Cl Influx Research Articles

New GABA-Targeting Therapies for the Treatment of Seizures and Epilepsy: I. Role of GABA as a Modulator of Seizure Activity and Recently Approved Medications Acting on the GABA System.

γ-Aminobutyric acid (GABA) is the most prevalent inhibitory neurotransmitter in the mammalian brain and has been found to play an important role in the pathogenesis or the expression of many neurological diseases, including epilepsy. Although GABA can act on different receptor subtypes, the component of the GABA system that is most critical to modulation of seizure activity is the GABAA-receptor-chloride (Cl-) channel complex, which controls the movement of Cl- ions across the neuronal membrane. In the mature brain, binding of GABA to GABAA receptors evokes a hyperpolarising (anticonvulsant) response, which is mediated by influx of Cl- into the cell driven by its concentration gradient between extracellular and intracellular fluid. However, in the immature brain and under certain pathological conditions, GABA can exert a paradoxical depolarising (proconvulsant) effect as a result of an efflux of chloride from high intracellular to lower extracellular Cl- levels. Extensive preclinical and clinical evidence indicates that alterations in GABAergic inhibition caused by drugs, toxins, gene defects or other disease states (including seizures themselves) play a causative or contributing role in facilitating or maintaning seizure activity. Conversely, enhancement of GABAergic transmission through pharmacological modulation of the GABA system is a major mechanism by which different antiseizure medications exert their therapeutic effect. In this article, we review the pharmacology and function of the GABA system and its perturbation in seizure disorders, and highlight how improved understanding of this system offers opportunities to develop more efficacious and better tolerated antiseizure medications. We also review the available data for the two most recently approved antiseizure medications that act, at least in part, through GABAergic mechanisms, namely cenobamate and ganaxolone. Differences in the mode of drug discovery, pharmacological profile, pharmacokinetic properties, drug-drug interaction potential, and clinical efficacy and tolerability of these agents are discussed.

Zircon constraints on gold enrichment in a high-silica porphyry, Yixingzhai region, north of the Trans-North China Orogen

The Hewan granite porphyry is a newly discovered, highly evolved Au deposit in north China. Two types magmatic zircon are found in the porphyry. Type I is CL-dark and only occurs in the core of some zircons. Type II is CL-bright and mantles Type I zircon or occurs as individual grains. The two Types are close in age, i.e. 138.0 ± 2.2 Ma for Type I and 135.9 ± 2.1 Ma for Type II. Type I zircon is enriched in Y, U, Th, Nb, Ta, P, REE, while Type II zircon is relatively depleted in these elements, which reflects a reverse in the magma evolution. Whole-rock geochemistry and the presence of xenocrystic zircons (2522 ± 18 Ma), combined with εHf(t) (−20.2 to −22.3) and δ18O (5.3 to 6.6 ‰) for Type I zircons, suggests that the Hewan granite porphyry was mainly derived from the melting of the surrounding older, country rock gneiss. The higher εHf(t) (−16.5 to −19.4) and lower δ18O (5.0 to 6.0‰) of Type II zircon indicates that Type II zircon formed during infiltration of a later fluid. This later fluid also had a higher oxygen fugacity (Ce/Ce* Z = 248 to 3663) and a higher H2O activity. We propose that the Hewan granite porphyry originated from the shallow melting of regional gneisses due to heat from underplating mafic magmas related to a subduction event. Type I zircons, which formed during this event where later mantled or totally replaced by Type II zircon due to an influx of Cl-, S-, and Au-bearing fluids from these crystallizing underplating mafic magmas. The high F and pH in this shallow magma promoted decoupling of AuCl− in the upwelling fluids, which allowed for the deposition of elemental Au.

