Zingerone Modulates Neuronal Voltage-Gated Na+ and L-Type Ca2+ Currents.

Zingerone (ZO), a nontoxic methoxyphenol, has been demonstrated to exert various important biological effects. However, its action on varying types of ionic currents and how they concert in neuronal cells remain incompletely understood. With the aid of patch clamp technology, we investigated the effects of ZO on the amplitude, gating, and hysteresis of plasmalemmal ionic currents from both pituitary tumor (GH3) cells and hippocampal (mHippoE-14) neurons. The exposure of the GH3 cells to ZO differentially diminished the peak and late components of the INa. Using a double ramp pulse, the amplitude of the INa(P) was measured, and the appearance of a hysteresis loop was observed. Moreover, ZO reversed the tefluthrin-mediated augmentation of the hysteretic strength of the INa(P) and led to a reduction in the ICa,L. As a double ramp pulse was applied, two types of voltage-dependent hysteresis loops were identified in the ICa,L, and the replacement with BaCl2-attenuated hysteresis of the ICa,L enhanced the ICa,L amplitude along with the current amplitude (i.e., the IBa). The hysteretic magnitude of the ICa,L activated by the double pulse was attenuated by ZO. The peak and late INa in the hippocampal mHippoE-14 neurons was also differentially inhibited by ZO. In addition to acting on the production of reactive oxygen species, ZO produced effects on multiple ionic currents demonstrated herein that, considered together, may significantly impact the functional activities of neuronal cells.

Depressive effectiveness of vigabatrin (γ-vinyl-GABA), an antiepileptic drug, in intermediate-conductance calcium-activated potassium channels in human glioma cells.

BACKGROUND: Vigabatrin (VGB) is an approved non-traditional antiepileptic drug that has been revealed to have potential for treating brain tumors; however, its effect on ionic channels in glioma cells remains largely unclear. METHODS: With the aid of patch-clamp technology, we investigated the effects of VGB on various ionic currents in the glioblastoma multiforme cell line 13-06-MG. RESULTS: In cell-attached configuration, VGB concentration-dependently reduced the activity of intermediate-conductance Ca2+-activated K+ (IKCa) channels, while DCEBIO (5,6-dichloro-1-ethyl-1,3-dihydro-2H-benzimidazol-2-one) counteracted the VGB-induced inhibition of IKCa channels. However, the activity of neither large-conductance Ca2+-activated (BKCa) nor inwardly rectifying K+ (KIR) channels were affected by the presence of VGB in human 13-06-MG cells. However, in the continued presence of VGB, the addition of GAL-021 or BaCl2 effectively suppressed BKCa and KIR channels. CONCLUSIONS: The inhibitory effect of VGB on IKCa channels demonstrated in the current study could be an important underlying mechanism of VGB-induced antineoplastic (e.g., anti-glioma) actions.

Characterization of the Inhibitory Effect of Gastrodigenin and Gastrodin on M-type K+ Currents in Pituitary Cells and Hippocampal Neurons.

Gastrodigenin (HBA) and gastrodin (GAS) are phenolic ingredients found in Gastrodia elata Blume (GEB), a traditional Chinese herbal medicine. These compounds have been previously used to treat cognitive dysfunction, convulsion, and dizziness. However, at present, there is no available information regarding their potential ionic effects in electrically excitable cells. In the current study, the possible effects of HBA and GAS on different ionic currents in pituitary GH3 cells and hippocampal mHippoE-14 neurons were investigated using the patch-clamp technique. The addition of HBA or GAS resulted in the differential inhibition of the M-type K+ current (IK(M)) density in a concentration-dependent manner in GH3 cells. HBA resulted in a slowing of the activation time course of IK(M), while GAS elevated it. HBA also mildly suppressed the density of erg-mediated or the delayed-rectifier K+ current in GH3 cells. Neither GAS nor HBA (10 µM) modified the voltage-gated Na+ current density, although they suppressed the L-type Ca2+ current density at the same concentration. In hippocampal mHippoE-14 neurons, HBA was effective at inhibiting IK(M) density as well as slowing the activation time course. Taken together, the present study provided the first evidence that HBA or GAS could act on cellular mechanisms, and could therefore potentially have a functional influence in various neurologic disorders.

