Ca2+ and electrolyte mobilization following agonist application to the pancreatic beta cell line HIT.
We have investigated intracellular Ca2+ mobilization in oscillations of cytoplasmic Ca2+ in response to glucagon-like peptide 1 (GLP-1) and glucose in clonal HIT insulinoma cells with a confocal laser-scanning microscope (CLSM). We also used electron probe X-ray microanalysis to determine the GLP-1- and glucose-induced changes in electrolyte levels in the cytoplasm and insulin granules of the cells. GLP-1 produced 10- to 35-s duration oscillations in cytoplasmic Ca2+ concentration ([Ca2+]i), both with and without Ca2+ in the extracellular solution, suggesting that Ca2+ is mobilized from intracellular Ca2+ stores, namely secretory granules. Glucose caused 1- to 3-min duration oscillatory increases in [Ca2+]i when the extracellular solution contained Ca2+. When the cells were cultured without Ca2+ (no Ca2+ added, 1 mM EGTA), an oscillatory [Ca2+]i increase of amplitude and short duration (12-35 s) was produced by 11 mM glucose, and the oscillation was inhibited by ruthenium red. X-ray microanalysis showed that stimulation with glucose increased the total Ca concentration in the cytoplasm and decreased it in the insulin granules with and without Ca2+ in the extracellular solution. The application of glucose significantly decreased K, and increased Na and C1 in the cytoplasm when the extracellular solution contained Ca2+. Our result also suggests that the [Ca2+]i oscillation induced by glucose is involved in the release of Ca2+ from intracellular Ca2+ stores through the ryanodine receptor, which is blocked by ruthenium red, and/or through the inositol trisphosphate receptor that may be present in the membrane of insulin granules.
Nakagaki
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I
Department of Physiology, Hyogo College of Medicine, Nishinomiya, Hyogo, Japan. ph1004@hyo-med.ac.jp
Sasaki
S
S
Hori
S
S
Kondo
H
H
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D003593:Cytoplasm / Q000378:metabolism; D003594:Cytoplasmic Granules / Q000378:metabolism; D004573:Electrolytes / Q000378:metabolism*; D004577:Electron Probe Microanalysis; D005110:Extracellular Space / Q000378:metabolism; D005934:Glucagon / Q000494:pharmacology*; D052216:Glucagon-Like Peptide 1; D007340:Insulinoma; D007515:Islets of Langerhans / Q000187:drug effects* / Q000378:metabolism* / Q000648:ultrastructure; D018613:Microscopy, Confocal; D008854:Microscopy, Electron; D010190:Pancreatic Neoplasms; D010446:Peptide Fragments / Q000494:pharmacology*; D011498:Protein Precursors / Q000494:pharmacology*; D014407:Tumor Cells, Cultured
D004573:Electrolytes; D010446:Peptide Fragments; D011498:Protein Precursors; D052216:Glucagon-Like Peptide 1; D005934:Glucagon; D002118:Calcium