GABAergic Regulation of Astroglial Gliotransmission through Cx43 Hemichannels

Gamma-Aminobutyric acid (GABA) is the primary inhibitory neurotransmitter in the brain. It is produced by interneurons and recycled by astrocytes. In neurons, GABA activates the influx of Cl- via the GABAA receptor or efflux or K+ via the GABAB receptor, inducing hyperpolarization and synaptic inhibition. In astrocytes, the activation of both GABAA and GABAB receptors induces an increase in intracellular Ca2+ and the release of glutamate and ATP. Connexin 43 (Cx43) hemichannels are among the main Ca2+-dependent cellular mechanisms for the astroglial release of glutamate and ATP. However, no study has evaluated the effect of GABA on astroglial Cx43 hemichannel activity and Cx43 hemichannel-mediated gliotransmission. Here we assessed the effects of GABA on Cx43 hemichannel activity in DI NCT1 rat astrocytes and hippocampal brain slices. We found that GABA induces a Ca2+-dependent increase in Cx43 hemichannel activity in astrocytes mediated by the GABAA receptor, as it was blunted by the GABAA receptor antagonist bicuculline but unaffected by GABAB receptor antagonist CGP55845. Moreover, GABA induced the Cx43 hemichannel-dependent release of glutamate and ATP, which was also prevented by bicuculline, but unaffected by CGP. Gliotransmission in response to GABA was also unaffected by pannexin 1 channel blockade. These results are discussed in terms of the possible role of astroglial Cx43 hemichannel-mediated glutamate and ATP release in regulating the excitatory/inhibitory balance in the brain and their possible contribution to psychiatric disorders.

Silicate Inhibits the Cytosolic Influx of Chloride in Protoplasts of Wheat and Affects the Chloride Transporters, TaCLC1 and TaNPF2.4/2.5.

Chloride is an essential nutrient for plants, but high concentrations can be harmful. Silicon ameliorates both abiotic and biotic stresses in plants, but it is unknown if it can prevent cellular increase of chloride. Therefore, we investigated the influx of Cl− ions in two wheat cultivars different in salt sensitivity, by epifluorescence microscopy and a highly Cl−-sensitive dye, MQAE, N-[ethoxycarbonylmethyl]-6-methoxy-quinolinium bromide, in absence and presence of potassium silicate, K2SiO3. The Cl−-influx was higher in the salt-sensitive cv. Vinjett, than in the salt-tolerant cv. S-24, and silicate pre-treatment of protoplasts inhibited the Cl−-influx in both cultivars, but more in the sensitive cv. Vinjett. To investigate if the Cl−-transporters TaCLC1 and TaNPF2.4/2.5 are affected by silicate, expression analyses by RT-qPCR were undertaken of TaCLC1 and TaNPF 2.4/2.5 transcripts in the absence and presence of 100 mM NaCl, with and without the presence of K2SiO3. The results show that both transporter genes were expressed in roots and shoots of wheat seedlings, but their expressions were differently affected by silicate. The TaNPF2.4/2.5 expression in leaves was markedly depressed by silicate. These findings demonstrate that less chloride accumulates in the cytosol of leaf mesophyll by Si treatment and increases salt tolerance.

Phagosomal chloride dynamics in the alveolar macrophage

SummaryAcidification in intracellular organelles is tightly linked to the influx of Cl− counteracting proton translocation by the electrogenic V-ATPase. We quantified the dynamics of Cl− transfer accompanying cargo incorporation into single phagosomes in alveolar macrophages (AMs). Phagosomal Cl− concentration and acidification magnitude were followed in real time with maximal acidification achieved at levels of approximately 200 mM. Live cell confocal microscopy verified that phagosomal Cl− influx utilized predominantly the Cl− channel CFTR. Relative levels of elemental chlorine (Cl) in hard X-ray fluorescence microprobe (XFM) analysis within single phagosomes validated the increase in Cl− content. XFM revealed the complex interplay between elemental K content inside the phagosome and changes in Cl− during phagosomal particle uptake. Cl− -dependent changes in phagosomal membrane potential were obtained using second harmonic generation (SHG) microscopy. These studies provide a mechanistic insight for screening studies in drug development targeting pulmonary inflammatory disease.

