Voltage-gated sodium channels are critical for the generation and conduction of nerve impulses. Recent studies show that in primary sensory neurons, the expression and dynamic regulation of several sodium channel subtypes play important roles in neuropathic pain. A number of SCN9A (encoding Nav1.7) gene point mutations are related with human genetic pain disorders. Transgenic and specific knockout techniques have revealed that Nav1.3, Nav1.8, Nav1.9 are important for the development and maintenance of neuropathic pain condition. Specific blockers of these sodium channels have been demonstrated to be effective in alleviating allodynia and hyperalgesia. Here we reviewed the roles of sodium channels in neuropathic pain, which may be applicable for the development of new drugs with enhanced efficacy for neuropathic pain treatment.
Animals ; Humans ; Neuralgia ; genetics ; metabolism ; physiopathology ; Neurons ; metabolism ; physiology ; Sodium Channels ; genetics ; metabolism ; physiology

Thiazide is known to decrease urinary calcium excretion. We hypothesized that thiazide shows different hypocalciuric effects depending on the stimuli causing hypercalciuria. The hypocalciuric effect of hydrochlorothiazide (HCTZ) and the expression of transient receptor potential vanilloid 5 (TRPV5), calbindin-D(28K), and several sodium transporters were assessed in hypercalciuric rats induced by high calcium diet and vitamin D3. Urine calcium excretion and the expression of transporters were measured from 4 groups of Sprague-Dawley rats; control, HCTZ, high calcium-vitamin D, and high calcium-vitamin D with HCTZ groups. HCTZ decreased urinary calcium excretion by 51.4% in the HCTZ group and only 15% in the high calcium-vitamin D with HCTZ group. TRPV5 protein abundance was not changed by HCTZ in the high calcium-vitamin D with HCTZ group compared to the high calcium-vitamin D group. Protein abundance of NHE3, SGLT1, and NKCC2 decreased in the hypercalciuric rats, and only SGLT1 protein abundance was increased by HCTZ in the hypercalciuric rats. The hypocalciuric effect of HCTZ is attenuated in high calcium and vitamin D-induced hypercalciuric rats. This attenuation seems to have resulted from the lack of HCTZ's effect on protein abundance of TRPV5 in severe hypercalciuric condition induced by high calcium and vitamin D.
Animals ; Calcium/therapeutic use/urine ; Calcium Channels/genetics/metabolism ; Cholecalciferol/*toxicity ; Hydrochlorothiazide/*therapeutic use ; Hypercalciuria/chemically induced/*drug therapy ; Rats ; Rats, Sprague-Dawley ; Sodium Chloride Symporter Inhibitors/*therapeutic use ; Sodium-Glucose Transporter 1/genetics/metabolism ; Sodium-Hydrogen Antiporter/genetics/metabolism ; Sodium-Potassium-Chloride Symporters/genetics/metabolism ; TRPV Cation Channels/genetics/metabolism

OBJECTIVETo observe the effects of chronic lead exposure on mRNA and protein expression of ASIC1a, ASIC2a, ASIC2b in hippocampus of baby-rats.

METHODSThe Wistar pregnant rats were randomly divided into 3 groups fed with distilled water or lead contained water (0.2% and 1.0% lead acetate) respectively, 5 rats in each group. The lead-exposure ranged from the 0 day of pregnancy to the offspring weaned. Then the baby-rats were fed with lead water like their mothers and killed at postnatal day 8 or 50. Atomic absorption spectrometry was used to determine lead content in the brain. RT-PCR and Western blotting were used to observe mRNA and protein expression of ASIC1a, ASIC2a and ASIC2b in their hippocampus respectively.

RESULTSThe brain lead content of test groups was higher than that of the control group (P < 0.01), and the lead content of the postnatal day 50 was higher than that in postnatal day 8 (P < 0.01). Compared with the control group, ASIC1a mRNA expression of 1.0% lead exposure in the hippocampus was uptrend (P < 0.01), ASIC1a protein expression of each test group was downtrend (P < 0.05), while for ASIC2a and ASIC2b mRNA and protein, there was no significant differences observed (P > 0.05).

CONCLUSIONASIC1a expression in hippocampus can be changed by chronic lead exposure.