Calreticulin regulates vascular endothelial growth factor-A mRNA stability in gastric cancer cells.

Calreticulin (CRT) and vascular endothelial growth factor-A (VEGF-A) are crucial for angiogenesis, and mediate multiple malignant behaviors in gastric cancer. In this study, we report that CRT is positively correlated with VEGF-A in gastric cancer patients. Moreover, high expressions of both CRT and VEGF-A are markedly associated with the pathological stage, progression, and poor prognosis in the patients. Therefore, we sought to elucidate the mechanism by which CRT affects VEGF-A in gastric cancer. Firstly, we demonstrate the novel finding that knockdown of CRT reduced VEGF-A mRNA stability in two gastric cancer cell lines, AGS and MKN45. The AU-Rich element (ARE) is believed to play a crucial role in the maintenance of VEGF-A mRNA stability. Luciferase reporter assay shows that knockdown of CRT significantly decreased the activity of renilla luciferase with VEGF-A ARE sequence. Additionally, competition results from RNA-binding/electrophoretic mobility shift assay indicate that CRT forms an RNA-protein complex with the VEGF-A mRNA by binding to the ARE. In addition, the proliferation rate of human umbilical vein endothelial cells (HUVEC) was significantly reduced when treated with conditioned medium from CRT knockdown cells; this was rescued by exogenous VEGF-A recombinant protein. Our results demonstrate that CRT is involved in VEGF-A ARE binding protein complexes to stabilize VEGF-A mRNA, thereby promoting the angiogenesis, and progression of gastric cancer.

The Novel Direct Modulatory Effects of Perampanel, an Antagonist of AMPA Receptors, on Voltage-Gated Sodium and M-type Potassium Currents.

Perampanel (PER) is a selective blocker of AMPA receptors showing efficacy in treating various epileptic disorders including brain tumor-related epilepsy and also potential in treating motor neuron disease. However, besides its inhibition of AMPA-induced currents, whether PER has any other direct ionic effects in different types of neurons remains largely unknown. We investigated the effects of PER and related compounds on ionic currents in different types of cells, including hippocampal mHippoE-14 neurons, motor neuron-like NSC-34 cells and U87 glioma cells. We found that PER differentially and effectively suppressed the amplitude of voltage-gated Na+ currents (INa) in mHippoE-14 cells. The IC50 values required to inhibit peak and late INa were 4.12 and 0.78 µM, respectively. PER attenuated tefluthrin-induced increases in both amplitude and deactivating time constant of INa. Importantly, PER also inhibited the amplitude of M-type K+ currents (IK(M)) with an IC50 value of 0.92 µM. The suppression of IK(M) was attenuated by the addition of flupirtine or ZnCl2 but not by L-quisqualic acid or sorafenib. Meanwhile, in cell-attached configuration, PER (3 µM) decreased the activity of M-type K+ channels with no change in single-channel conductance but shifting the activation curve along the voltage axis in a rightward direction. Supportively, PER suppressed IK(M) in NSC-34 cells and INa in U87 glioma cells. The inhibitory effects of PER on both INa and IK(M), independent of its antagonistic effect on AMPA receptors, may be responsible for its wide-spectrum of effects observed in neurological clinical practice.

Risk of ischemic stroke after discharge from inpatient surgery: Does the type of surgery matter?

