Polarographic measurement of oxygen uptake by astrocytes in primary cultures using the tissue-culture flask as the respirometer chamber

Abstract

Oxygen uptake was measured in primary cultures of astrocytes from the brain hemispheres of newborn DBA mice by the aid of an oxygen electrode inserted directly into the culture flastk, i.e. using the flasks, completely filled with MEM medium, as the respirometer chamber. The respiration was inibitally intense (300 μmol per hr per 100 mg protein) but delined somewhat during the 6 hr of measurement, probably due to a depletion of intermediary metabolites released to the large surplus of medium. The respiratory rates were approximately identical in the presence of a CO₂/ bicarbonate and a HEPES buffer. Exposure to a high concentration of potassium led to a transient stimulation of the oxygen uptake of almost 100%, a response that was very easily observed using the present method. Since no mechanical damage was inflicted upon the cells, culturing could be continued, if so desired, after the measurement.