Acid Sensing Ion Channels ; Animals ; Female ; Hippocampus ; drug effects ; metabolism ; Lead ; toxicity ; Nerve Tissue Proteins ; genetics ; metabolism ; Pregnancy ; Prenatal Exposure Delayed Effects ; genetics ; RNA, Messenger ; genetics ; Rats ; Rats, Wistar ; Sodium Channels ; genetics ; metabolism

OBJECTIVETo investigate the epithelial sodium channel (ENaC) subunit mRNA expression in acutely isolated rat alveolar type II (ATII) cells.

METHODSAcutely isolated ATII cells from 20 SD rats were purified and ENaC alpha, beta, gamma-subunit mRNA levels were determined by reverse transcription-polymerase chain reaction (RT-PCR).

RESULTSmRNA expressions of all the subunits were detected in the ATII cells, and ENaC alpha-subunit mRNA showed significantly higher expression than beta- and gamma-subunit mRNAs, and the expressions of the latter two mRNAs were comparable.

CONCLUSIONAs the predominant ENaC subunit expressed at the mRNA level in rat ATII cells, the alpha-subunit of ENAC plays an important role in alveolar fluid clearance.

Animals ; Cells, Cultured ; Epithelial Sodium Channels ; genetics ; metabolism ; Male ; Pulmonary Alveoli ; cytology ; metabolism ; RNA, Messenger ; genetics ; metabolism ; Rats ; Rats, Sprague-Dawley

OBJECTIVETo explore the protective mechanism of electroacupuncture (EA) at "Neiguan" (PC 6) on ischemic myocardial injury, and to explain the response patterns and characteristics of the specific effect of acupoints along meridians in sodium channel in the level of cardiac organ.

METHODSA total of 60 SPF male rats were randomly divided into a blank group, a model group, a non-acupoint group, a Neiguan group and a Lieque group, 12 cases in each one. Except the blank group, rats in the remaining group were treated with subcutaneous injection of isoprenaline to establish the model of myocardial ischemia. Rats in the Neiguan group, Lieque group and non- acupoint group were treated with EA, dilatational wave, with a frequency of 2 Hz/20 Hz. The intensity was 2-3 mA. The needles were retained for 20 min per time, once a day for consecutive 7 days. In the blank group and control group, the rats were grasped and fixed at the treating time each day. The western-blot method was used to test the expression of voltage-gated sodium channel alpha subunit (Nav 1.5), protein tyrosine kinase (PTKs) and protein tyrosine phosphatase (PTPs).

RESULTSThe expression of Nav 1.5 and PTKs in the model group was lower than that in the blank group (both P<0. 01); the expression in the Neiguan group and Lieque group was higher than that in the model group (all P < 0.01); the expression of Nav 1.5 and PTKs in the Neiguan group was higher than that in the Lieque group (both P < 0.01). The expression of PTPs in the model group and non-acupoint group was higher than that in the blank group (both P < 0.01); the expression of PTPs in the Neiguan group and Lieque group was significantly down-regulated, which was lower than the model group (both P < 0.01); the down-regulation in the Neiguan group was significantly different from that in the Lieque group (P < 0.05).

CONCLUSIONEA at "Neiguan" (PC 6), by down-regulating the expression of PTPs, up-regulating the expression of Nav 1.5 and PTKs, is likely to achieve the aim of regulation on sodium channel activity and calcium overload, further to improve myocardial ischemia, which provides experimental basis for the theory of the specific effect of acupoints along meridians.

Acupuncture Points ; Animals ; Disease Models, Animal ; Electroacupuncture ; Humans ; Male ; Myocardial Ischemia ; genetics ; metabolism ; therapy ; Myocardium ; metabolism ; Rats ; Rats, Sprague-Dawley ; Sodium Channels ; genetics ; metabolism

OBJECTIVETo investigate changes of two sodium channels, PN(3) and NaN, during orofacial pain by occlusal trauma in rat.

METHODSExpressions of PN(3) mRNA and NaN mRNA in trigeminal ganglion were tested during various periods of persistent occlusal trauma with reverse transcription-polymerase chain reaction (RT-PCR).

RESULTSIn groups, including control, occlusal trauma groups, PN(3) mRNA and NaN mRNA were all expressed in trigeminal ganglion neurons. In the control group, there were similar density values bilaterally. In the occlusal trauma group, the density values in gel electrophoresis of PN(3) mRNA and NaN mRNA on the intervention side were slightly greater than those on the control side.