OBJECTIVE: Stroke is a well-known and devastating complication during the perioperative period. However, detailed stroke risk profiles within 90 days in patients discharged without stroke after inpatient surgery are not fully understood. Using the case-crossover design, we aimed to evaluate the risk of ischemic stroke in these patients. METHODS: We included adult patients with the first hospitalization for ischemic stroke between 2011 and 2012 from 23 million enrollees in the National Health Insurance Research Database. Admission date of the hospitalization was defined as the case day and exactly 365 days before the admission date as the control day. The exposure was the last hospitalization for surgery within 1-30, 31-60, or 61-90 days (case period) before the case day or similar time intervals (control period) before the control day. Surgical types were grouped based on the International Classification of Diseases procedure codes. We performed conditional logistic regression adjusting for time-varying variables to determine the relationship between surgery and subsequent stroke, and case-time-control analyses to examine whether the results were confounded by the time-trend in surgery. RESULTS: A total of 56596 adult patients (41% female, mean age 69 years) comprised the study population. After adjustment was made for confounding variables, an association between stroke and prior inpatient surgery within 30 days was observed (adjusted odds ratio 1.44; 95% confidence interval 1.29-1.61). Cardiothoracic, vascular, digestive surgery, and musculoskeletal surgery within 30 days independently predicted ischemic stroke in the case-crossover analysis. In the case-time-control analysis, inpatient surgery remained an independent risk factor for ischemic stroke, whereas only cardiothoracic, vascular, and digestive surgery independently predicted ischemic stroke. CONCLUSIONS: Surgery as a whole independently increased the risk of ischemic stroke within 30 days. Among various types of surgery, cardiothoracic, vascular, and digestive surgery significantly increased the risk of ischemic stroke.

The Novel Effect of Immunomodulator-Glatiramer Acetate on Epileptogenesis and Epileptic Seizures.

BACKGROUND/AIMS: Immunological mechanisms can be triggered as a response to central nervous system insults and can lead to seizures. In this study an investigation was made to determine if glatiramer acetate (GA), an immunomodulator currently used in the treatment of multiple sclerosis, could protect rats from pilocarpine-induced seizures and chronic epilepsy. METHODS: Two groups of adult male Sprague-Dawley rats, experimental (GA) and control, were used in the study. The systemic IL-1α and IL-1ß levels at baseline were checked as well as status epilepticus (SE), and the spontaneous recurrent seizure (SRS) stage by enzyme-linked immunosorbent assay. The GA group was given GA (150 µg/kg, ip) and the control group was given a saline injection prior to pilocarpine-induced seizures. Seizure susceptibility, severity and mortality were evaluated, using Racine seizure classification and hippocampal damage was evaluated by Nissl staining. The GA group received GA (150 µg/kg/day, ip) daily after SE, and the chronic spontaneous seizures were evaluated by long-term video recording, and mossy fiber sprouting was evaluated by Timm staining. The IL-1α and IL-1ß levels were correlated with seizure activities. The TNF-α level in the hippocampus was determined at the SRS stage by immunohistochemistry. The effect of GA on ionic currents and action potentials (APs) in NG108-15 differentiated neurons was investigated using patch-clamp technology. RESULTS: It was found that latency to severe seizures was significantly longer in the GA (p < 0.01) group, which also had SE of shorter duration and less frequent SRS (p < 0.01). GA attenuated acute hippocampal neuron loss and chronic mossy fiber sprouting in the CA3 and the SRS-reduction correlated with the reduction of IL-1α, but not with IL-1ß or TNF-α levels. Mechanistically, GA reduced the peak amplitude of voltage-gated Na+ current (INa), with a negative shift in the inactivation curve of INa and reduced the amplitude of APs along with decreased firing of APs. CONCLUSION: GA might serve as a neuroexcitability modulator which attenuates pilocarpine-induced acute and chronic excitotoxicity. Sodium channel attenuation was partially independent of the immunomodulatory effect.

Multiple Actions of Rotenone, an Inhibitor of Mitochondrial Respiratory Chain, on Ionic Currents and Miniature End-Plate Potential in Mouse Hippocampal (mHippoE-14) Neurons.

