Hypoxia is a major cause of neonatal encephalopathy and seizures, and an increased neuronal excitability may be an underlying mechanism. To determine the role of Ca2+-activated K+ channels in hyperexcitability, we measured large unitary conductance (>200 pS, BKca) currents in symmetrical 140/140 mM K+ using inside-out configuration in CA1 pyramidal cells acutely dissociated from the hippocampus of rats exposed to normoxia or hypoxia (at 10% inspired 02) for 4 weeks after birth. About 53% of the patches contained BKca channels in the normoxic group, but only 20% in the hypoxic one. There were no differences in channel conductance or reversal potential between the groups. Yet, the open probability of BKca channels was much less in hypoxic neurons than that in the control, because of a decrease in channel open time and a prolongation of the closed time. These were partially recovered by an oxidizing but not by reducing agent, suggesting an involvement of redox mechanism. Results indicate that the Ca2+-activated K+ channel activities in hippocampal CA1 neurons are modulated by hypoxia during maturation. The reduction in BKca activity may contribute to hypoxia-induced neuronal hyperexcitability.

Using inside-out configuration of patch clamp techniques, ATP modulation of BKca channels was studied in hippocampal CAI pyramidal neurons of adult rat. Intracellular ATP application markedly increased BKca channel activity, and this ATP-produced increase in BK, channel activity was characterized by a higher opening frequency with no changes in channel open times. In the presence of specific inhibitor against protein kinase A, H-89, ATP did not induce any increase in the channel activity. Furthermore, adding H-89 after addition of ATP reversed the modulation produced by ATE In contrast, protein kinase C inhibitor chelerythrine exerted no apparent effects on ATP-induced channel activation. The present study suggests that BKca channels from hippocampal CA1 pyramidal neurons could be modulated by ATP via a functionally associated orotein kinase A-like orotein.

Redox regulation of BKca channels was studied in CA1 pyramidal neurons of adult rat hippocampus by using inside-out configuration of patch clamp. Intracellular application of oxidizing agent 5, 5-dithio-bis (2-nitrobenzoic acid)(DTNB) markedly increased activity of BKca channels and this stimulating action persisted even after washout. In contrast, the reducing agent dithiothreitol (DTT) had no apparent effects on channel activity but could reverse the pre-exposure of DTNB-induced enhancement. The increase in channel activity produced by DTNB was due to shortened closed time as well as prolonged open time. The effects exerted by another redox couple glutathione disulphide and its reducing form were similar as DTNB and DTT. The present results indicate that BKca channels in CA1 pyramidal neurons can be modulated by intracellular redox potential, and that augmentation of BKca channels by oxidative stress might contribute to the postischemic electrophysiological alterations of CA1 pyramidal neurons.

Buyang Huanwu Decoction (BYHWD), a traditional Chinese medicine, has been developed as a drug to be used for treatment of stroke for hundreds of years. However, the underlying mechanisms remain unknown. In the present study, the effects of BYHWD on delayed neuronal death of hippocampus after transient forebrain ischemia were examined in rats. Transient forebrain ischemia in a duration of 15 rain was induced with the four-vessel occlusion method. BYHWD (per 6.65g/kg) was given orally to rats twice each day for 7 days before ischemia. In BYHWD-pretreated rats, the neuronal injury in the hippocampal CA1 region was significantly less than that of controls. Oral administration of BYHWD also markedly attenuated the number of TUNEL-positive neurons and suppressed the expression of caspase-3p20, a product of catalytically active caspase-3, in the CA1 region. Our results suggest that an inhibition of caspase-3 and apoptosis by BYHWD may oartially account for its neuroorotection against ischemic iniury in the hippocampal CA1 region.