OBJECTIVE: To observe the mechanism of "" acupuncture for the opening of ATP sensitive potassium channel (K) against cerebral ischemia reperfusion injury in rats. METHODS: Eighty-four rats were randomly divided into a sham-operation group, a model group, an electroacupuncture (EA) group, an EA+K blocker group, 21 rats in each group. 10 μL intracerebral injection with glipizide (1 μmol/5 μL) was used in the EA+K blocker group. The cerebral ischemia reperfusion model was established by Zea Longa's suture method in the model group, the EA group and the EA+K blocker group. Rats in the sham-operation group were received the same surgery but without nylon filament insertion. Acupuncture (20 min a time) was performed at "Neiguan" (PC 6), "Shuigou" (GV 26) and "Sanyinjiao" (SP 6) in the EA group and the EA+K blocker group at 10:00 and 16:00 for 3 days, firstly 90 min after model establishment. EA (2 Hz/15 Hz, 1 mA) was connected at the affected "Neiguan" (PC 6) and "Sanyinjiao" (SP 6). The same fixation was used in the sham-operation group and the model group, without EA. Neurological function was assessed by Zausinger's neurologic assessment scale. 2, 3, 5-triphenyltetrazolium chloride (TTC) staining was used to detect infarct volume. Neurocyte apoptosis in the hippocampus was detected by flow cytometry and the protein expressions of B lymphocytoma-2 gene (Bcl-2) and B cell lymphoma factor-associated X protein (Bax) were measured by Western-blot. RESULTS: In comparison with the model group, the neurological score of the EA group increased (<0.01); the infarction volume and the hippocampal neuron's total apoptosis rate of the EA group decreased (both <0.05); the protein expression of Bcl-2 and Bcl-2/Bax of the EA group increased (<0.05, <0.01); and the protein expression of Bax of the EA group decreased (<0.01). Compared with the EA group, the neurological score of the EA+K blocker group decreased (<0.05); the total apoptosis rate of hippocampus neurons of the EA+K blocker group increased (<0.05); the expression of Bcl-2 protein of the EA+K blocker group reduced (<0.05); the expression of Bax protein of the EA+K blocker group increased (<0.05). CONCLUSION "" acupuncture has brain protective effect on rats with focal cerebral ischemia reperfusion injury. The mechanism may be related to regulating the opening of K channels and decreasing the apoptosis of neurons.
Acupuncture Points ; Animals ; Brain Ischemia ; Electroacupuncture ; KATP Channels ; Rats ; Rats, Sprague-Dawley ; Reperfusion Injury

ATP-sensitive potassium channels (K) are energy sensors on the plasma membrane. By sensing the intracellular ADP/ATP ratio of β-cells, pancreatic K channels control insulin release and regulate metabolism at the whole body level. They are implicated in many metabolic disorders and diseases and are therefore important drug targets. Here, we present three structures of pancreatic K channels solved by cryo-electron microscopy (cryo-EM), at resolutions ranging from 4.1 to 4.5 Å. These structures depict the binding site of the antidiabetic drug glibenclamide, indicate how Kir6.2 (inward-rectifying potassium channel 6.2) N-terminus participates in the coupling between the peripheral SUR1 (sulfonylurea receptor 1) subunit and the central Kir6.2 channel, reveal the binding mode of activating nucleotides, and suggest the mechanism of how Mg-ADP binding on nucleotide binding domains (NBDs) drives a conformational change of the SUR1 subunit.
Adenosine Triphosphate ; metabolism ; Amino Acid Sequence ; Animals ; Binding Sites ; Cryoelectron Microscopy ; Ligands ; Mesocricetus ; Mice ; Models, Molecular ; Nucleotides ; metabolism ; Pancreas ; metabolism ; Potassium Channels, Inwardly Rectifying ; chemistry ; metabolism ; Protein Binding ; Protein Multimerization ; Protein Structure, Quaternary ; Protein Subunits ; chemistry ; metabolism ; Sf9 Cells ; Spodoptera ; Sulfonylurea Receptors ; chemistry ; metabolism