State-dependent inhibition of GABA receptor channels by the ectoparasiticide fluralaner

γ-Aminobutyric acid (GABA) receptors (GABARs) are ligand-gated Cl− channels, which cause an influx of Cl− that inhibits excitation in postsynaptic cells upon activation. GABARs are important targets for drugs and pest control chemicals. We previously reported that the isoxazoline ectoparasiticide fluralaner inhibits GABA-induced currents in housefly (Musca domestica) GABARs by binding to the putative binding site in the transmembrane subunit interface. In the present study, we investigated whether fluralaner inhibits the GABA response in the GABAR activated state, the resting state, or both, using two-electrode voltage clamp electrophysiology protocols. We found that inhibition progresses over time to steady-state levels by repeated short applications of GABA during fluralaner perfusion. The GABA response was not impaired by fluralaner treatment in the GABAR resting state. However, once inhibited, the GABA response was not restored by repeated applications of GABA. These findings suggest that fluralaner might reach the binding site of the activated conformation of GABARs in a stepwise fashion and tightly bind to it.

Severe impairment of salivation in Na+/K+/2Cl- cotransporter (NKCC1)-deficient mice.

The salivary fluid secretory mechanism is thought to require Na(+)/K(+)/2Cl(-) cotransporter-mediated Cl(-) uptake. To directly test this possibility we studied the in vivo and in vitro functioning of acinar cells from the parotid glands of mice with targeted disruption of Na(+)/K(+)/2Cl(-) cotransporter isoform 1 (Nkcc1), the gene encoding the salivary Na(+)/K(+)/2Cl(-) cotransporter. In wild-type mice NKCC1 was localized to the basolateral membranes of parotid acinar cells, whereas expression was not detected in duct cells. The lack of functional NKCC1 resulted in a dramatic reduction (>60%) in the volume of saliva secreted in response to a muscarinic agonist, the primary in situ salivation signal. Consistent with defective Cl(-) uptake, a loss of bumetanide-sensitive Cl(-) influx was observed in parotid acinar cells from mice lacking NKCC1. Cl(-)/ HCO(3)(-) exchanger activity was increased in parotid acinar cells isolated from knockout mice suggesting that the residual saliva secreted by mice lacking NKCC1 is associated with anion exchanger-dependent Cl(-) uptake. Indeed, expression of the Cl(-)/ HCO(3)(-) exchanger AE2 was enhanced suggesting that this transporter compensates for the loss of functional Na(+)/K(+)/2Cl(-) cotransporter. Furthermore, the ability of the parotid gland to conserve NaCl was abolished in NKCC1-deficient mice. This deficit was not associated with changes in the morphology of the ducts, but transcript levels for the alpha-, beta-, and gamma-subunits of the epithelial Na(+) channel were reduced. These data directly demonstrate that NKCC1 is the major Cl(-) uptake mechanism across the basolateral membrane of acinar cells and is critical for driving saliva secretion in vivo.

Proton Transport through E. coli CLC Chloride/Proton Antiporter in the Presence of Bound Fluoride

A prototypical CLC antiporter from E. coli, EcCLC, regulates the influx of Cl− and efflux of H+ across membranes. Interestingly, it does not transport F-, even though F- has a smaller radius than Cl-. Here we apply multiscale quantum-mechanical/molecular-mechanical (QM/MM) simulations to study H+ migration via the Grotthuss mechanism through the transmembrane domain of EcCLC with F− present in the central binding site. Our preliminary data suggest that the F- does not prevent the protonation of the external gating residue E148, which rotates inward into the pore to receive the migrating H+. However, the protonated E148 is stabilized by F- in the pore through hydrogen-bonding interactions via 0-2 bridging water molecules. Some trajectories even show back and forth exchanges of H+ between the F- and E148. This picture differs significantly from our previous simulations of H+ transfer in the presence of a bound Cl-, where such strong interactions are not found. These strong interactions imply that it is difficult to separate the F- and protonated E148, therefore disrupting the antiporter operating cycle. Our results agree well with the “lock down” mechanism previously proposed by Miller and coworkers based on experiments (Lim, et al. Nat. Chem. Biol. 2013, 9, 721-725).