CONCLUSIONSThe stimulation of occlusal trauma upregulates expressions of PN(3) mRNA and NaN mRNA, which suggests the signal occurring and conduction of chronic pain by occlusal trauma have the same molecular mechanism of sodium channel as inflammatory pain.

Animals ; Dental Occlusion, Traumatic ; physiopathology ; Facial Pain ; etiology ; Male ; NAV1.8 Voltage-Gated Sodium Channel ; RNA, Messenger ; biosynthesis ; Rats ; Rats, Sprague-Dawley ; Sodium Channels ; biosynthesis ; genetics ; Trigeminal Ganglion ; metabolism

Kv2.1 channel currents in pancreatic beta-cells are thought to contribute to action potential repolarization and thereby modulate insulin secretion. Because of its central role in this important physiological process, Kv2.1 channel is a promising target for the treatment of type 2 diabetes. Jingzhaotoxin-XI (JZTX-XI) is a novel peptide neurotoxin isolated from the venom of the spider Chilobrachys jingzhao. Two-microelectrode voltage clamp experiments had showed that the toxin inhibited Kv2.1 potassium currents expressed in Xenopus Laevis oocytes. In order to investigate the structure-function relationship of JZTX-XI, the natural toxin and a mutant of JZTX-XI in which Arg3 was replaced by Ala, were synthesized by solid-phase chemistry method with Fmoc-protected amino acids on the PS3 automated peptide synthesizer. Reverse-phase high performance liquid chromatography (RP-HPLC) and matrix assisted laser desorption/ ionization time-of-flight mass spectrometry (MALDI-TOF/TOF MS) were used to monitor the oxidative refolding process of synthetic linear peptides to find the optimal renaturation conditions of these toxins. The experiments also proved that the relative molecular masses of refolded peptides were in accordance with their theoretical molecular masses. RP-HPLC chromatogram of co-injected native and refolded JZTX-XI was a single peak. Under the whole-cell patch-clamp mode, JZTX-XI could completely inhibit hKv2.1 and hNav1.5 channels currents expressed in HEK293T cells with IC50 values of 95.8 nmol/L and 437.1 nmol/L respectively. The mutant R3A-JZTX-XI could also inhibit hKv2.1 and hNav1.5 channel currents expressed in HEK293T cells with IC50 values of 1.22 micromol/L and 1.96 micromol/L respectively. However, the prohibitive levels of R3A-JZTX-XI on hKv2.1 and hNav1.5 channels were reduced by about 12.7 times and 4.5 times respectively, indicating that Arg3 was a key amino acid residue relative to the hKv2.1 channel activity of JZTX-XI, but it is also an amino acid residue correlated with the binding activity of JZTX-XI to hNav1.5 channel. Our findings should be helpful to develop JZTX-XI into a molecular probe and drug candidate targeting to Kv2.1 potassium channel in the pancreas.
Animals ; HEK293 Cells ; Humans ; Insulin-Secreting Cells ; metabolism ; Mutant Proteins ; genetics ; pharmacology ; NAV1.5 Voltage-Gated Sodium Channel ; metabolism ; Neurotoxins ; chemical synthesis ; genetics ; pharmacology ; Protein Refolding ; Shab Potassium Channels ; antagonists & inhibitors ; metabolism ; Sodium Channel Blockers ; pharmacology ; Spider Venoms ; genetics ; pharmacology ; Transfection

OBJECTIVE: To explore the effect of nerve growth factor(NGF) and interferon regulatory factor-1(IRF-1) on sodium current change of sensory neuron in rat pheochromocytoma cells. METHODS: Sensory neuron rat pheochromocytoma cells were stimulated by different concentrations of NGF(0-200 ng/mL), the IRF-1 mRNA levels were examined by real-time PCR, and the activation of IRF-1 was examined by Western blot. The sodium current change was recorded by patch clamp. RESULTS: Low concentration of NGF improved the sodium current, which was concentration dependent. When exposed to high concentration of NGF, the expression of IRF-1 mRNA in PC-12 was improved. Low concentration of NGF resulted in IRF-1 intronuclear transporting, and the expression was not affected. Sodium current did not occur in PC-12 cells when IRF-1 was blocked. CONCLUSION NGF can improve the sodium current in PC-12 cells concentration-dependently, and the improvement is regulated by IRF-1.
Animals ; Interferon Regulatory Factor-1 ; genetics ; metabolism ; Nerve Growth Factor ; pharmacology ; PC12 Cells ; RNA, Messenger ; genetics ; metabolism ; Rats ; Real-Time Polymerase Chain Reaction ; Sodium Channels ; drug effects

OBJECTIVEThis study sought to isolate and identify the proteins that interact with ataxin-3, to confirm the interacted domain, and to provide new clues for exploring the function of ataxin-3 and the pathogenesis of spinocerebellar ataxia type 3 and Machado-Joseph disease (SCA3/MJD).

