Severe Preeclampsia Uterine Artery Smooth Muscle Cells Large Conductance Changes In Calcium Activated Potassium Channels

objective： Severe preeclampsia belongs to the category ofpregnancy hypertension disease. The continuous increasing tension ofresistance vessel is the fundamental pathological features of gestationalhypertension disease, the tension of peripheral vascular resistance increased,and the ion channels on vascular smooth muscle cells (VSMCs) are involvedin the regulation of vascular tension. It is necessary to investigate Whether theBKCachannels change under the Severe preeclampsia condition.So thisexperiment using the patch clamp technique from the level of ion channelresearch whether severe preeclampsia is associated with BKCachannel change,provide a theoretical basis for the pathogenesis of the disease and prevention.Methods: We collected the samples from the patient who deliveried in ourhospital from June2012to March2012,not associated with othercomplications. The need for cesarean section surgery were divided into twogroups: The severe preeclampsia late pregnancy8cases, The normal latepregnancy8cases. Muscle tissues from the uterine section which containsabundant blood sinus, separation of uterine arterial smooth muscle cells in twosteps of acute enzyme.Select the cell membrane integrity, smooth, refractionand better smooth muscle cell experiment. patch clamp single channelrecording technique:The current was recorded on single myocyte in symmetrichigh potassium solution by patch clamp single channel recording technique. Single channel current was amplified and filtered by patch clamp amplifier(CEZ-2200), then imported into the computer, pClamp6.0software was usedto record the digital data and Clampfit10.1software was used to analyze data.Compare the change of BKCacurrent signal in the uterine artery smoothmuscle cells between the pregnant woman In normal pregnancy and severepreeclampsia. Results:1. Acute enzyme separation of Uterine arterialsmooth muscle cells: The single uterine arterial smooth muscle cell whichwas dissociated by enzyme solution can maintain its physiological functions,and it can be used in patch clamp experiments2. The properties of singlechannel currents of BKCachannels in Uterine arterial smooth muscle cellswere as follows:(1)Single channel currents of BKCawere recorded ininside-out patch. The value of single channel conductance was207.2±12.7pS.(2)The channels had distinct voltage dependent and calcium dependentcharacteristics.(3)In outside-out patch (Vm=+30mV),the activation ofBKCa channel could be blocked by200nM IbTX completely.3. comperedthe single channel currents of the normotensive pregnant group BKCachannelwith the severe preeclampsia group: in cell-attached patch (Vm=+40mV),the The mean open probability of the Normotensive pregnant group was0.007±0.0017, The amplitude of voltage was7.627±0.727pA, the mean opentime was7.216±1.003ms ms, the mean close time was879.445±149.716ms，the The mean open probability of the Severe preeclampsia group was0.0017±0.0005, The amplitude of voltage was4.399±0.464pA, the mean open time was1.642±0.348ms, the mean close time was930.791±264.157ms.compered with the Normotensive pregnant group,the the mean open time andthe amplitude of voltage of the Severe preeclampsia group significantlydecreased,the to of the Severe preeclampsia group significantly decreased also.the mean close time of the Severe preeclampsia group No significantdecreased. Conclusion:1. This experiment adopts the two-step acute enzymeseparation to dissociated pregnancy uterine arterioles, then we got the smoothmuscle cells which were suitable for the perforated patch clamp experiment.2.Large-conductance Ca2+-activated potassium channels (BKCa) are highlyexpressed in the smooth muscle cells which isolated from small uterus artery,the BKCachannel had the characteristics of large conductance value, calciumdependented and voltage dependented. the BKCachannel can be blocked bythe specificity inhibitor.3. In cell-attached patch, compared with the normalthird trimester pregnant women, the activity of BKCachannel in uterinearteriolar smooth muscle cells was reduced in the patients with severepreeclampsia, This suggests that the significant reduction in BKCachannelexpression had taken part in the occurrence of severe preeclampsia.