More Rapid Sleep Onset with Lingual-Spray vs Oral-Tablet Delivery Zolpidem

Insomnia and related sleep disorders (somnipathies) affect a large segment of the population, and result in a significant negative impact on quality of life and reduced or lost productivity. The speed of sleep onset is a critical characteristic of successful pharmacotherapeutic intervention for insomnia. Zolpidem, a non-benzodiazepine benzodiazepine receptor agonist (nBzRA) is widely used to treat insomnia. Although not itself a benzodiazepine (BZD), zolpidem has high binding affinity for the benzodiazepine receptor, which acts as a positive allosteric modulator of the GABAA receptor complex. It therefore increases the neuronal transmembrane influx of Cl- ions, thereby decreasing neuronal excitability and promoting sleep. In this four-way crossover, dose-ranging, multiple-treatment study, a lingual spray formulation of zolpidem was safe and well-tolerated and yielded more rapid pharmacokinetics (mean plasma concentration) and efficacy (visual analog scale and digit symbol substitution test) compared to oral tablets.

In silico toxicology protocols and software platforms

γ-Aminobutyric acid (GABA) receptors (GABARs) are ligand-gated Cl− channels, which cause an influx of Cl− that inhibits excitation in postsynaptic cells upon activation. GABARs are important targets for drugs and pest control chemicals. We previously reported that the isoxazoline ectoparasiticide fluralaner inhibits GABA-induced currents in housefly (Musca domestica) GABARs by binding to the putative binding site in the transmembrane subunit interface. In the present study, we investigated whether fluralaner inhibits the GABA response in the GABAR activated state, the resting state, or both, using two-electrode voltage clamp electrophysiology protocols. We found that inhibition progresses over time to steady-state levels by repeated short applications of GABA during fluralaner perfusion. The GABA response was not impaired by fluralaner treatment in the GABAR resting state. However, once inhibited, the GABA response was not restored by repeated applications of GABA. These findings suggest that fluralaner might reach the binding site of the activated conformation of GABARs in a stepwise fashion and tightly bind to it.

A role for the cystic fibrosis transmembrane conductance regulator in the nitric oxide-dependent release of Cl− from acidic organelles in amacrine cells

γ-Amino butyric acid (GABA) and glycine typically mediate synaptic inhibition because their ligand-gated ion channels support the influx of Cl- However, the electrochemical gradient for Cl- across the postsynaptic plasma membrane determines the voltage response of the postsynaptic cell. Typically, low cytosolic Cl- levels support inhibition, whereas higher levels of cytosolic Cl- can suppress inhibition or promote depolarization. We previously reported that nitric oxide (NO) releases Cl- from acidic organelles and transiently elevates cytosolic Cl-, making the response to GABA and glycine excitatory. In this study, we test the hypothesis that the cystic fibrosis transmembrane conductance regulator (CFTR) is involved in the NO-dependent efflux of organellar Cl- We first establish the mRNA and protein expression of CFTR in our model system, cultured chick retinal amacrine cells. Using whole cell voltage-clamp recordings of currents through GABA-gated Cl- channels, we examine the effects of pharmacological inhibition of CFTR on the NO-dependent release of internal Cl- To interfere with the expression of CFTR, we used clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 genome editing. We find that both pharmacological inhibition and CRISPR/Cas9-mediated knockdown of CFTR block the ability of NO to release Cl- from internal stores. These results demonstrate that CFTR is required for the NO-dependent efflux of Cl- from acidic organelles.NEW & NOTEWORTHY Although CFTR function has been studied extensively in the context of epithelia, relatively little is known about its function in neurons. We show that CFTR is involved in an NO-dependent release of Cl- from acidic organelles. This internal function of CFTR is particularly relevant to neuronal physiology because postsynaptic cytosolic Cl- levels determine the outcome of GABA- and glycinergic synaptic signaling. Thus the CFTR may play a role in regulating synaptic transmission.

