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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:
    15/08/1993
  • Pagination:
    35, 42
  • Journal:
    Biochem J
  • Volume:
    294 ( Pt 1)
  • Status:
    Published
  • Country:
    England
  • Print ISSN:
    0264-6021
  • Language:
    eng
  • 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
Abstract
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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