RESUMEN
Objective To observe dynamic changes of Kir2. 3 mRNA in the hippocampus of rats with chronic temporal lobe epilepsy, and to discuss the relationship between Kir2. 3 expression and the pathogenesis of chronic temporal lobe epilepsy. Methods We used pilocarpine to induce status epilepticus (SE) in rats,which became chronic temporal lobe epileptic rats in 2 weeks. The expression of Kir2.3 mRNA was measured by reverse transcription polymerase chain reaction (RT-PCR) at the time points of 0, 6, 72 hours and 2 weeks after SE. Results The ratios of Kir2. 3 mRNA to β-actin of normal control and 0, 6, 72 hours, 2 weeks after SE were 0. 080 ± 0. 030, 0. 103 ± 0. 045, 0. 164 ± 0. 026, 0. 132 ± 0. 024, and 0. 011 ± 0. 008, respectively. The ratio was significantly higher 6 and 72 hours after SE and significantly lower 2 weeks after SE than that of the normal control. Conclusions Two weeks after SE, when the rats had spontaneous recurrent seizures, the expression rate of Kir2.3 reached a turning point, which possibly became the basis of epileptogenesis.
RESUMEN
Aim To study the regulatory effects of PMA,a PKC activator,on Kir 2.3 channel function expressed in Xenopus oocytes and COS-7 cells,and PIP2 involvement in these regulations.Methods Kir 2.3 channel was expressed in Xenopus oocytes and COS-7 cell by RNA microinjection and DNA transfection using calcium phosphate precipitate,respectively.Two-electrode-voltage-clamp and whole-cell patch clamp were used to record the Kir 2.3 current in Xenopus oocytes and COS-7 cell.The PIP2 hydrolysis was detected by confocal microscopy.Results PMA significantly inhibited Kir 2.3 current in Xenopus oocytes.But PMA had no effect on the Kir 2.3 current expressed in COS-7 cell,in which activation of M1 receptor,however,induced a significant inhibition of Kir 2.3 current.It was reported recently that PMA could trigger the PIP2 hydrolysis in membrane of oocytes.Thus PKC inhibition of Kir 2.3 current seen in oocytes could be the result of PIP2 hydrolysis.Following the same line,the inability of PKC inhibition of Kir 2.3 current seen in COS-7 cells would suggest PKC could not induce PIP2 hydrolysis in these cells. This hypothesis was tested by monitoring the PIP2 level in COS-7 cell membrane by confocal microscopy.Dynamic changes in membrane PIP2 level were imaged using GFP fluorescence signal that had been tagged to the PLC?1PH domain known to be able to bind PIP2 specifically. There was no significant change of PIP2 level on COS-7 cell membrane after longtime treatment of PMA,whereas again,the activation of M1 receptor by ACh induced a significant change in the PIP2 level.These results were in perfect agreement with the electrophysiological results.Conclusions PMA,through activation of PKC,inhibited Kir 2.3 current expressed in Xenopus oocytes but not in COS-7 cells.Similarly PMA induced significant reduction in membrane PIP2 level in Xenopus oocytes but not in COS-7 cells. PIP2 hydrolysis plays an important role in PKC-induced inhibition of the Kir channel currents.