BACKGROUND/AIMS: Rotenone (Rot) is known to suppress the activity of complex I in the mitochondrial chain reaction; however, whether this compound has effects on ion currents in neurons remains largely unexplored. METHODS: With the aid of patch-clamp technology and simulation modeling, the effects of Rot on membrane ion currents present in mHippoE-14 cells were investigated. RESULTS: Addition of Rot produced an inhibitory action on the peak amplitude of INa with an IC50 value of 39.3 µM; however, neither activation nor inactivation kinetics of INa was changed during cell exposure to this compound. Addition of Rot produced little or no modifications in the steady-state inactivation curve of INa. Rot increased the amplitude of Ca2+-activated Cl- current in response to membrane depolarization with an EC50 value of 35.4 µM; further addition of niflumic acid reversed Rot-mediated stimulation of this current. Moreover, when these cells were exposed to 10 µM Rot, a specific population of ATP-sensitive K+ channels with a single-channel conductance of 18.1 pS was measured, despite its inability to alter single-channel conductance. Under current clamp condition, the frequency of miniature end-plate potentials in mHippoE-14 cells was significantly raised in the presence of Rot (10 µM) with no changes in their amplitude and time course of rise and decay. In simulated model of hippocampal neurons incorporated with chemical autaptic connection, increased autaptic strength to mimic the action of Rot was noted to change the bursting pattern with emergence of subthreshold potentials. CONCLUSIONS: The Rot effects presented herein might exert a significant action on functional activities of hippocampal neurons occurring in vivo.

Evidence for the Inhibition by Temozolomide, an Imidazotetrazine Family Alkylator, of Intermediate-Conductance Ca2+-Activated K+ Channels in Glioma Cells.

BACKGROUND: Temozolomide (TMZ), an oral alkylator of the imidazotetrazine family, is used to treat glioma. Whether this drug has any ionic effects in glioma cells remains largely unclear. METHODS: With the aid of patch-clamp technology, we investigated the effects of TMZ on the ionic currents in U373 glioma cells. The mRNA expression of KCNN4 (KCa3.1) in U373 glioma cells and TMZ's effect on K+ currents in these KCNN4 siRNA-transfected U373 cells were investigated. RESULTS: In whole-cell recordings, TMZ decreased the amplitude of voltage-dependent K+ currents (IK) in U373 cells. TMZ-induced IK inhibition was reversed by ionomycin or 1-ethyl-2-benzimidazolinone (1-EBIO). In cell-attached configuration, TMZ concentration-dependently reduced the activity of intermediate-conductance Ca2+-activated K+ (IKCa) channels with an IC50 value of 9.2 µM. Chlorzoxazone or 1-EBIO counteracted the TMZ-induced inhibition of IKCa channels. Although TMZ was unable to modify single-channel conductance, its inhibition of IKCa channels was weakly voltage-dependent and accompanied by a significant prolongation in the slow component of mean closed time. However, neitherlarge-conductance Ca2+-activated (BKCa) nor inwardly rectifying K+ (Kir) channels were affected by TMZ. In current-clamp mode, TMZ depolarized the cell membrane and 1-EBIO reversed TMZ-induced depolarization. TMZ had no effect on IK in KCNN4 siRNA-transfected U373 cells. CONCLUSION: In addition to the DNA damage it does, its inhibitory effect on IKCa channels accompanied by membrane depolarization could be an important mechanism underlying TMZ-induced antineoplastic actions.

Subjective memory evaluation before and after temporal lobe epilepsy surgery.

Subjective memory (SM), a self-evaluation of memory, in contrast to objective memory (OM) measured by neuropsychological testing, is less well studied in patients with epilepsy. We assessed SM before and after temporal lobectomy. The Frequency of Forgetting 10 scale (FOF-10), developed to evaluate SM in dementia, was given before and one year after temporal lobectomy. Reliability and validity for use in epilepsy were first assessed. Measures of depression (CES-D) and neuroticism (PANAS) were done before and after surgery as well as complete neuropsychological assessment of OM. Correlation analysis between FOF-10 results and all the other variables was implemented. In 48 patients the FOF-10 was reliable and valid showing high internal consistency in all items (Cronbach's alpha >0.82) and high reproducibility (p<0.01). The FOF-10 also correlated with the memory assessment clinics self rating scale (MAC-S) (p<0.01). FOF-10 scores improved or were unchanged postoperatively in 28 patients (58%) and worsened in 20 (42%). The FOF-10 did not significantly correlate with memory scores from neuropsychological testing but did correlate with perceived word finding difficulty (p<0.001) and postoperative depression (p<0.05). A reduction in number of antiepileptic drugs (AEDs) after surgery distinguished those with improved postoperative SM. No correlation was found between SM and neuroticism, side of surgery or number of seizures. The FOF-10 is a brief and reliable measure of subjective memory in patients with epilepsy. Perceived memory impairment reflects more emotional state, language problems and quantity of AEDs than actual defects in memory function. These results would potentially be useful in presurgical counselling and management of memory issues after temporal lobe surgery.

