3-nitropropionic acid increases the intracellular Ca2+ in cultured astrocytes by reverse operation of the Na+-Ca2+ exchanger

The effect of 3-nitropropionic acid (3-NPA) on intracellular calcium ([Ca2+](i)) in cultured cortical and striatal astrocytes was examined using a calcium imaging technique with fura-2. 3-NPA (1.7 mM) increased the [Ca2+](i) in cortical and striatal astrocytes. The latency for the onset of the rise in [Ca2+](i) in cortical astrocytes was 22.7 +/- 0.8 min and was significantly longer in striatal astrocytes (39.2 +/- 2.4 min; P < 0.001). The maximal increase in [Ca2+](i) for both astrocytes was seen after 50 min and remained at that level even after extensive washing, The maximal responses were about 125 and 140% of the initial values for striatal and cortical cells, respectively. Pretreatment (2-3 h) with creatine (25 mM) significantly delayed the onset of increase in [Ca2+](i) by 3-NPA in cortical (39.8 +/- 3.7 min) and in striatal (57.8 +/- 2.5 min) astrocytes from the respective untreated cells (P < 0.05). However, the [Ca2+](i) increase was similar to that of the untreated cells at 60 or 90 min. The increase in [Ca2+](i) by 3-NPA was not observed in Ca2+-free or low-Na+ medium, but there was rather a 10-15% decrease under the Ca2+-free condition (P < 0.05), Superfusion of normal Ca2+ (2 nM) medium after exposure of the cells to 3-NPA in Ca2+-free medium increased the [Ca2+](i) dramatically and reversibly. This increase was significantly attenuated in the presence of 2',4'-dichlorobenzamil (100 mu M), nifedipine (10 mu M), or Ni2+ (2 mM) or after exposure to amiloride (1 mM). The blockade was total ill the case of 2',4'-dichlorobenzamil, The results indicate that the 3-NPA-induced increase in [Ca2+](i) in astrocytes was due to an influx of Ca2+ by the reverse operation of the Na+-Ca2+ exchanger system, which may be responsible for the gliotoxic action of 3-NPA. (C) 1997 Academic Press.