To investigate the electrophysiology mechanisms of new anxiolytic and antidepressant drug: 4-butyl-alpha-agarofuran (AF-5), patch clamp-recording was used to test the effects of AF-5 on voltage-dependent sodium currents, voltage-dependent potassium currents, L-type voltage-dependent calcium currents and GABA dependent Cl(-) currents in primary cultured rat cortical neurons. Effects of AF-5 on Kv2.1 currents, expressed stably in HEK293 cells, were also tested. Our results showed that, delayed rectifier potassium currents (I(K(DR, L-type voltage-dependent calcium currents (I(LC-ca)) in primary cultured rat cortical neurons and Kv2.1 currents in HEK293 cells were significantly inhibited by AF-5, with IC50 as 6.17, 4.4 and 5.29 micromol x L(-1) respectively. However, voltage-dependent sodium currents (I(Na)), GABA dependent Cl(-) currents and transient outward potassium currents (I(K(A)) in primary cultured rat cortical neurons were not significantly blocked by AF-5. Our results concluded that, blocked I(K(DR)) and I(L-Ca) currents may be one of the mechanisms of anxiolytic and antidepression actions of AF-5.
Animals ; Antidepressive Agents ; pharmacology ; Calcium Channels, L-Type ; drug effects ; Cells, Cultured ; Cerebral Cortex ; cytology ; Chloride Channels ; drug effects ; Delayed Rectifier Potassium Channels ; drug effects ; HEK293 Cells ; Humans ; Neurons ; cytology ; Patch-Clamp Techniques ; Potassium Channels, Voltage-Gated ; drug effects ; Rats ; Rats, Wistar ; Sesquiterpenes ; pharmacology ; Shab Potassium Channels ; drug effects ; Voltage-Gated Sodium Channels ; drug effects

AIMTo investigate mRNA expression changes of voltage-gated outward potassium channel subtypes in cultured rat hippocampal neurons after chronic exposure to beta-amyloid-petitde25-35 (beta-AP25-35).

METHODSmRNA expression was detected by RT-PCR, comparative expression levels were determined by imaging densitometer.

RESULTSDelayed rectifying (Kv2.1, Kv1.5), transient outward (Kv1.4, Kv4.2) and large conductance calcium-activated (rSlo) potassium channel mRNA were expressed in cultured rat hippocampal. In the presence of beta-AP25-35 3 mumol.L-1 for 24 h, the relative expression level of Kv2.1 was significantly increased (n = 3, P < 0.05); the other subtypes were not changed obviously (n = 3, P > 0.05). The increase of Kv2.1 mRNA mainly happened between 24 and 36 h after exposure to beta-AP25-35. After exposure to beta-AP25-35 for 60 h, Kv2.1 mRNA decreased significantly (n = 3, P < 0.01).

CONCLUSIONThe upregulation of Kv2.1 on transcription levels may be involved in the enhancement of delayed rectifying outward potassium (Ik) current induced by beta-AP25-35.

Amyloid beta-Peptides ; toxicity ; Animals ; Animals, Newborn ; Cell Division ; drug effects ; Cells, Cultured ; Delayed Rectifier Potassium Channels ; Female ; Gene Expression ; drug effects ; Hippocampus ; cytology ; Male ; Neurons ; drug effects ; metabolism ; Peptide Fragments ; toxicity ; Potassium Channels ; biosynthesis ; genetics ; Potassium Channels, Voltage-Gated ; RNA, Messenger ; biosynthesis ; drug effects ; Rats ; Rats, Wistar ; Shab Potassium Channels

AIMTo study the effects of 17beta-estradiol on Kv2.1 potassium channel current and delayed rectifier potassium current (IK) in cultured rat hippocampal neurons.

METHODSThe effects of 17beta-estradiol on Kv2.1 channel current and IK in cultured rat hippocampal neurons were observed using the whole cell patch clamp techniques.

