Two-pore-domain potassium channels (K2P) family is widely expressed in many human cell types and organs, which has important regulatory effect on physiological processes. K2P is sensitive to a variety of chemical and physical stimuli, and they have also been critically implicated in transmission of neural signal, ion homeostasis, cell development and death, and synaptic plasticity. Aberrant expression and dysfunction of K2P channels are involved in a range of diseases, including autoimmune, central nervous system, cardiovascular disease and others. The scope of this review is to give a detailed overview of the structure, function, pharmacological regulation, and related diseases of TREK-1 channels, a member of the K2P family.
Potassium Channels, Tandem Pore Domain/genetics* ; Humans ; Animals ; Cardiovascular Diseases/physiopathology* ; Autoimmune Diseases/metabolism* ; Central Nervous System Diseases/physiopathology*

OBJECTIVETo construct the acid-sensitive potassium hannel-3(TASK3) eukaryotic expression plasmid and to establish a stable SH-SY5Y cell line expressing enhanced green fluorescent protein (EGFP)-tagged TASK3.

METHODSTASK3 coding region was subcloned into pEGFP-N1 plasmid to construct a recombinant vector alled pEGFP-TASK3. The correct recombinant expressing plasmid was transfected with X-feet transfection reagent to SH-SY5Y cells. The cell line stably expressiing EGFP tagged-TASK3 gene was established by screening with antibiotic G418 and fluorescence microscope. The expression and localization of the EGFP tagged-TASK3 fusion protein was detected by Western blot and confocal microscope. Exposure of the SH-SY5Y cell line expressing stably TASK3-eGFP fusion proteins was exposed to different pH media (7.0, 6.7, 6.4, 6.1) for 24 h, the cell viability was assessed with cell counting Kit-8 (CCK-8).

RESULTSAll the results of identification by PCR, digestion with restriction endonuclease and sequencing indicated that the recombinant eukaryotic expression plasmid pEGFP-TASK3 was constructed correctly. The stable SH-SY5Y cell line expressing EGFP tagged-TASK3 fusion protein was successfully established. Exposure of the wild type SH-SY5Y cells and the stable SH-SY5Y-GFP tag-TASK3 cell line to different pH media (7.0, 6.7, 6.4, 6.1) for 24 h, the cell viability of two group cells significantly reduced with pH declining, and the difference was statistically significant (P < 0.05). Compared with wild type SH-SY5Y cells, the cell viability of stable SH-SYSY-GFP tag-TASK3 cell line increased significantly with the same pH media, and the difference was statistically significant (P < 0.05).

CONCLUSIONThe eukaryotic expression vector pEGFP-TASK3 is successfully constructed and the cell line stably expressing TASK3-eGFP fusion is established which is important for their fundamental research and potential applications.

Blotting, Western ; Cell Line ; Gene Expression ; Genetic Vectors ; Green Fluorescent Proteins ; genetics ; Humans ; Plasmids ; Polymerase Chain Reaction ; Potassium Channels, Tandem Pore Domain ; genetics ; Transfection

Major depression disorder is an increasing heavy burden in modem society, but its pathological mechanism is still vague. Recent evidence indicated that two pore potassium channel, TREK1, is one of the important drug targets of antidepressants. The structural and functional research progress of TREK1 potassium channel were reviewed with an emphasis on its roles in anti-depression, neuronal protection, and neuronal plasticity. The complicated interactions between TREK1 potassium channel and monoamine transmitters-receptors were also reviewed and future directions to explore the underline mechanism were also discussed.
Animals ; Antidepressive Agents ; pharmacology ; Depressive Disorder, Major ; genetics ; metabolism ; physiopathology ; Drug Delivery Systems ; Gene Knockout Techniques ; Humans ; Neuronal Plasticity ; Polymorphism, Genetic ; Potassium Channels, Tandem Pore Domain ; genetics ; metabolism ; physiology ; Receptors, Serotonin ; metabolism ; Receptors, Serotonin, 5-HT4 ; Serotonin ; pharmacology

This study was aimed to examine the effect of TREK-1 silencing on the function of astrocytes. Three 21-nucleotide small interfering RNA (siRNA) duplexes (siT1, siT2, siT3) targeting TREK-1 were constructed. Cy3-labeled dsRNA oligmers were used to determine the transfection efficiency in cultured astrocytes. TREK-1-specific siRNA duplexes (siT1, siT2, siT3) at the optimal concentration were transfected into cultured astrocytes, and the most efficient siRNA was identified by the method of immunocytochemical staining and Western blotting. The proliferation of astrocytes tranfected with TREK-1-targeting siRNA under hypoxia condition was measured by fluorescence-activated cell sorting (FACS). The results showed that TREK-1 was expressed in cultured astrocytes. The dsRNA oligmers targeting TREK-1 could be transfected efficiently in cultured astrocytes and down-regulate the expression of TREK-1 in astrocytes. Moreover, the down-regulation of TREK-1 in astrocytes contributed to the proliferation of astrocytes under hypoxia condition as determined by cell cycle analysis. It was concluded that siRNA is a powerful technique that can be used to knockdown the expression of TREK-1 in astrocytes, which helps further investigate the function of TREK-1 channel in astrocytes under physicological and pathological condition.
Animals ; Astrocytes ; physiology ; Cells, Cultured ; Gene Silencing ; physiology ; Potassium Channels ; Potassium Channels, Tandem Pore Domain ; genetics ; RNA Interference ; physiology ; RNA, Small Interfering ; genetics ; Rats

Animals ; Arachidonic Acid ; pharmacology ; Brain ; metabolism ; Brain Ischemia ; metabolism ; Female ; Humans ; Neuroprotective Agents ; pharmacology ; Ovary ; metabolism ; Potassium Channels ; chemistry ; genetics ; metabolism ; Potassium Channels, Tandem Pore Domain ; chemistry ; genetics ; metabolism ; Riluzole ; pharmacology

AIMTo study mRNA expression alteration of two-pore potassium channels in the brain of beta-amyloid peptide25-35 (beta-AP25-35)-induced memory impaired rats.

METHODSMemory impairments induced in rats by single icv injection of beta-AP25-35 (2 mmol.L-1) 5 microL were assessed in the Morris water maze test. The mRNA expression levels of three two-pore potassium channels TREK-1, TREK-2 and TRAAK were detected in rat cerebral cortex and hippocampus by reverse transcription-polymerase chain reaction (RT-PCR).

RESULTSIn the Morris water maze test, the escape latencies of the beta-AP25-35-treated rats were longer than those of the control group in 1st, 2nd and 4th training day, suggesting that the memory of beta-AP25-35-treated rats was obviously impaired. Compared with the control group, the mRNA levels of TREK-1, TREK-2 and TRAAK in the hippocampus of the beta-AP25-35-treated rats were increased by 40.0%, 27.9% and 18.9%, respectively; while no significant change of TREK-1, TREK-2 and TRAAK mRNA levels was observed in the cortex.

CONCLUSIONThe mRNA expression levels of two-pore potassium channels were increased significantly in the brain of beta-AP25-35-induced memory impaired rats.

Amyloid beta-Peptides ; Animals ; Cerebral Cortex ; metabolism ; Hippocampus ; metabolism ; Male ; Maze Learning ; Memory Disorders ; chemically induced ; metabolism ; Peptide Fragments ; Potassium Channels ; biosynthesis ; genetics ; Potassium Channels, Tandem Pore Domain ; RNA, Messenger ; biosynthesis ; genetics ; Rats ; Rats, Sprague-Dawley ; Reverse Transcriptase Polymerase Chain Reaction