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Keywords:

  • adenosine;
  • hippocampal slice cultures;
  • ischaemia;
  • metabotropic;
  • potassium current

Abstract

The role of guanosine triphosphate-binding proteins (G-proteins) in the generation of the outward current during transient oxygen-glucose deprivation (OGD) was investigated in CA3 pyramidal cells in rat hippocampal organotypic slice cultures using the single-electrode voltage-clamp technique with KMeSO4-filled microelectrodes. To simulate ischaemia, brief chemical OGD (2 mm 2-deoxyglucose and 3 mm NaN3 for

4–9 min) was used, which induced an outward K+ current associated with an increase in input conductance. OGD failed to induce the outward current under conditions where G-protein function was disrupted by loading cells with guanosine 5′-O-(2-thiodiphosphate) [GDPβS] or after prolonged injection of guanosine 5′-O-(3-thiotriphosphate) [GTPγS]. However, in slices treated with pertussis toxin (PTX), OGD still elicited the outward current, indicating that PTX-insensitive G-proteins are involved. Consistent with this insensitivity to PTX, neither adenosine receptors nor GABAB (γ-aminobutyric acid) receptors, which operate via PTX-sensitive G-proteins, mediate the OGD-induced outward current. When adenosine receptors or GABAB receptors were blocked with 1,3-dipropyl-8-p-sulphophenylxanthine (DPSPX, 5 μm) or CGP 52 432 (10 μm), respectively, the OGD-induced response was not modified. The response also persisted following pretreatment of slice cultures with tetanus toxin to prevent vesicular release of neurotransmitters and neuromodulators from presynaptic terminals.

 Both PTX-sensitive and PTX-insensitive G-protein-mediated responses were suppressed during OGD. The inward current induced by the metabotropic glutamate receptor agonist 1S, 3R-1-aminocyclopentane-1,3-dicarboxylate (1S,3R-ACPD) and the outward current elicited by adenosine or baclofen were strongly or completely attenuated. In contrast, the ionotropic α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) response was not affected. These findings suggest that during OGD there is a functional uncoupling of receptors from G-proteins, and a direct receptor-independent activation of PTX-insensitive G-proteins leading to an increase in membrane K+ conductance.