RESULTS17beta-Estradiol was shown to reduce the amplitude of Kv2.1 current and IK in concentration-dependent manners. The IC50s of 17beta-estradiol blocking Kv2.1 and IK were 2.4 and 4.0 micromol x L(-1), respectively. 17beta-Estradiol (3 micromol x l(-1)) significantly shifted the steady-state activation and inactivation curves of Kv2.1 current to negative potentials. However, it only produced the shift of the steady-state activation curve of IK to the negative potential without effect on the steady-state inactivation of IK.

CONCLUSION17beta-Estradiol inhibits Kv2.1 and IK of hippocampus at similar level. The inhibition of 17beta-estradiol on IK current may be partially via blocking Kv2.1 current.

Animals ; Animals, Newborn ; Cell Line ; Cells, Cultured ; Delayed Rectifier Potassium Channels ; Dose-Response Relationship, Drug ; Embryo, Mammalian ; Estradiol ; pharmacology ; Female ; Hippocampus ; cytology ; physiology ; Humans ; Kidney ; cytology ; Male ; Neurons ; cytology ; physiology ; Patch-Clamp Techniques ; Potassium Channels, Voltage-Gated ; drug effects ; Rats ; Rats, Wistar ; Shab Potassium Channels

Animals ; Cerebral Cortex ; cytology ; physiology ; Delayed Rectifier Potassium Channels ; physiology ; Neurons ; physiology ; Patch-Clamp Techniques ; Rats ; Rats, Sprague-Dawley

BACKGROUNDPrevious studies demonstrated general anesthetics affect potassium ion channels, which may be one of the mechanisms of general anesthesia. Because the effect of etomidate on potassium channels in rat hippocampus which is involved in memory function has not been studied, we investigated the effects of etomidate on both delayed rectifier potassium current (I(K(DR))) and transient outward potassium current (I(K(A))) in acutely dissociated rat hippocampal pyramidal neurons.

METHODSSingle rat hippocampal pyramidal neurons from male Wistar rats of - 10 days were acutely dissociated by enzymatic digestion and mechanical dispersion according to the methods of Kay and Wong with slight modification. Voltage-clamp recordings were performed in the whole-cell patch clamp configuration. Currents were recorded with a List EPC-10 amplifier and data were stored in a computer using Pulse 8.5. Student's paired two-tail t test was used for data analysis.

RESULTSAt the concentration of 100 micromol/L, etomidate significantly inhibited I(K(DR)) by 49.2% at +40 mV when depolarized from -110 mV (P < 0.01, n = 8), while did not affect I(K(A)) (n = 8, P > 0.05). The IC(50) value of etomidate for blocking I(K(DR)) was calculated as 5.4 micromol/L, with a Hill slope of 2.45. At the presence of 10 micromol/L etomidate, the V1/2 of activation curve was shifted from (17.3 +/- 1.5) mV to (10.7 +/- 2.9) mV (n = 8, P < 0.05), the V1/2 of inactivation curve was shifted from (-18.3 +/- 2.2) mV to (-45.3 +/- 9.4) mV (n = 8, P < 0.05). Etomidate 10 micromol/L shifted both the activation curve and inactivation curve of I(K(DR)) to negative potential, but mainly affected the inactivation kinetics.

CONCLUSIONSEtomidate potently inhibited I(K(DR)) but not I(K(A)) in rat hippocampal pyramidal neurons. I(K(DR)) was inhibited by etomidate in a concentration-dependent manner, while I(K(A)) remained unaffected.