OBJECTIVE: Corticotropin-releasing hormone (CRH) is a crucial regulator of human pregnancy and parturition. Adenosine triphosphate (ATP)-sensitive potassium (KATP) channels are important for regulating myometrial quiescence during pregnancy. We investigated regulatory effects of different concentrations of CRH on KATP channel expression in human myometrial smooth muscle cells (HSMCs) in in vitro conditions. METHODS: After treating HSMCs with different concentrations of CRH (1, 10, 102, 103, 104 pmol/L), mRNA and protein expression of KATP channel subunits (Kir6.1 and SUR2B) was analyzed by reverse transcription-polymerase chain reaction and western blot. We investigated which CRH receptor was involved in the reaction and measured the effects of CRH on intracellular Ca2+ concentration when oxytocin was administered in HSMCs using Fluo-8 AM ester. RESULTS: When HSMCs were treated with low (1 pmol/L) and high (103, 104 pmol/L) CRH concentrations, KATP channel expression significantly increased and decreased, respectively. SUR2B mRNA expression at low and high CRH concentrations was significantly antagonized by antalarmin (CRH receptor-1 antagonist) and astressin 2b (CRH receptor-2 antagonist), respectively; however, Kir6.1 mRNA expression was not affected. After oxytocin treatment, the intracellular Ca2+ concentration in CRH-treated HSMCs was significantly lowered in low concentration of CRH (1 pmol/L), but not in high concentration of CRH (103 pmol/L), compared to control. CONCLUSION: Our data demonstrated the regulatory effect was different when HSMCs were treated with low (early pregnancy-like) and high (labor-like) CRH concentrations and the KATP channel expression showed significant increase and decrease. This could cause inhibition and activation, respectively, of uterine muscle contraction, demonstrating opposite dual actions of CRH.
Adenosine Triphosphate ; Adenosine ; Animals ; Blotting, Western ; Corticotropin-Releasing Hormone ; Female ; Humans ; In Vitro Techniques ; KATP Channels ; Mice ; Myocytes, Smooth Muscle ; Myometrium ; Oxytocin ; Parturition ; Potassium Channels ; Potassium ; Pregnancy ; Receptors, Corticotropin-Releasing Hormone ; RNA, Messenger

OBJECTIVETo analyze the clinical characteristics of an infant with neonatal diabetes mellitus (NDM) and to sequence the ABCC8 gene of this family in order to provide a theoretical basis for the diagnosis and treatment.

METHODSThe clinical data of the patient was collected, and the proband and his direct relatives within three generations were sequenced.

RESULTSThe patient was 1-month-old, random blood glucose was more than 27.8 mmol/L, C-peptide was 33.8 pmol/L, blood gas analysis was pH 7.16, HCO3.9 mmol/L and urine alkone was 3+. Genetic testing revealed that the patient, his father, elder brother and grandmother have carried heterozygous mutation c.2690A>T(p.D897V) of the ABCC8 gene. Fluid infusion, intravenous administration of insulin and other supportive therapies were provided. After the correction of acidosis, subcutaneous insulin injection were uesd to control the blood glucose. Eight months later, blood glucose was pooly controlled. After combined with glibenclamide, blood glucose was under control.

CONCLUSIONThe patient carries a heterozygous mutation c.2690A>T(p.D897V) of ABCC8 gene, which is a novel mutation. Glibenclamide was partly effective for the patient.

Blood Glucose ; genetics ; Diabetes Mellitus ; genetics ; Follow-Up Studies ; Humans ; Infant ; Infant, Newborn ; Infant, Newborn, Diseases ; genetics ; Male ; Mutation ; genetics ; Sulfonylurea Receptors ; genetics

ATP-sensitive potassium channels (K), as an inward rectifying potassium channel, are widely distributed in many types of tissues. Kare activated by the depletion of ATP level and the increase in oxidative stress in cells. The activity of Kcouples cell metabolism with electrical activity and results in membrane hyperpolarization. Kare ubiquitously distributed in the brain, including substantia nigra, hippocampus, hypothalamus, cerebral cortex, dorsal nucleus of vagus and glial cells, and participate in neuronal excitability, mitochondria homeostasis and neurotransmitter release. Accumulating lines of evidence suggest that Kare the major contributing factors in the pathogenesis of Parkinson's disease (PD). This review discussed the association of Kwith the pathogenic processes of PD by focusing on the roles of Kon the degeneration of dopaminergic neurons, the functions of mitochondria, the firing pattern of dopaminergic neurons in the substantia nigra, the α-synuclein secretion from striatum, and the microglia activation.
Dopaminergic Neurons ; Humans ; KATP Channels ; Mitochondria ; Oxidative Stress ; Parkinson Disease ; Synaptic Transmission

