INTRACELLULAR CALCIUM STORES AND INOSITOL 1,4,5-TRISPHOSPHATE RECEPTOR IN RAT-LIVER CELLS

The D-myo-inositol 1,4,5-trisphosphate [Ins(1,4,5)P-3] receptor was lo calized by immunofluorescence experiments in situ in liver cryosection s. Two anti-Ins(1,4,5)P, receptor antibodies (against the 14 C-termina l residues of the type 1 receptor or against the entire cerebellar rec eptor) weakly decorated the whole cytoplasm, and a more intense labell ing was observed at the periphery of the hepatocytes, particularly ben eath the canalicular and the sinusoidal domains of the plasma membrane (PM). Antibodies against calreticulin, the Ca2+ pump (SERCA2b) or end oplasmic reticulum (ER) membranes homogeneously labelled the cytoplasm and the subplasmalemmal area. These data indicate that the ER can be divided into at least two specialized subregions: one is located throu ghout most of the cytoplasm and contains markers of the rough ER (RER) , calreticulin, SERCA2b and a low density of Ins(1,4,5)P-3 receptor, a nd the other is confined to the periphery of the cells and contains ca lreticulin, Ca2+ pump, RER markers and a high density of Ins(1,4,5)P-3 receptor. A membrane fraction enriched in Ins(1,4,5)P-3 receptor and in markers of the PM was immune-adsorbed with the antibody against the C-terminal end of the Ins(1,4,5)P-3 receptor and pelleted with Sephar ose protein A. The immune-isolated material was enriched in Ins(1,4,5) P-3 receptor, but none of the markers of the ER or of the PM could be detected. This suggests that the Ins(1,4,5)P-3 receptor is localized o n discrete domains of the ER membrane beneath the canalicular and the sinusoidal membranes, where it was found at higher densities than the other markers.