Effect of adherence, cell morphology, and lipopolysaccharide on potassium conductance and passive membrane properties of murine macrophage J774.1 cells

Abstract

The effects of adherence, cell morphology, and lipopolysaccharide on electrical membrane properties and on the expression of the inwardly rectifying K conductance in J774.1 cells were investigated. Whole-cell inwardly rectifying K currents (Ki), membrane capacitance (Cm), and membrane potential (Vm) were measured using the patch-clamp technique. Specific Ki conductance (GKi, whole-cell Ki conductance corrected for leak and normalized to membrane capacitance) was measured as a function of time after adherence, and was found to increase almost twofold one day after plating. Membrane potential (Vm) also increased from -42 +/- 4 mV (n = 32) to -58 +/- 2 mV (n = 47) over the same time period. GKi and Vm were correlated with each other; GL (leak conductance normalized to membrane capacitance) and Vm were not. The magnitudes of GKi and Vm 15 min to 2 hr after adherence were unaffected by the presence of 100 microM cycloheximide, but the increase in GKi and Vm that normally occurred between 2 and 8 hr after adherence was abolished by cycloheximide treatment. Membrane properties were analyzed as a function of cell morphology, by dividing cells into three categories ranging from small round cells to large, extremely spread cells. The capacitance of spread cells increased more than twofold within one day after adherence, which indicates that spread cells inserted new membrane. Spread cells had more negative resting membrane potentials than round cells, but GKi and GL were not significantly different. Lipopolysaccharide-(LPS; 1 or 10 micrograms/ml) treated cells showed increased Cm compared to control cells plated for comparable times. In contrast to the effect of adherence, LPS-treated cells exhibited a significantly lower GKi than control cells, indicating that the additional membrane did not have as high a density of functional GKi channels. We conclude that both adherence and LPS treatment increase the total surface membrane area of J774 cells and change the density of Ki channels. In addition, this study demonstrates that membrane area and density of Ki channels can vary independently of one another.