BACKGROUND AND OBJECTIVES: Adenosine triphosphate (ATP)-sensitive potassium (K(ATP)) channels play an important role in myocardial protection. We examined the effects of thromboxane A₂ on the regulation of K(ATP) channel activity in single ventricular myocytes. SUBJECTS AND METHODS: Single ventricular myocytes were isolated from the hearts of adult Institute of Cancer Research (ICR) mice by enzymatic digestion. Single channel activity was recorded by excised inside-out and cell-attached patch clamp configurations at -60 mV holding potential during the perfusion of an ATP-free K-5 solution. RESULTS: In the excised inside-out patches, the thromboxane A₂ analog, U46619, decreased the K(ATP) channel activity in a dose-dependent manner; however, the thromboxane A₂ receptor antagonist, SQ29548, did not significantly attenuate the inhibitory effect of U46619. In the cell-attached patches, U46619 inhibited dinitrophenol (DNP)-induced K(ATP) channel activity in a dose-dependent manner, and SQ29548 attenuated the inhibitory effects of U46619 on DNP-induced K(ATP) channel activity. CONCLUSION: Thromboxane A₂ may inhibit K(ATP) channel activity, and may have a harmful effect on ischemic myocardium.
15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid ; Adenosine Triphosphate ; Adenosine* ; Adult ; Animals ; Digestion ; Heart ; Humans ; KATP Channels ; Mice* ; Muscle Cells* ; Myocardium ; Perfusion ; Potassium Channels* ; Potassium*

BACKGROUND AND OBJECTIVES: Adenosine triphosphate (ATP)-sensitive potassium (K(ATP)) channels play an important role in myocardial protection. We examined the effects of thromboxane A₂ on the regulation of K(ATP) channel activity in single ventricular myocytes. SUBJECTS AND METHODS: Single ventricular myocytes were isolated from the hearts of adult Institute of Cancer Research (ICR) mice by enzymatic digestion. Single channel activity was recorded by excised inside-out and cell-attached patch clamp configurations at -60 mV holding potential during the perfusion of an ATP-free K-5 solution. RESULTS: In the excised inside-out patches, the thromboxane A₂ analog, U46619, decreased the K(ATP) channel activity in a dose-dependent manner; however, the thromboxane A₂ receptor antagonist, SQ29548, did not significantly attenuate the inhibitory effect of U46619. In the cell-attached patches, U46619 inhibited dinitrophenol (DNP)-induced K(ATP) channel activity in a dose-dependent manner, and SQ29548 attenuated the inhibitory effects of U46619 on DNP-induced K(ATP) channel activity. CONCLUSION: Thromboxane A₂ may inhibit K(ATP) channel activity, and may have a harmful effect on ischemic myocardium.
15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid ; Adenosine Triphosphate ; Adenosine* ; Adult ; Animals ; Digestion ; Heart ; Humans ; KATP Channels ; Mice* ; Muscle Cells* ; Myocardium ; Perfusion ; Potassium Channels* ; Potassium*

OBJECTIVETo investigate the effect of curcumin on oligomer formation and mitochondrial ATP-sensitive potassium channels (mitoKATP) induced by overexpression or mutation of α-synuclein.

METHODSRecombinant plasmids α-synuclein-pEGFP-A53T and α-synuclein-pEGFP-WT were transfected into PC12 cells by lipofectamin method, and intervened by application of curcumin (20 μmol/L) and 5-hydroxydecanoate (5-HD). Oligomer formation in the cultured cells was identified by Western blotting and Dot blotting. Cytotoxicity and apoptosis of the PC12 cells were measured by lactate dehydrogenase (LDH) and JC-1 assays. mitoKATP were identified by Western blotting and whole cell patch clamp.

RESULTSCurcumin has significantly reduced the oligomer formation induced by overexpression or mutation of α-synuclein in the cultured cells. LDH has decreased by 36.3% and 23.5%, and red/green fluorescence ratio of JC-1 was increased respectively by 48.46% and 50.33% after application of curcumin (P<0.05). Protein expression of Kir6.2 has decreased and mitoKATP channel current has significantly increased (P<0.05).

CONCLUSIONCurcumin can inhibit α-synuclein gene overexpression or mutation induced α-synuclein oligomers formation. It may block apoptosis induced by wild-type overexpression or mutation of α-synuclein. By stabilizing mitochondrial membrane potential. Opening of mitoKATP channel may have been the initiating protective mechanism of apoptosis induced by wild-type overexpression or mutation of α-synuclein. Curcumin may antagonize above cytotoxicity through further opening the mitoKATP channel.