Pregabalin attenuates excitotoxicity in diabetes.

Diabetes can exacerbate seizures and worsen seizure-related brain damage. In the present study, we aimed to determine whether the standard antiepileptic drug pregabalin (PGB) protects against pilocarpine-induced seizures and excitotoxicity in diabetes. Adult male Sprague-Dawley rats were divided into either a streptozotocin (STZ)-induced diabetes group or a normal saline (NS) group. Both groups were further divided into subgroups that were treated intravenously with either PGB (15 mg/kg) or a vehicle; all groups were treated with subcutaneous pilocarpine (60 mg/kg) to induce seizures. To evaluate spontaneous recurrent seizures (SRS), PGB-pretreated rats were fed rat chow containing oral PGB (450 mg) for 28 consecutive days; vehicle-pretreated rats were fed regular chow. SRS frequency was monitored for 2 weeks from post-status epilepticus day 15. We evaluated both acute neuronal loss and chronic mossy fiber sprouting in the CA3 area. In addition, we performed patch clamp recordings to study evoked excitatory postsynaptic currents (eEPSCs) in hippocampal CA1 neurons for both vehicle-treated rats with SRS. Finally, we used an RNA interference knockdown method for Kir6.2 in a hippocampal cell line to evaluate PGB's effects in the presence of high-dose ATP. We found that compared to vehicle-treated rats, PGB-treated rats showed less severe acute seizure activity, reduced acute neuronal loss, and chronic mossy fiber sprouting. In the vehicle-treated STZ rats, eEPSC amplitude was significantly lower after PGB administration, but glibenclamide reversed this effect. The RNA interference study confirmed that PGB could counteract the ATP-sensitive potassium channel (KATP)-closing effect of high-dose ATP. By opening KATP, PGB protects against neuronal excitotoxicity, and is therefore a potential antiepileptogenic in diabetes. These findings might help develop a clinical algorithm for treating patients with epilepsy and comorbid metabolic disorders.

Characterizing the effects of Eugenol on neuronal ionic currents and hyperexcitability.

RATIONALE: Eugenol (EUG, 4-allyl-2-methoxyphenol), the main component of essential oil extracted from cloves, has various uses in medicine because of its potential to modulate neuronal excitability. However, its effects on the ionic mechanisms remains incompletely understood. OBJECTIVES: We aimed to investigate EUG's effects on neuronal ionic currents and excitability, especially on voltage-gated ion currents, and to verify the effects on a hyperexcitability-temporal lobe seizure model. METHODS: With the aid of patch-clamp technology, we first investigated the effects of EUG on ionic currents in NG108-15 neuronal cells differentiated with cyclic AMP. We then used modified Pinsky-Rinzel simulation modeling to evaluate its effects on spontaneous action potentials (APs). Finally, we investigated its effects on pilocarpine-induced seizures in rats. RESULTS: EUG depressed the transient and late components of I(Na) in the neurons. It not only increased the degree of I(Na) inactivation, but specifically suppressed the non-inactivating I(Na) (I(Na(NI))). Its inhibition of I (Na(NI)) was reversed by tefluthrin. In addition, EUG diminished L-type Ca(2+) current and delayed rectifier K(+) current only at higher concentrations. EUG's effects on APs frequency reduction was verified by the simulation modeling. In pilocarpine-induced seizures, the EUG-treated rats showed no shorter seizure latency but a lower seizure severity and mortality than the control rats. The EUG's effect on seizure severity was occluded by the I(Na(NI)) antagonist riluzole. CONCLUSION: The synergistic blocking effects of I (Na) and I(Na(NI)) contributes to the main mechanism through which EUG affects the firing of neuronal APs and modulate neuronal hyperexcitability such as pilocarpine-induced temporal lobe seizures.

