1. The cell volume of suspended C6 glioma cells and primary cultured rat astrocytes was measured at normothermia (37 degrees C), and at mild (32 degrees C) and moderate (27 degrees C) hypothermia by flow cytometry with electrical cell sizing. 2. Under control conditions (37 degrees C), C6 glioma cells had a volume of 809 +/- 29 microm3. Moderate hypothermia (27 degrees C) led to rapid cell swelling, with a maximum volume of 113.1 +/- 1.3 % of control being achieved after 50 min. After rewarming to 37 degrees C, cell volume recovered very slowly and incompletely (to 107.2 +/- 0.4 % of control). Less severe hypothermia (32 degrees C) led to a smaller increase in cell volume (108.7 +/- 0.5 % of control). 3. The maximal cell swelling response and the kinetics of swelling were similar in C6 glioma cells and primary cultured astrocytes. 4. Hypothermia-induced cell swelling was dependent on the presence of extracellular Na+ and was reduced by the Na+-H+ antiporter inhibitor EIPA. 5. The underlying mechanisms of hypothermia-induced cell swelling are an intracellular accumulation of Na+ by (1) differential effects of hypothermia on the membrane permeabilities of Na+ and K+ and (2) activation of the Na+-H+ antiporter by a shift of its activation curve to a more alkaline value.