A kainate receptor subunit promotes the recycling of the neuron-specific K+-Cl− co-transporter KCC2 in hippocampal neurons

Synaptic inhibition depends on a transmembrane gradient of chloride, which is set by the neuron-specific K+-Cl- co-transporter KCC2. Reduced KCC2 levels in the neuronal membrane contribute to the generation of epilepsy, neuropathic pain, and autism spectrum disorders; thus, it is important to characterize the mechanisms regulating KCC2 expression. In the present study, we determined the role of KCC2-protein interactions in regulating total and surface membrane KCC2 expression. Using quantitative immunofluorescence in cultured mouse hippocampal neurons, we discovered that the kainate receptor subunit GluK2 and the auxiliary subunit Neto2 significantly increase the total KCC2 abundance in neurons but that GluK2 exclusively increases the abundance of KCC2 in the surface membrane. Using a live cell imaging assay, we further determined that KCC2 recycling primarily occurs within 1-2 h and that GluK2 produces an ∼40% increase in the amount of KCC2 recycled to the membrane during this time period. This GluK2-mediated increase in surface recycling translated to a significant increase in KCC2 expression in the surface membrane. Moreover, we found that KCC2 recycling is enhanced by protein kinase C-mediated phosphorylation of the GluK2 C-terminal residues Ser-846 and Ser-868. Lastly, using gramicidin-perforated patch clamp recordings, we found that the GluK2-mediated increase in KCC2 recycling to the surface membrane translates to a hyperpolarization of the reversal potential for GABA (EGABA). In conclusion, our results have revealed a mechanism by which kainate receptors regulate KCC2 expression in the hippocampus.

Effects of whole-tree harvest on soil-water chemistry at five conifer sites in Sweden

Logging residues, such as tops and branches, can provide a useful biofuel for large-scale energy production. However, increasing the harvest intensity may affect the soil nutrient stores and water quality. Here, effects on soil-water chemistry after stem-only and whole-tree harvesting were investigated using data from five experimental sites in Sweden, representing medium- to high-fertility sites. They were located in recharge areas on mineral soil and harvested between the years 1995 and 2001. Soil-water samples had previously been collected from below the main part of the root zone in study plots subjected to stem-only or whole-tree harvest. Soil-water chemistry data from the five sites were jointly analyzed by ANOVA using seasonal mean concentrations from the first six seasons after clear-cutting. The concentrations of NO3–-N, K+, and Mg2+were significantly different (p < 0.05) between the two harvest regimes, indicating lower levels after whole-tree harvest than after stem-only harvest. No significant differences were detected for electrical conductivity, pH, or the concentrations of NH4+-N, Al, Ca2+, SO42–-S, and Cl–. Measurements at one site suggested that the logging residues left on the ground increased the influx of Cl–to the soil. Simple budget calculations indicated that the nutrient export by logging-residue harvest was greater than the export by leaching during the regeneration phase.

Neonatal seizures and disruption to neurotransmitter systems

Neonatal seizures and disruption to neurotransmitter systems

Activation of band 3 mediates group A Streptococcus streptolysin S-based beta-haemolysis.

Streptococcus pyogenes, or group A Streptococcus (GAS), is a human bacterial pathogen that can manifest as a range of diseases from pharyngitis and impetigo to severe outcomes such as necrotizing fasciitis and toxic shock syndrome. GAS disease remains a global health burden with cases estimated at over 700 million annually and over half a million deaths due to severe infections(1). For over 100 years, a clinical hallmark of diagnosis has been the appearance of complete (beta) haemolysis when grown in the presence of blood. This activity is due to the production of a small peptide toxin by GAS known as streptolysin S. Although it has been widely held that streptolysin S exerts its lytic activity through membrane disruption, its exact mode of action has remained unknown. Here, we show, using high-resolution live cell imaging, that streptolysin S induces a dramatic osmotic change in red blood cells, leading to cell lysis. This osmotic change was characterized by the rapid influx of Cl(-) ions into the red blood cells through SLS-mediated disruption of the major erythrocyte anion exchange protein, band 3. Chemical inhibition of band 3 function significantly reduced the haemolytic activity of streptolysin S, and dramatically reduced the pathology in an in vivo skin model of GAS infection. These results provide key insights into the mechanism of streptolysin S-mediated haemolysis and have implications for the development of treatments against GAS.