Reversible postvaccination paraneoplastic encephalomyelitis in a patient with lung adenocarcinoma.

Encephalomyelitis occurs in paraneoplastic syndrome and acute disseminated encephalomyelitis through different autoimmune mechanisms. No postvaccinal encephalomyelitis other than acute disseminated encephalomyelitis has been reported in patients with malignancy. A 68-year-old woman was admitted because of a headache followed by a gait disturbance and psychomotor retardation 2 days after she had received an influenza vaccination followed by abulia, limb rigidity and hyperreflexia of both legs, and meningeal irritation. Cerebrospinal fluid studies showed increased intracranial pressure, elevated immunoglobulins G and A, and pleocytosis. Contrasted brain magnetic resonance imaging revealed ventriculomegaly and multiple symmetric leptomeningeal enhancement, without demyelinating changes or cortical ribbon signs. Somatosensory evoked potentials and nerve conduction velocity studies suggested myelitis. Encephalomyelitis was diagnosed on the basis of clinical and laboratory examinations. The etiological survey identified a lung adenocarcinoma. Both the encephalomyelitis and the lung adenocarcinoma simultaneously progressed after the vaccination and then, after targeted therapy for lung cancer, simultaneously subsided. In conclusion, postinfluenza-vaccination paraneoplastic encephalomyelitis may occur in patients with lung adenocarcinoma.

Diazoxide reduces status epilepticus neuron damage in diabetes.

Diabetic hyperglycemia is associated with seizure severity and may aggravate brain damage after status epilepticus. Our earlier studies suggest the involvement of ATP-sensitive potassium channels (K(ATP)) in glucose-related neuroexcitability. We aimed to determine whether K(ATP) agonist protects against status epilepticus-induced brain damage. Adult male Sprague-Dawley rats were divided into two groups: the streptozotocin (STZ)-induced diabetes (STZ) group and the normal saline (NS) group. Both groups were treated with either diazoxide (15 mg/kg, i.v.) (STZ + DZX, NS + DZX) or vehicle (STZ + V, NS + V) before lithium-pilocarpine-induced status epilepticus. We evaluated seizure susceptibility, severity, and mortality. The rats underwent Morris water-maze tests and hippocampal histopathology analyses 24 h post-status epilepticus. A multi-electrode recording system was used to study field excitatory postsynaptic synaptic potentials (fEPSP). RNA interference (RNAi) to knockdown Kir 6.2 in a hippocampal cell line was used to evaluate the effect of diazoxide in the presence of high concentration of ATP. Seizures were less severe (P < 0.01), post-status epilepticus learning and memory were better (P < 0.05), and neuron loss in the hippocampal CA3 area was lower (P < 0.05) in the STZ + DZX than the STZ + V group. In contrast, seizure severity, post-status epilepticus learning and memory, and hippocampal CA3 neuron loss were comparable in the NS + DZX and NS + V groups. fEPSP was lower in the STZ + DZX but not in the NS + DZX group. The RNAi study confirmed that diazoxide, with its K(ATP)-opening effects, could counteract the K(ATP)-closing effect by high dose ATP. We conclude that, by opening K(ATP), diazoxide protects against status epilepticus-induced neuron damage during diabetic hyperglycemia.

Diabetic hyperglycemia aggravates seizures and status epilepticus-induced hippocampal damage.

