As a noninvasive neuromodulation technique, transcranial magnetic stimulation (TMS) is widely used in the clinical treatment of neurological and psychiatric diseases, but the mechanism of its action is still unclear. The purpose of this paper is to investigate the effects of different frequencies of magnetic stimulation (MS) on neuronal excitability and voltage-gated potassium channels in the in vitro brain slices from the electrophysiological perspective of neurons. The experiment was divided into stimulus groups and control group, and acute isolated mice brain slices were applied to MS with the same intensity (0.3 T) at different frequencies (20 Hz and 0.5 Hz, 500 pulses) respectively in the stimulus groups. The whole-cell patch clamp technique was used to record the resting membrane potential (RMP), action potential (AP), voltage-gated potassium channels current of hippocampal dentate gyrus (DG) granule cells. The results showed that 20 Hz MS significantly increased the number of APs released and the maximum slope of a single AP, reduced the threshold of AP, half width and time to AP peak amplitude, and improved the excitability of hippocampal neurons. The peak currents of potassium channels were decreased, the inactivation curve of transient outward potassium channels shifted to the left significantly, and the time constant of recovery after inactivation increased significantly. 0.5 Hz MS significantly inhibited neuronal excitability and increased the peak currents of potassium channels, but the dynamic characteristics of potassium channels had little change. The results suggest that the dynamic characteristics of voltage-gated potassium channels and the excitability of hippocampal DG granule neurons may be one of the potential mechanisms of neuromodulation by MS.
Objective To observe the effects of high concentr at ion glucose on the calcium-activated potassium channel of rabbits′ retinal Müller cells. Methods The rabbits′retinal Müller cells were cultured in vitro under the condition of high concentration glucose, and identified by immunohistochemical staining and transmission electron microscopy. Patch-clamp technique was used to observe the changes of the calcium-activated potassium channel of retinal Müller cells caused by high concentration glucose at different time.Results High concentration glucose could inhibit the calcium-activated potassium channel of cultured retinal Müller cells in a time-dependent manner. Conclusion High concentration glucose may reduce the biological functions of Müller cells by inhibiting calcium-activated potassium channel. (Chin J Ocul Fundus Dis，2003，19：164-167)
Objective To investigate the role of Kv1. 5 in the pathogenesis of pulmonary hypertension simulated by hypobaria and hypoxia, and the effects of dichloroacetate ( DCA) on the Kv1. 5 expression in pulmonary arterial smooth muscle cells ( PASMCs ) and mean pulmonary arterial pressure ( mPAP) . Methods Twenty-four SD rats were randomly divided into a normal group ( N group) , a high altitude group ( HA group) , and a DCA treated group ( DCA group) . The N group were fed in normalconditions, the HA group and DCA group were fed in a hypobaria and hypoxia chamber simulated to an altitude of 5000 meters. In addition, the DCA group rats were gastric gavaged with DCA ( 70 mg · kg - 1 · d - 1 ) .Twenty-one days later, percentage of wall thickness ( WT% ) and percentage of wall area ( WA% ) of the pulmonary arteriole, mPAP, and the ratio of right ventricle / left ventricle and septum ( RV/ LV + S) were evaluated. Real-time PCR, immunohistochemistry and Western blot were carried out to detect the Kv1. 5 expression in PASMCs. Results In the HA group, WT% , and WA% of pulmonary arteriole, mPAP and RV/ ( LV + S) all increased significantly compared with the N group ( P lt;0. 01) . These changes in the DCA group were significantly lower than those in the HA group( P lt; 0. 01) . Furthermore, the protein and mRNA expression of Kv1. 5 in the PASMCs deceased significantly in the HA group compared with the N group( P lt;0. 01) , but recovered in the DCA group ( P lt;0. 01) . Conclusions The expression of Kv1. 5 in PASMCs is tremendously inhibited in rats fed in high altitude, which might be a important role of pulmonaryhypertension. DCA can inhibit the remodeling of pulmonary arterials probably by recovering Kv1. 5 expression.
