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Publication Detail
Substrate-dependent changes in mitochondrial function, intracellular free calcium concentration and membrane channels in pancreatic beta-cells.
  • Publication Type:
    Journal article
  • Publication Sub Type:
    Journal Article
  • Authors:
    Duchen MR, Smith PA, Ashcroft FM
  • Publication date:
  • Pagination:
    35, 42
  • Journal:
    Biochem J
  • Volume:
    294 ( Pt 1)
  • Status:
  • Country:
  • Print ISSN:
  • Language:
  • Keywords:
    Animals, Ascorbic Acid, Binding Sites, Calcium, Glucose, In Vitro Techniques, Islets of Langerhans, Male, Membrane Potentials, Mice, Mice, Inbred BALB C, Mitochondria, NAD, Potassium Channels, Spectrometry, Fluorescence, Tetramethylphenylenediamine
Microfluorimetric and patch-clamp techniques have been combined to determine the relationship between changes in mitochondrial metabolism, the activity of KATP channels and changes in intracellular free calcium concentration ([Ca2+]i) in isolated pancreatic beta-cells in response to glucose, ketoisocaproic acid (KIC) and the electron donor couple tetramethyl p-phenylenediamine (TMPD) and ascorbate. Exposure of cells to 20 mM glucose raised NAD(P)H autofluorescence after a delay of 28 +/- 1 s (mean +/- S.E.M., n = 30). The mitochondrial inner membrane potential, delta psi m (monitored using rhodamine 123 fluorescence), hyperpolarized with a latency of 49 +/- 6 s (n = 17), and the [Ca2+]i rose after 129 +/- 13 s (n = 5). The amplitudes of the metabolic changes were graded appropriately with glucose concentration over the range 2.5-20 mM. All variables responded to KIC with shorter latencies: NAD(P)H autofluorescence rose after a delay of 20 +/- 3 s (n = 5) and rhodamine 123 changed after 21 +/- 3 s (n = 6). The electron donor couple, TMPD with ascorbate, rapidly hyperpolarized delta psi m and raised [Ca2+]i. When [Ca2+]i was raised by sustained exposure to 20 mM glucose, TMPD had no further effect. TMPD also decreased whole-cell KATP currents and depolarized the cell membrane, measured with the perforated patch configuration. These data are consistent with a central role for mitochondrial oxidative phosphorylation in coupling changes in glucose concentration with the secretion of insulin.
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