THE UPTAKE OF ³⁶Cl INTO ASTROCYTES IN TISSUE CULTURE BY A POTASSIUM‐DEPENDENT, SATURABLE PROCESS

Abstract

Abstract— Previous suggestions that the K + ‐dependent uptake of chloride by mammalian cerebral cortex is localized to cortical astrocytes has been investigated in two lines of neural cells in tissue culture. Hamster astrocytes and mouse neuroblastoma cells were each cultured on small glass cover slips in Eagle's (astrocytes) or Dulbecco's (neuroblastoma) supplemented media. At 48 h, cultures were incubated at 37°C for 1–60 min in the presence of 36 Cl plus [ 3 H]inulin (as a measure of extracellular space). In media containing approximately 140 m m chloride, the rate of uptake of 36 Cl by the astrocytes was a function of the concentration of K + (range 5–54 m m ) in the medium, and the uptake was characterized by saturation kinetics and an apparent K m of 38·5 m m . The uptake was temperature and apparently energy dependent, significantly inhibited by 5 or 10 m m Br ‐ , I ‐ , SCN ‐ or ClO 4 ‐ , and competitively inhibited by 10 m m acetazolamide ( K i = 27·1 m m ). None of these characteristics were observed with neuroblastoma cultures studied under similar conditions. In chloride‐free media, the cellular K/Na ratio of the astrocytes shifted from the usual value of 4·55 to a value of 0·29, the culture medium became more alkaline than normal, and the cells spontaneously sloughed from the cover slips but remained normally viable. Our observations are the first to provide direct support for previous suggestions that there is a mediated, K + ‐dependent coupled cation, chloride and fluid uptake by mammalian cerebrocortical glia and that this uptake is an enzymatically catalysed process. The observations have been discussed in terms of a presumed central role for astrocytes in modulating the external ionic milieu of the neurons they surround and in terms of implications for epilepsy, stroke and cortical edema.