It has been reported previously that the neuronal excitability persistently suppresses and the amplitude of fast afterhyperpolarization (fAHP) increases in CAI pyramidal cells of rat hippocampus following transient forebrain ischemia. To understand the conductance mechanisms underlying these post-ischemic electrophysiological alterations, we compared differences in activities of large conductance Ca2+-activated potassium (BKca) channels in CA1 pyramidal cells acutely dissociated from hippocampus before and after ischemia by using inside-out configuration of patch clamp techniques. (1) The unitary conductance of BKc. channels in post-iscfiemic neurons (295pS) was higher than that in control neurons (245pS) in symmetrical 140/140mM K+ in inside-out patch; (2) the membrane depolarization for an e-fold increase in open probability (P) showed no significant differences between two groups while the membrane potential required to produce one-half of the maximum Po was more negative after ischemia, indicating no obvious changes in channel voltage dependence; (3) the [Ca2+] required to half activate BKca, channels was only 1uM in post-ischemic whereas 2 IxM in control neurons, indicating an increase in [Ca2+], sensitivity after ischemia; and (4) BKca channels had a longer open time and a shorter closed time after ischemia without significant differences in open frequency as compared to control. The present results indicate that enhanced activity of BKc. channels in CAI pyramidal neurons after ischemia may partially contribute to the post-ischemic decrease in neuronal excitability and increase in fAHR.

Pyramidal neurons in the CAI field of the hippocampus die a few days after transient cerebral ischemia.4,21 Excessive excitatory synaptic activation following reperfusion is thought to be responsible for such de-layed cell death. 4,24 However, it remains controversial whether excitatory synaptic transmission in the CAI field is increased following reperfusion.2'3'1'1u18'26 Here we report a novel postsynaptic potential evoked from CAI pyramidal neurons preceding cell death after transient forebrain ischemia with intracellular record-ing and staining techniques in vivo. This result indicates the dramatic alteration of synaptic transmission in CAI neurons after transient ischemia. The ischemia-induced postsynaptic potential may be associated with the postischemic neuronal injury. Copyright

The present study examined temporal changes in activity of large conductance, Ca2+-activated potassium (BKca) channels in postischemic CA1 pyramidal neurons at 2, 6, 24 and 48h after reperfusion. These changes in activity and possible cellular mechanisms were examined using the inside-out configuration of patch clamp. The unitary conductance of postischemic BKca channels increased transiently to 1.9% of the control at 2h after reperfusion, and recovered to the control level thereafter. A persistent increase in [Ca2+]i sensitivity of BKca channels was observed in postischemic CA1 neurons with the maximal sensitivity to [Ca2+]i at 6h after reperfusion while channel voltage- dependence showed no obvious changes. Kinetic analyses showed that the postischemic enhancement of BKca channel activity was due to longer open times and shorter closed times as there was no significant changes in opening frequency after ischemia. Glutathione disulphide markedly increased BKca channel activity in normal CA1 neurons, while reducing glutathione caused a decrease in BKca channel activity by reducing the sensitivity of this channel to [Ca2+]i in postischemic CA1 neurons. Similar modulatory effects on postischemic BKca channels were also observed with another redox couple, DTNB and DTT, suggesting an oxidation modulation of BKca channel function after ischemia. The present results indicate that a persistent enhancement in activity of BKca channels, probably via oxidation of channels, in postischemic CA1 pyramidal neurons may account for the decrease in neuronal excitability and increase in fAHP after ischemia. The ischemia-induced augmentation in BKca channel activity may be also associated with the postischemic neuronal injury.

Evoked postsynaptic potentials of CA1 pyramidal neurons in rat hippocampus were studied during 48h after severe ischemic insult using in vivo intracellular recording and staining techniques. Postischemic CAI neurons displayed one of three distinct response patterns following contralateral commissural stimulation. At early recirculation times (0-12h) approximately 50% of neurons exhibited, in addition to the initial excitatory postsynaptic potential, alate depolarizing postsynaptic potential lasting for more than 100ms. Application of dizocilpine maleate reduced the amplitude of late depolarizing postsynaptic potential by 60% Other CAI neurons recorded in this interval failed to develop late depolarizing postsynaptic potentials but showed a modest blunting of initial excitatory postsynaptic potentials (non-late depolarizing postsynaptic potential neuron). The proportion of recorded neurons with late depolarizing postsynaptic potential characteristics increased to more than 70% during 13-24h after reperfusion. Beyond 24h reperfusion, 20% of CA1 neurons exhibited very small excitatory postsynaptic potentials even with maximal stimulus intensity. The slope of the initial excitatory postsynaptic potentials in late depolarizing postsynaptic potential neurons increased to 150% of control values up to 12h after reperfusion indicating a prolonged enhancement of synaptic transmission. In contrast, the slope of the initial excitatory postsynaptic potentials in non-late depolarizing postsynaptic potential neurons decreased to less than 50% of preischemic values up to 24h after reperfusion indicating a prolonged depression of synaptic transmission. More late depolarizing postsynaptic potential neurons were located in the medial portion of CA1 zone where neurons are more vulnerable to ischemia whereas more non-late depolarizing postsynaptic potential neurons were located in the lateral portion of CA1 zone where neurons are more resistant to ischemia. The result from the present study suggests that late depolarizing postsynaptic potential and small excitatory postsynaptic potential neurons may be irreversibly injured while non-late depolarizing postsynaptic potential neurons may be those that survive the ischemic insult. Alterations of synaptic transmission may be associated with the pathogenesis of postischemic neuronal injury.

We have previously identified three distinct populations of CA1 pyramidal neurons after reperfusion based on differences in synaptic response, and named these late depolarizing postsynaptic potential neurons (enhanced synaptic transmission), non-late depolarizing postsynaptic potential and small excitatory postsynaptic neurons (depressed synaptic transmission). In the present study, spontaneous activity and membrane properties of CA I neurons were examined up to 48 h following-14 min ischemic depolarization using intracellular recording and staining techniques in vivo. In comparison with preis-chemic properties, the spontaneous firing rate and the spontaneous synaptic activity of CA1 neurons decreased significantly during reperfusion; spontaneous synaptic activity ceased completely 36-48h alter reperfusion, except for a low level of activity which persisted in non-late depolarizing postsynaptic potential neurons. Neuronal hyperactivity as indicated by increasing firing rate was never observed in the present study. The membrane input resistance and time constant decreased significantly in late depolariz-ing postsynaptic potential neurons at 24-48h reperfusion. In contrast, similar changes were not observed in non-late depolarizing postsynaptic potential neurons. The rheobase, spike threshold and spike frequency adaptation in late depolarizing postsynaptic potential neurons increased progressively following reperfu-sion. Only a transient increase in rheobase and spike threshold was detected in non-late depolarizing postsynaptic potential neurons and spike frequency adaptation remained unchanged in these neurons. The amplitude of fast afterhy perpolarization increased in all neurons after reperfusion, with the smallest increment in non-late depolarizing postsynaptic potential neurons. Small excitatory postsynaptic potential neurons shared similar changes to those of late depolarizing postsynaptic potential neurons. These results suggest that the enhancement and depression of synaptic transmission following ischemia are probably due to changes in synaptic efficacy rather than changes in intrinsic membrane properties. The neurons with enhanced synaptic transmission following ischemia are probably the degenerating neurons, while the neurons with depressed synaptic transmission may survive the ischemic insult.