Objective To investigate the effects of docosahexenoic acid (DHA) on large conductance Ca2+-activated K+ (BK) channels in normal retinal artery smooth muscle cells (RASMCs). Methods Cultured human RASMCs (6 th-8 th generations) were used to patch clamp experiment. The open probabihties (NP0) in BK channels with different concentrations (0.0, 1.0, 3.0, 5.0, 7.5, 10.0 μmol/L) of DHA were recorded by patch clamp technique in single channel configuration. RASMCs were intervened by different concentrations (0.0, 1.0, 5.0 μmol/L) of DHA as control group, low and high doses of DHA groups, respectively. The protein expressions of β subunit of BK channels in RASMCs from three groups were measured by Western blot. Results The NP0 of BK channels were 0.044 4±0.001 2, 0.081 2±0.004 2, 0.209 0±0.006 1, 0.310 5±0.005 3, 0.465 0±0.007 8 and 0.497 7±0.014 5 with perfusate of 0.0, 1.0, 3.0, 5.0, 7.5, 10.0 μmol/L DHA. DHA activated BK channels in a dose-dependent manner (F=2.621,P＜0.05). There was no significant difference in the protein expression of control group, low and high doses of DHA groups (F=11.657,P＞0.05). Conclusion DHA can directly activate BK channels, no increasing in subunit expression of BK channels.
Objective To explore the activity of Ca2 + -activated K+ ( KCa) inairwaysmoothmuscle cells( ASMCs) in a rat model of chronic obstructive pulmonary disease( COPD) , and to observe the effect of 11, 12-Epoxyeicosatrienoic acid( 11, 12-EETs) on the KCa channel of ASMCs. Methods Forty male Sprague-Dawley rats were randomly assigned to a COPD group and a normal control group. The rats in the COPD group were exposed to cigarette smoking in a relatively closed chamber to induce COPD. The ASMCs were isolated from small bronchi using an acute enzymatic digestion method. In the symmetrical high K+ solution,the KCa currents were separated with inside-out configuration using the patch clamp technique. The activity of KCa currents in ASMCs between the COPD group and the normal group were compared and the effect of 11, 12-EETs on KCa channel was recorded. The opening probability( Po) , opening time( To) and closing time ( Tc) of the KCa were measured. Results Compared with the normal group, Po of KCa in the COPD rats was much shorter ( 0. 084 ±0. 028 vs 0. 198 ±0. 029, P lt; 0. 01) , To was shorter [ ( 0. 732 ±0. 058) ms vs ( 1. 648 ±0. 152) ms, P lt; 0. 01] and Tc was longer[ ( 12. 259 ±2. 612) ms vs ( 6. 753 ±1. 237) ms, P lt;0. 01] . 11, 12-EETs can evoke the activity of KCa currents of ASMCs in the COPD rats while Po was increased( 0. 227 ±0. 059 vs 0. 084 ±0. 028, P lt; 0. 01) , To was much longer[ ( 2. 068 ±0. 064) ms vs ( 0. 732 ±0. 058) ms, P lt; 0. 01] , and Tc was shorter [ ( 4. 273 ±0. 978) ms vs ( 12. 259 ±2. 612) ms, P lt;0. 01] .Conclusions The results suggest that the decreasing of KCa activity plays an important role in the development of COPD. 11,12-EETs can directly evoke the activity of KCa channel in COPD rats, thus relax the airway smooth muscles.
Objective To investigate Kir4.1 expressions in Muuml;ller cells under high glucose conditions and treatment of pigment epitheliumderived factor (PEDF）. Methods Cultured rat Muuml;ller cells were divided into control group (5 mmol/L glucose), high glucose group (25 mmol/L glucose), PEDF treatment group (25 mmol/L glucose+100 ng/ml PEDF) and intervention control group（25 mmol/L glucose+phosphate buffer solution）. Kir4.1 expressions were measured by Western blot and real-time reverse transcription polymerase chain reaction (RT-PCR). Reactive oxygen species (ROS) productions were measured using 2prime;7prime;dichlorofluorescin diacetate and glutathione peroxidase （GPx）expressions were studied by real-time RT-PCR. Results By Western blot and real-time RT-PCR, it was found the expressions of Kir4.1 decreased obviously under high glucose conditions (real-time RT-PCR: t=4.12, P＜0.05; Western blot: t=3.53,P＜0.05); simultaneously, ROS generation was increased （t=3.76,P＜0.05）and GPx level was decreased （t=3.18,P＜0.05）. PEDF treatment inhibited the high glucose-induced Kir4.1 down regulation (real-time RT-PCR: t=3.66, P＜0.05; Western blot: t=6.43,P＜0.01) and decreased ROS generations （t=4.11,P＜0.05） and increased GPx levels (t=5.12,P＜0.01）. Conclusions The high glucose can supress Kir4.1 expressions in Muuml;ller cells by oxidative stress, and PEDF can ameliorate these effects.