Anesthetics, Intravenous ; pharmacology ; Animals ; Delayed Rectifier Potassium Channels ; drug effects ; physiology ; Etomidate ; pharmacology ; Male ; Potassium Channels ; drug effects ; physiology ; Pyramidal Cells ; drug effects ; physiology ; Rats ; Rats, Wistar

BACKGROUND: This study aimed to explore the effects of norepinephrine on the potassium currents of rat medial vestibular nuclear neurons by using the whole-cell patch clamp technique. METHODS: Sprague-Dawley rats aged 14 to 16 days were anesthetized with ether and decapitated. After enzymatic digestion, the portion of the medial vestibular nucleus neurons were obtained by micropunching. The dissociated neurons were transferred into a recording chamber mounted on an inverted microscope and potassium currents were recorded by standard patch-clamp techniques under voltage-clamp modes. RESULTS: Norepinephrine inhibited the whole potassium currents of the medial vestibular nuclear neurons. Outward potassium currents were more suppressed in 4 mM 4-aminopyridine and norepinephrine added solutions than 4 mM 4-aminopyridine only, but were not suppressed in 10 mM tetraethylammonium and norepinephrine added solutions compared to 10 mM tetraethylammonium only. CONCLUSIONS: These results suggest that norepinephrine blocks the delayed rectifier potassium channels in medial vestibular nuclear neurons.
4-Aminopyridine ; Animals ; Delayed Rectifier Potassium Channels ; Digestion ; Ether ; Neurons* ; Norepinephrine* ; Patch-Clamp Techniques ; Potassium* ; Rats* ; Rats, Sprague-Dawley ; Tetraethylammonium ; Vestibular Nuclei

AIMTo observe the change of potassium current on cultured neurons differentiated from hippocampus neural stem cells of the newborn rat.

METHODSNeural stem cells from newborn rat hippocampus were cultured in vitro and passaged continuously. Differentiation of the cell was induced by serum and removing mitogens. After differentiation cells were plated on plastic dishes and cultured for 1 d, 7 d, 14 d and 21 d. Whole-cell voltage patch clamp recording was used respectively to detect voltage-dependent K+ current.

RESULTSAfter 1 d culture, no current was detected, and on the 7th d, 14th d, 21st d after differentiation, the amplitude of K+ currents was (18.077 +/- 2.789)pA/pF, (13.099 +/- 2.742)pA/pF, (34.045 +/- 8.067)pA/pF at +50 mV. The recorded K+ current included two components that could be blocked by TEA and 4-AP separately, assumed the slowly inactivating delayed rectifier K+ current (IK) and the fast inactivating transient outward K+ current (IA).

CONCLUSIONThe function of potassium channels on the hippocampus neural stem cells of the newborn rat approaches mature gradually when the time of differentiation becomes longer in vitro.

Animals ; Animals, Newborn ; Cells, Cultured ; Delayed Rectifier Potassium Channels ; physiology ; Hippocampus ; cytology ; Neural Stem Cells ; cytology ; metabolism ; physiology ; Patch-Clamp Techniques ; Potassium Channels ; physiology ; Potassium Channels, Inwardly Rectifying ; physiology ; Rats ; Rats, Sprague-Dawley

AIMTo investigate the effect of benzyltetrahydropalmatine (BTHP) on the rapidly activating component of delayed rectifier K+ current (Ikr) in single guinea pig ventricular myocytes.

METHODSWhole-cell patch clamp technique was used to record Ikr.

RESULTSIkr was blocked by 1-100 mumol.L-1 BTHP in concentration-, voltage-, and specifically frequency-dependent fashion, with IC50 of 13.5 mumol.L-1 (95% confidence range: 11.2-15.8 mumol.L-1). 30 mumol.L-1 BTHP reduced Ikr and Ikr.tail by (31 +/- 4)% and (36 +/- 5)% (n = 6, P < 0.01), respectively. The time constant for deactivation (tau') of the tail current was decreased by 30 mumol.L-1 BTHP from (238 +/- 16) ms to (196 +/- 14) ms, while drug had no any effect on the time constant for activation (tau) of Ikr,tail.

CONCLUSIONBTHP inhibited Ikr in a frequency-dependent fashion.

