EFFECTS OF TRANSIENT OXYGEN-GLUCOSE DEPRIVATION ON G-PROTEINS AND G-PROTEIN-COUPLED RECEPTORS IN RAT CA3 PYRAMIDAL CELLS IN-VITRO

The role of guanosine triphosphate-binding proteins (G-proteins) in th e generation of the outward current during transient oxygen-glucose de privation (OGD) was investigated in CA3 pyramidal cells in rat hippoca mpal organotypic slice cultures using the single-electrode voltage-cla mp technique with KMeSO4-filled microelectrodes. To simulate ischaemia , brief chemical OGD (2 mM 2-deoxyglucose and 3 mM NaN3 for 4-9 min) w as used, which induced an outward K+ current associated with an increa se in input conductance. OGD failed to induce the outward current unde r conditions where G-protein function was disrupted by loading cells w ith guanosine 5'-O-(2-thiodiphosphate) [GDP beta S] or after prolonged injection of guanosine 5'-O-(3-thiotriphosphate) [GTP gamma S]. Howev er, in slices treated with pertussis toxin (PTX), OGD still elicited t he outward current, indicating that PTX-insensitive G-proteins are inv olved. Consistent with this insensitivity to PTX, neither adenosine re ceptors nor GABA(B) (gamma-aminobutyric acid) receptors, which operate via PTX-sensitive G-proteins, mediate the OGD-induced outward current . When adenosine receptors or GABA(B) receptors were blocked with 1,3- dipropyl-8-p-sulphophenylxanthine (DPSPX, 5 mu M) or CGP 52 432 (10 mu M), respectively, the OGD-induced response was not modified. The resp onse also persisted following pretreatment of slice cultures with teta nus toxin to prevent vesicular release of neurotransmitters and neurom odulators from presynaptic terminals. Both PTX-sensitive and PTX-insen sitive G-protein-mediated responses were suppressed during OGD. The in ward current induced by the metabotropic glutamate receptor agonist 1S , 3R-1-aminocyclopentane-1,3-dicarboxylate (1S,3R-ACPD) and the outwar d current elicited by adenosine or baclofen were strongly or completel y attenuated. In contrast, the ionotropic alpha-amino-3-hydroxy-5-meth yl-4-isoxazole propionic acid (AMPA) response was not affected. These findings suggest that during OGD there is a functional uncoupling of r eceptors from G-proteins, and a direct receptor-independent activation of PTX-insensitive G-proteins leading to an increase in membrane K+ c onductance.