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Oxidative stress activates a novel non-selective cation channel in insulin-secreting cells

Single channel recordings from CRI-G1 insulin-secreting cells were used to characterize a novel ion channel. The presence of both Ca<sup>2+</sup> and β-NAD<sup>+</sup> at the cytoplasmic aspect of the membrane are required for channel activity. This is the first ion channel described which requires internal β-NAD<sup>+</sup> for activity (thus termed NS<sub>NAD</sub>). The channel was found to be permeable to all monovalent (Na<sup>+</sup>, K<sup>+ </sup>and Cs<sup>+</sup>) and divalent cations tested (Ca<sup>2+</sup>, Mg<sup>2+</sup>, Ba<sup>2+</sup>, and Mn<sup>2+</sup>). The slope conductance is relatively large (70 - 90pS) compared to other non-selective cation channels and also has extremely slow kinetics (open and closed times in the range of seconds). Whole-cell voltage clamp experiments illustrate that internal β-NAD<sup>+</sup> activates a cation current consistent with activation of the NS<sub>NAD</sub> channel. Similar to the single NS<sub>NAD</sub> channel, the β-NAD<sup>+</sup>-activated current was sensitive to the internal concentrations of both Ca<sup>2+</sup> and β-NAD<sup>+</sup>. The non-selective nature of this cation current was confirmed by replacement of the internal K<sup>+</sup> with Cs<sup>+</sup> which did not diminish the β-NAD<sup>+</sup>-activated current. Additionally, replacement of external cations with the impermeant NMDG abolished the β-NAD<sup>+</sup>-activated current. The diabetogenic agent alloxan was found to irreversibly depolarize CRI-GI cells by opening a non-selective cation channel with characteristics similar to the NS<sub>NAD</sub> channel. The channel activated by alloxan is characterized by a slope conductance of approximately 70 pS and very slow (seconds) kinetics. Channel activity is lost upon excision of the patch, but can be re-activated by the application of internal β-NAD<sup>+</sup>. The mechanism of alloxan-induced depolarization and channel activation appears to be through the production of reactive oxygen species (ROS). This data indicates that oxidative stress generated by both alloxan and H<sub>2</sub>O<sub>2</sub> causes the activation of the NS<sub>NAD</sub> channel which results in irreversible collapse of the membrane potential and massive Ca<sup>+</sup> influx leading to eventual cell death. This may represent a component of the destruction of pancreatic β-cells during type I diabetes and possibly other pathologies in which oxidative stress is implicated.

Identiferoai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:265376
Date January 1998
CreatorsHerson, Paco S.
PublisherUniversity of Aberdeen
Source SetsEthos UK
Detected LanguageEnglish
TypeElectronic Thesis or Dissertation

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