Animals ; Anti-Arrhythmia Agents ; pharmacology ; Berberine Alkaloids ; pharmacology ; Cell Separation ; Delayed Rectifier Potassium Channels ; Female ; Guinea Pigs ; Heart Ventricles ; cytology ; metabolism ; Male ; Myocytes, Cardiac ; drug effects ; metabolism ; Patch-Clamp Techniques ; Potassium Channels ; drug effects ; metabolism ; Potassium Channels, Voltage-Gated

OBJECTIVETo correlate delayed rectifier K(+) channel to cyclooxygenase-2 (COX-2) in onco genesis of human gastric cancer cell.

METHODSHuman COX-2 encoding gene was cloned with RT-PCR strategy and its antisense recombinant eukaryotic expression vector was constructed. COX-2 highly expressed human gastric cancer cell line SGC7901 was stably transfected with the antisense vector. The whole-cell recording technique of perforated patch clamp was employed to observe the change of delayed rectifier K(+) current (I(k)) of SGC7901 after gene transfer or treatment with COX-2 inhibitor indomethacin. MTT was also performed to determine the effect of delayed rectifier K(+) channel inhibitors on cell growth.

RESULTSStably transfected cell (7901-AS) was obtained and a down-regulated expression of COX-2 protein and mRNA in the cell was achieved. Patch clamp recording showed that both SGC7901 and 7901-AS cells had a typical delayed rectifier K(+) current. However, I(k) was significantly lower (P < 0.01) in transfected cell or cell treated with indomethacin at each test potential. The altered I(k) could be entirely recovered after drug removal from the cells. K(+) channel blockers tetraethylammonium (TEA) and 4-aminopyridine (4-AP) could retard the growth of SGC7901 and the transfected cell in a dose-dependent manner.

CONCLUSIONDelayed rectifier K(+) channel, existing in human gastric cancer cell line SGC7901, is related to the growth of the cell. The highly expressed COX-2 may affect the biological behavior of gastric cancer cell by regulating this ion channel.

Cell Division ; physiology ; Cyclooxygenase 2 ; Delayed Rectifier Potassium Channels ; Humans ; Isoenzymes ; metabolism ; Membrane Proteins ; Potassium Channels ; metabolism ; Potassium Channels, Voltage-Gated ; Prostaglandin-Endoperoxide Synthases ; metabolism ; Stomach Neoplasms ; enzymology ; metabolism ; Tumor Cells, Cultured

AIMTo study the effect of magnesium sulfate on transient outward K+ current (IA) and delayed rectifier K+ current (IK) in freshly dissociated hippocampal neurons of rats.

METHODSThe whole-cell patch clamp techniques were used.

RESULTSMagnesium sulfate reversibly reduced the amplitudes of IA and IK in a concentration-dependent and voltage-dependent, but not frequency-dependent manner. Half-blocking concentration (IC50) on IA and IK were 6.30 mmol.L-1 and 7.60 mmol.L-1, respectively. Magnesium sulfate (6 mmol.L-1) affected the activation process of IA and IK. Before and after application of the drug, the half-activation voltages of IA were (7 +/- 6) mV and (-7 +/- 11) mV (n = 10, P < 0.01), and the half-activation voltages of IK were (20 +/- 6) mV and (28 +/- 4) mV (n = 10, P < 0.01), but the slope factors were not changed. In addition, magnesium sulfate (6 mmol.L-1) also affected the inactivation process of IA. Before and after application of the drug, the half-inactivation voltages of IA were (-65 +/- 5) mV and (-89 +/- 6) mV (n = 10, P < 0.01).

CONCLUSIONMagnesium sulfate inhibited IA and IK in freshly dissociated hippocampal neurons of rats, which might contribute to protect the central neuronal system (CNS) against damages induced by ischemia and oxygen deprivation.

Animals ; Cell Separation ; Delayed Rectifier Potassium Channels ; Female ; Hippocampus ; cytology ; Magnesium Sulfate ; pharmacology ; Male ; Neurons ; drug effects ; physiology ; Neuroprotective Agents ; pharmacology ; Patch-Clamp Techniques ; Potassium Channels ; drug effects ; metabolism ; Potassium Channels, Voltage-Gated ; Rats ; Rats, Wistar