ObjectiveTo observe the changes in open probability and protein expression of large conductance Ca2+-activated K+ (BK) channel in retinal vascular smooth muscle cells (RVSMCs) of diabetic rats. MethodsStreptozotocin (STZ)-induced rat diabetic animal model was established by STZ injection intraperitoneally.RVSMCs were isolated by enzyme digestion. The BK currents in control and diabetic groups were recorded by patch clamp technique in single channel configuration. BK channel protein expression in control and diabetic group were measured by Western blot. ResultsCompared with control group, the NP0 of BK channels in diabetic group were significantly increased (t=4.260, P < 0.05). Compared with control group, there was no significant difference inα-subunit protein expression in diabetic group in RVSMCs (t=10.126, P > 0.05); however, β1-subunit protein expression was remarkably increased in diabetic group (t=5.146, P < 0.05). ConclusionThe NP0 of BK channels andβ1-subunit protein expression are increased in RVSMCs of diabetic rats.
Objective To investigate the role of mitochondrial adenosine triphosphatesensitive potassium channel(mitoKATP) in immature myocardial ischemic preconditioning, and to provide evidence for immature myocardial protection. Methods Langendorff isolated heart infused model was used in the experiment. Twentyfour rabbits (aged from 14 to 21 days) were randomly divided into 4 groups：ischemiareperfusion group（I/R group）, myocardial ischemic preconditioning group(E1 group), 5hydroxydecanoate(5-HD) group (E2 group) and Diazoxide (Diaz) group(E3 group). Hemodynamics recovery rate, myocardial water content(MWC), the leakage rates of serum creatine kinase and lactate dehydrogenase, adenosine triphosphate content, superoxide dismutase activity, malondialdehyde content, myocardial cell Ca2+ content and myocardial mitochondrial Ca2+ content, myocardial mitochondrial Ca2+-ATPase activity, the adenosine triphosphate(ATP) synthesizing ability of myocardial mitochondria were tested, and myocardial ultrastructure was observed via electron microscopy. Results The hemodynamics recovery rate, myocardial water content(P＜0.05), adenosine triphosphate content, superoxide dismutase activity, myocardial mitochondrial Ca2+-adenosine triphosphyatase(ATPase) activity and the ATP synthesizing ability of myocardial mitochondria of the rabbits in E1 and E3 group were significantly better than that in I/R group and E2 group(P＜0.05). Malondialdehyde content, the leakage rates of serum creatine kinase and lactate dehydrogenase, myocardial cell Ca2+ content and myocardial mitochondrial Ca2+ content of the rabbits in E1 group and E3 group were significantly lower than that in I/R group and E2 group (P＜0.05). The myocardial ultrastructure injury in E1 and E3 group were significantly reduced compared with that in I/R and E2 group. Conclusion Myocardial ischemic preconditioning has significant protective effects on immature myocardium. Its mechanism may be related to the activation of mitoKATP.
Objective To investigate the effects of diazoxide （DIA）cardioplegic solution on the reduction of donor cardiomyocyte apoptosis, Methods In a Krebs-Henseleit （KH） solution perfused isolated rabbit heart Langendorff model, 32 rabbit hearts were divided into four groups with simple random sampling （8 rabbits in each group ）： DIA group （50μmol/L diazoxide mixed in KH solution）,STH group （ST, Thomas＇ solution）, 5-HD group （50μmol/L diazoxide and 100μmol/L 5-hydroxydecanoic acid mixed in KH solution）, KH group （KH solution）, The rabbit hearts of each group underwent 6 hours of hypothermic （4 C） storage in the corresponding cardioplegic solution. Left ventricular developed pressure （LVDP）, maximal values of positive rate of left ventricular pressure （＋dp/dtmax） were measured before and after storage, The post-storage values of LVDP and ＋dp/dtmax were expressed as the percentage of pre-storage control values. Apoptotic cardiomyocytes were detected by the TdT- mediated dUTP-biotin nick end labeling （TUNEL）. Malonaldehyde （MDA） contents and adenosine triphosphate （ATP） contents were also measured after storage. Results Recovery rates of LVDP, ＋dp/dtmax, and ATP contents in DIA group were higher than those of other 3 groups respectively（P〈0. 05）, Cardiomyocytes apoptosis percentage and MDA content were lower than other 3 groups respectively（P〈0. 05）, Conclusions Diazoxide cardioplegic solution can protect the isolated hearts and this may be relates to opening selective mitochondrial KATP channels. The selective mitochondrial KATP channel antagonist 5-hydroxydecanoic acid can block the cardioprotective effect of diazoxide.