higher proportion of closing events associated with short time constants, i.e., the probability of the channel closing into a long-lived closed state was decreased. The decay phase of the ensemble average also was changed; the proportion of the curve described by the slower time constant was almost doubled. Thus, the delayed rectifier from NG108-15 cells can exist in at least three open and six closed states, and changes in membrane lipid composition may have subtle effects on the gating kinetics of the channel.