Validation of confocal laser scanning microscopy for detecting intracellular calcium heterogeneity in liver slices

To investigate changes in the intracellular Ca(2+) ([Ca(2+)]i) in liver lobules under aerobic and hypoxic conditions, we measured [Ca(2+)]i in liver slices using a confocal laser scanning microscope (CLSM). The liver lobule is divided into 3 equal parts between the central vein and portal area, Zones 1, 2, and 3 from the portal side. [Ca(2+)]i in each zone of cultured rat liver lobules was measured by CLSM and a fluorescent Ca(2+) indicator (Rhod 2 AM). After the culture solution was changed to an Na(+)-free solution under aerobic conditions, the percentage of cells showing an increase in [Ca(2+)]i was 66.0+/-9.7% in Zone 1, 70.0+/-10.5% in Zone 2, and 94.0+/-9.7% in Zone 3. The percentage was significantly higher in Zone 3 than in Zones 1 and 2 (P< .01). Under hypoxic conditions, the percentage of cells showing an increase in [Ca(2+)]i was 6.0+/-9.7% in Zone 1, 8.0+/-10.3% in Zone 2, and 10.0+/-10.5% in Zone 3. There were no differences among the 3 zones. In all zones, the percentage was higher under aerobic conditions than under hypoxic conditions (P< .01). These results indicated that the increase in [Ca(2+)]i in liver lobules was heterogeneous. Measurement of [Ca(2+)]i in liver slices by CLSM was considered useful for studying heterogeneity between liver lobules, as well as between liver cells.