Histochemical and physiological study of delayd neuronal death

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1.Making plasmamembrane Ca^<2+>-ATPase antibody, immunohistochemistryUsing Ca^<2+>-ATPase type II antibody we made a immunohistochemical mapping of that enzyme in normal rat and gerbil brain. In hippocampus, the distribution of the enzyme is similar to that one which we have detected enzymehistochemically. That is, the enzyme is diffusely located on the plasma membrane of pyramidal neurons, axon, dendrite. There is no differences in distributional density between CA1, CA3 and dentate gyrus.2.Physiological study of gerbil hippocampal slicesWe studied N-methyl-D-aspartate (NMDA) receptor-mediated synaptic potentials in CA1 pyramidal neurons using hippocampal slices of the gerbils after transient forebrain ischemia. In the presence of 6-cyanc 7-nitroquinoxaline-2,3-dione (CNQX) and bicucullin, stimulation on Schaffer collateral/commissural fibers induced field excitatory postsynaptic potentials (fEPSP) activated by NMDA receptors. We found that in many slices after ischemia, low frequency … More stimulation (0.1-10Hz) to input fibers caused repeated depression and potentiation of the NMDA-mediated fEPSP.The cyclic changes in fEPSP amplitude were dependent on stimulus frequency, ranging from 0.08 to 2.5 cycle/min. The cyclic changes were blocked by application of 1 bis (o-aminophenoxy) ethane-N,N,N', N'-tetraacety1, tetraacetoxymethy1 ester (BAPTA-AM), a membrane permeable Ca^<2+> chelator, but they were little affected by application of vera-pamil or by reducing Ca^<2+>in bathing solution. Intracellular recordings showed periodic depolarizations of membrane potential synchroniz with depression of EPSP.The cyclic phenomenon was significantly attenuated by application of 1-(5-soquinolinylsulfony1)-2-methylpiperazine (H-7) and K252a, protein kinase C (PKC) antagonist.These results suggest that stimulus dependent NMDA-receptor activation, medi-ated by PKC,takes place the postischemic CA1 neurons and the cyclic change may reflect abnormal intracellular Ca^<2+> signaling process towards neuronal degeneration re-sulted in periodic membrane depolarization.3.Potential mapping of the gerbil hippocampus stimulated on the contralateral commisural fibersWe made potential mapping of postischemic gerbil hippocampus by recording EPSP induced by contralateral commisural fiber stimulation. We revealed that CA1 pyramidal neurons are in the hyper excitatory state in the early stage (2-8h) following ischemic insult and LTP is significantly increased in that period compared with the non-ischemic group. This is the first report of the abnormal physiological conditions in the early postischemic period in vivo. Less