Animals ; Apoptosis ; drug effects ; Cell Line ; Curcumin ; pharmacology ; Humans ; KATP Channels ; chemistry ; genetics ; metabolism ; Mitochondria ; drug effects ; genetics ; metabolism ; Mutation ; drug effects ; PC12 Cells ; Parkinson Disease ; drug therapy ; genetics ; metabolism ; physiopathology ; Rats ; alpha-Synuclein ; genetics

AMP-activated protein kinase (AMPK) is a metabolic sensor activated during metabolic stress and it regulates various enzymes and cellular processes to maintain metabolic homeostasis. We previously reported that activation of AMPK by glucose deprivation (GD) and leptin increases KATP currents by increasing the surface levels of KATP channel proteins in pancreatic beta-cells. Here, we show that the signaling mechanisms that mediate actin cytoskeleton remodeling are closely associated with AMPK-induced KATP channel trafficking. Using F-actin staining with Alexa 633-conjugated phalloidin, we observed that dense cortical actin filaments present in INS-1 cells cultured in 11 mM glucose were disrupted by GD or leptin treatment. These changes were blocked by inhibiting AMPK using compound C or siAMPK and mimicked by activating AMPK using AICAR, indicating that cytoskeletal remodeling induced by GD or leptin was mediated by AMPK signaling. AMPK activation led to the activation of Rac GTPase and the phosphorylation of myosin regulatory light chain (MRLC). AMPK-dependent actin remodeling induced by GD or leptin was abolished by the inhibition of Rac with a Rac inhibitor (NSC23766), siRac1 or siRac2, and by inhibition of myosin II with a myosin ATPase inhibitor (blebbistatin). Immunocytochemistry, surface biotinylation and electrophysiological analyses of KATP channel activity and membrane potentials revealed that AMPK-dependent KATP channel trafficking to the plasma membrane was also inhibited by NSC23766 or blebbistatin. Taken together, these results indicate that AMPK/Rac-dependent cytoskeletal remodeling associated with myosin II motor function promotes the translocation of KATP channels to the plasma membrane in pancreatic beta-cells.
AMP-Activated Protein Kinases/metabolism ; Actins/*metabolism ; Animals ; Cell Line ; Glucose/metabolism ; Insulin-Secreting Cells/*metabolism ; KATP Channels/*metabolism ; Leptin/metabolism ; Myosin Type II/*metabolism ; Phosphorylation ; Rats ; *Signal Transduction ; rac GTP-Binding Proteins/*metabolism

The aim of the present study is to investigate the expressions of ATP-sensitive K(+) channels (KATP) in pulmonary artery smooth muscle cells (PASMCs) and the relationship with p38 MAPK signal pathway in rats. Male SD rat PASMCs were cultured in vitro, and a model of hypoxia and hypercapnia was reconstructed. PASMCs were divided to normal (N), hypoxia-hypercapnia (H), hypoxia-hypercapnia+DMSO incubation (HD), hypoxia-hypercapnia+SB203580 (inhibitor of p38 MAPK pathway) incubation (HS) and hypoxia-hypercapnia+Anisomycin (agonist of p38 MAPK pathway) incubation (HA) groups. Western blot was used to detect the protein expression of SUR2B and Kir6.1; semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) was used to detect the mRNA expression of SUR2B and Kir6.1. The results demonstrated that: (1) Compared with N, H, HD and HS groups, the expressions of Kir6.1 mRNA and protein in PASMCs of HA group were decreased significantly (P < 0.01), but there were no differences among N, H, HD and HS groups (P > 0.05); (2) Compared with N group, the expressions of SUR2B mRNA and protein in H, HD, HS and HA groups were increased significantly (P < 0.05), but there were no differences among H, HD, HS and HA groups (P > 0.05). The results imply that: (1) Hypoxia-hypercapnia, SB203580 didn't change the expressions of Kir6.1 mRNA and protein in PASMCs, but Anisomycin decreased the expressions of Kir6.1 mRNA and protein, so Kir6.1 may be regulated by the other subfamily of MAPK pathway; (2) Hypoxia-hypercapnia raised SUR2B mRNA and protein expressions in PASMCs, but SB203580 and Anisomycin did not affect the changes, so the increasing of SUR2B mRNA and protein induced by hypoxia-hypercapnia may be not depend on p38 MAPK pathway.
Animals ; Anisomycin ; pharmacology ; Cell Hypoxia ; Cells, Cultured ; Hypercapnia ; Imidazoles ; pharmacology ; KATP Channels ; metabolism ; MAP Kinase Signaling System ; Male ; Myocytes, Smooth Muscle ; metabolism ; Pulmonary Artery ; cytology ; Pyridines ; pharmacology ; Rats ; Rats, Sprague-Dawley ; Sulfonylurea Receptors ; metabolism ; p38 Mitogen-Activated Protein Kinases ; antagonists & inhibitors