Plasma Membrane Activities Retained after Lethal Heat Shock

Abstract

Cultures of Chinese hamster ovary (CHO) cells were examined to determine if heat killing could be attributed to severe damage in the plasma membrane. Three independent transport activities of the plasma membrane were measured. Glucose transport into the cells (measured with the non-metabolizable analogue 3-O-methyl-d-glucose) was stimulated rather than inhibited by heat. Most of the stimulation was found after non-toxic heat doses. Although amino acid transport (measured with the non-metabolizable analogue 2-aminoisobutyric acid) was slightly inhibited by heat, heat-sterilized cells were able to accumulate high intracellular concentrations. Cellular uptake of the nucleoside uridine was unaffected for at least 4 h after heating. In contrast, its incorporation into RNA was immediately inhibited. To further study plasma membrane damage, cells were either heated or treated with drugs which localize to the plasma membrane, ionophore A23187 or amphotericin B. The mode of cell killing by heat was radically different from that of the two drugs: heat-sterilized cells retained a phase-bright morphology and excluded the viability dye trypan blue while drug-killed cells rapidly became phase-dark and absorbed the dye. These results add to a growing list of plasma membrane activities which are retained in heat-sterilized cells, and suggest that the initial thermal damage responsible for cell killing is at an alternate site(s).