Epileptic seizures in diabetic hyperglycemia (DH) are not uncommon. This study aimed to determine the acute behavioral, pathological, and electrophysiological effects of status epilepticus (SE) on diabetic animals. Adult male Sprague-Dawley rats were first divided into groups with and without streptozotocin (STZ)-induced diabetes, and then into treatment groups given a normal saline (NS) (STZ-only and NS-only) or a lithium-pilocarpine injection to induce status epilepticus (STZ + SE and NS + SE). Seizure susceptibility, severity, and mortality were evaluated. Serial Morris water maze test and hippocampal histopathology results were examined before and 24 h after SE. Tetanic stimulation-induced long-term potentiation (LTP) in a hippocampal slice was recorded in a multi-electrode dish system. We also used a simulation model to evaluate intracellular adenosine triphosphate (ATP) and neuroexcitability. The STZ + SE group had a significantly higher percentage of severe seizures and SE-related death and worse learning and memory performances than the other three groups 24 h after SE. The STZ + SE group, and then the NS + SE group, showed the most severe neuronal loss and mossy fiber sprouting in the hippocampal CA3 area. In addition, LTP was markedly attenuated in the STZ + SE group, and then the NS + SE group. In the simulation, increased intracellular ATP concentration promoted action potential firing. This finding that rats with DH had more brain damage after SE than rats without diabetes suggests the importance of intensively treating hyperglycemia and seizures in diabetic patients with epilepsy.

The synergistic inhibitory actions of oxcarbazepine on voltage-gated sodium and potassium currents in differentiated NG108-15 neuronal cells and model neurons.

Oxcarbazepine (OXC), one of the newer anti-epileptic drugs, has been demonstrating its efficacy on wide-spectrum neuropsychiatric disorders. However, the ionic mechanism of OXC actions in neurons remains incompletely understood. With the aid of patch-clamp technology, we first investigated the effects of OXC on ion currents in NG108-15 neuronal cells differentiated with cyclic AMP. We found OXC (0.3-30 microm) caused a reversible reduction in the amplitude of voltage-gated Na+ current (INa). The IC50 value required for the inhibition of INa by OXC was 3.1 microm. OXC (3 microm) could shift the steady-state inactivation of INa to a more negative membrane potential by approximately -9 mV with no effect on the slope of the inactivation curve, and produce a significant prolongation in the recovery of INa inactivation. Additionally, OXC was effective in suppressing persistent INa (INa(P)) elicited by long ramp pulses. The blockade of INa by OXC does not simply reduce current magnitude, but alters current kinetics. Moreover, OXC could suppress the amplitude of delayed rectifier K+ current (IK(DR)), with no effect on M-type K+ current (IK(M)). In current-clamp configuration, OXC could reduce the amplitude of action potentials and prolong action-potential duration. Furthermore, the simulations, based on hippocampal pyramidal neurons (Pinsky-Rinzel model) and a network of the Hodgkin-Huxley model, were analysed to investigate the effect of OXC on action potentials. Taken together, our results suggest that the synergistic blocking effects on INa and IK(DR) may contribute to the underlying mechanisms through which OXC affects neuronal function in vivo.

Glucose and hippocampal neuronal excitability: role of ATP-sensitive potassium channels.

Hyperglycemia-related neuronal excitability and epileptic seizures are not uncommon in clinical practice. However, their underlying mechanism remains elusive. ATP-sensitive K(+) (K(ATP)) channels are found in many excitable cells, including cardiac myocytes, pancreatic beta cells, and neurons. These channels provide a link between the electrical activity of cell membranes and cellular metabolism. We investigated the effects of higher extracellular glucose on hippocampal K(ATP) channel activities and neuronal excitability. The cell-attached patch-clamp configuration on cultured hippocampal cells and a novel multielectrode recording system on hippocampal slices were employed. In addition, a simulation modeling hippocampal CA3 pyramidal neurons (Pinsky-Rinzel model) was analyzed to investigate the role of K(ATP) channels in the firing of simulated action potentials. We found that incremental extracellular glucose could attenuate the activities of hippocampal K(ATP) channels. The effect was concentration dependent and involved mainly in open probabilities, not single-channel conductance. Additionally, higher levels of extracellular glucose could enhance neuropropagation; this could be attenuated by diazoxide, a K(ATP) channel agonist. In simulations, high levels of intracellular ATP, used to mimic increased extracellular glucose or reduced conductance of K(ATP) channels, enhanced the firing of action potentials in model neurons. The stochastic increases in intracellular ATP levels also demonstrated an irregular and clustered neuronal firing pattern. This phenomenon of K(ATP) channel attenuation could be one of the underlying mechanisms of glucose-related neuronal hyperexcitability and propagation.