METHODSYeast two-hybrid screen (MATCHMAKER GAL4 Two-Hybrid System 3) and regular molecular biologic techniques were undertaken to screen human brain cDNA library with mutant ataxin-3 bait. Two baits from both normal and mutant C-terminus of ataxin-3 were created by subcloned methods to determine which domain of ataxin-3 interacts with the putative associated proteins and to find out optimal candidate proteins that interact with C-terminus of ataxin-3. Confocal microscope was used to observe whether ataxin-3 co-localized with the obtained interacting proteins in mammalian cells.

RESULTSFive novel ataxin-3 interacting proteins were obtained, among which were three known proteins, namely human rhodopsin guanosine diphosphate dissociation inhibitor alpha, small ubiquitin-like modifier 1, and human neuronal amiloride-sensitive cation channel 2; the other two were unknown. Interacting domain analysis revealed that an unknown protein interacted with the C-terminus near the polyglutamine tract of ataxin-3, the other four all interacted with the N-terminus. In the nucleus of SH-SY5Y cell, small ubiquitin-like modifier 1 co-localized with the wild-type ataxin-3 and with the intranuclear aggregates formed by the mutant ataxin-3.

CONCLUSIONAn unknown protein probably interacting with C-terminus of ataxin-3 is firstly discovered, and the initiative findings suggest first that the interaction of small ubiquitin-like modifier 1 with N-terminus of ataxin-3 and the relevant sumoylation probably participate in the post-translation modifying of ataxin-3 and in the pathogenesis of SCA3/MJD.

Acid Sensing Ion Channels ; Ataxin-3 ; Cell Line, Tumor ; Green Fluorescent Proteins ; genetics ; metabolism ; Humans ; Microscopy, Confocal ; Mutation ; Nerve Tissue Proteins ; genetics ; metabolism ; Nuclear Proteins ; genetics ; metabolism ; Plasmids ; genetics ; Protein Binding ; Recombinant Fusion Proteins ; genetics ; metabolism ; Repressor Proteins ; genetics ; metabolism ; SUMO-1 Protein ; genetics ; metabolism ; Sodium Channels ; genetics ; metabolism ; Transfection ; Two-Hybrid System Techniques

OBJECTIVETo observe the effects of ginsenoside Rg1 on the functional expression of human neural stem cells (hNSCs).

METHODThe membrane electrophysiological properties and sodium and potassium ion channels in the hNSCs induced by Rg1 were analyzed using the whole-cell patch-clamp.

RESULTOn the 7th day, the neuron-like cells derived from ginsenoside Rg1 (20 mg x L(-1))-induced NSCs show: (1) The resting membrane potential: (-45.70 +/- 2.63) mV, the membrane capacitance: (26.89 +/- 1.91) pF, the membrane input impedance: (877.51 +/- 20.44) MH (P < 0.05 compared with the control group, respectively); (2) The detection rate of inward sodium current which is rapidly activated and inactivated in voltage-dependence was 50%, and its average peak value was (711.48 +/- 158.03) pA (P < 0.05 compared with the control group); (3) The outward potassium currents were composed of rapidly activated and inactivated transient outward potassium current and delayed rectifier outward potassium current, and its average peak value was (1 070.42 +/- 177.18) pA (P < 0.05 compared with the control group).

CONCLUSIONGinsenoside Rg1 can promote the functional expression and maturity of hNSCs.

Cells, Cultured ; Gene Expression ; drug effects ; Ginsenosides ; pharmacology ; Humans ; Membrane Potentials ; drug effects ; Neural Stem Cells ; cytology ; drug effects ; Patch-Clamp Techniques ; Plant Extracts ; pharmacology ; Potassium Channels ; genetics ; metabolism ; Sodium Channels ; genetics ; metabolism