The opening effect of pregabalin on ATP-sensitive potassium channels in differentiated hippocampal neuron-derived H19-7 cells.

PURPOSE: Adenosine triphosphate (ATP)-sensitive K+ (KATP) channels can couple an intracellular metabolic state to an electrical activity, which is important in the control of neuronal excitability and seizure propagation. We investigated whether the newer antiepileptic drug, pregabalin (PGB), could exert effects on KATP channels in differentiated hippocampal neuron-derived H19-7 cells. METHODS: The inside-out configuration of the patch-clamp technique was used to investigate KATP channel activities in H19-7 cells in the presence of PGB. Effects of various compounds known to alter KATP channel activities were compared. RESULTS: The activity of KATP channels in these cells was characterized. The single-channel conductance from a linear current-voltage relation was 78 +/- 2 pS (n = 8) with a reversal potential of 63 +/- 2 mV (n = 8), similar to that of KATP channels reported in pancreatic beta cells. 2,4-Dinitrophenol activated channel activity, but the further addition of glucose (20 mM) or glibenclamide (30 microM) could offset these increments. PGB significantly opened these KATP channel activities in a concentration-dependent fashion with a median effective concentration (EC50) value of 18 microM. A significant increase was noted in the mean open lifetime of KATP channels in the presence of PGB (1.71 +/- 0.04 to 5.62 +/- 0.04 ms). CONCLUSIONS: This study suggests that in differentiated hippocampal neuron-derived H19-7 cells, the opening effect on KATP channels could be one of the underlying mechanisms of PGB in the reduction of neuronal excitability.

Inhibitory action of ICI-182,780, an estrogen receptor antagonist, on BK(Ca) channel activity in cultured endothelial cells of human coronary artery.

ICI-182,780 is known to be a selective inhibitor of the intracellular estrogen receptors. The effect of ICI-182,780 on ion currents was studied in cultured endothelial cells of human coronary artery. In whole-cell current recordings, ICI-182,780 reversibly decreased the amplitude of K(+) outward currents. The decrease in outward current caused by ICI-182,780 could be counteracted by further application of magnolol or nordihydroguaiaretic acid, yet not by 17beta-estradiol. Under current-clamp condition, ICI-182,780 (3microM) depolarized the membrane potentials of the cells, and magnolol (10 microM) or nordihydroguaiaretic acid (10 microM) reversed ICI-182,780-induced depolarization. In inside-out patches, ICI-182,780 added to the bath did not alter single-channel conductance of large-conductance Ca(2+)-activated K(+) channels (BK(Ca) channels), but decreased their open probability. ICI-182,780 reduced channel activity in a concentration-dependent manner with an IC(50) value of 3 microM. After BK(Ca) channel activity was suppressed by 2-methoxyestradiol (3 microM), subsequent application of ICI-182,780 (3 microM) did not further reduce the channel activity. The application of ICI-182,780 shifted the activation curve of BK(Ca) channels to positive potentials. Its decrease in the open probability primarily involved a reduction in channel open duration. ICI-182,780 also suppressed the proliferation of these endothelial cells with an IC(50) value of 2 microM. However, in coronary smooth muscle cells, a bell-shaped concentration-response curve for the ICI-182,780 effect on BK(Ca) channel activity was observed. This study provides evidence that ICI-182,780 can inhibit BK(Ca) channels in vascular endothelial cells in a mechanism unlikely to be linked to its anti-estrogen activity. The inhibitory effects on these channels may partly contribute to the underlying mechanisms by which ICI-182,780 affects endothelial function.