Lotus Leaf Alkaloid Extract Displays Sedative-Hypnotic and Anxiolytic Effects through GABAA Receptor.

Lotus leaves have been used traditionally as both food and herbal medicine in Asia. Open-field, sodium pentobarbital-induced sleeping and light/dark box tests were used to evaluate sedative-hypnotic and anxiolytic effects of the total alkaloids (TA) extracted from the herb, and the neurotransmitter levels in the brain were determined by ultrafast liquid chromatography-tandem mass spectrometry. The effects of picrotoxin, flumazenil, and bicuculline on the hypnotic activity of TA, as well as the influence of TA on Cl(-) influx in cerebellar granule cells, were also investigated. TA showed a sedative-hypnotic effect by increasing the brain level of γ-aminobutyric acid (GABA), and the hypnotic effect could be blocked by picrotoxin and bicuculline, but could not be antagonized by flumazenil. Additionally, TA could increase Cl(-) influx in cerebellar granule cells. TA at 20 mg/kg induced anxiolytic-like effects and significantly increased the concentrations of serotonin (5-HT), 5-hydroxyindoleacetic acid (5-HIAA), and dopamine (DA). These data demonstrated that TA exerts sedative-hypnotic and anxiolytic effects via binding to the GABAA receptor and activating the monoaminergic system.

Pharmacological inhibition of cation-chloride cotransporters for neurological diseases.

γ-Aminobutyric acid (GABA) is a major neurotransmitter and plays important roles in both the developing and mature central nervous system (CNS). One way that GABA can act is by binding to fast, ionotropic GABAA receptors in neurons. The binding of GABA to GABAA receptors causes a conformational change that opens ion channels, allowing the passage of Cl– ions. In mature adults, intracellular Cl– concentration [Cl–]i) is low, and the opening of these ion channels triggers influx of Cl– ions, causing hyperpolarization and neuronal inhibition (Blaesse, 2009). However, in the developing nervous system, [Cl–]i is high, and the binding of GABA to GABAA receptors induces a depolarizing excitatory response (Figure 1). This leads to the stimulation of voltage-dependent Ca2+ channels, which is important for proper neuronal proliferation and differentiation in their circuitry development. Gradual changes in [Cl–]i during development, known as the “GABA shift”, determine the strength and polarity of GABA-mediated activity. The precise regulation of [Cl–]i is determined by two major cation-chloride cotransporters (CCCs), the inwardly directed Na-K-2Cl cotransporter (NKCC1) and the Cl– extruding K-Cl cotransporter (KCC2) (Blaesse, 2009). NKCC1 is expressed in a wide range of neurons and glia, but especially in embryonic ventricular zones, which suggests an important role in neuronal proliferation. KCC2 is located in plasma membrane of both somata and dendrites of neurons in various brain regions, including cortical pyramidal neurons, thalamic relay cells, and auditory brainstem neurons (Blaesse, 2009). In embryonic and early postnatal life, neurons show high expression of NKCC1and low expression of KCC2 (Figure 1). NKCC1 functions in Cl– influx and raises the [Cl–]i during the early development of the CNS, which enables GABAA receptor-mediated neuronal depolarization and excitation. NKCC1 expression rapidly decreases during the first year of life to levels common in an adult, while KCC2 expression rises (Dzhala et al., 2005). KCC2 functions in Cl– efflux and lowering [Cl–]i, which leads to the dominance of GABAA receptor-mediated hyperpolarization and inhibition response in the maturing brain (Figure 1).

Mechanisms of Na+ uptake, ammonia excretion, and their potential linkage in native Rio Negro tetras (Paracheirodon axelrodi, Hemigrammus rhodostomus, and Moenkhausia diktyota).

Mechanisms of Na(+) uptake, ammonia excretion, and their potential linkage were investigated in three characids (cardinal, hemigrammus, moenkhausia tetras), using radiotracer flux techniques to study the unidirectional influx (J in), efflux (J out), and net flux rates (J net) of Na(+) and Cl(-), and the net excretion rate of ammonia (J Amm). The fish were collected directly from the Rio Negro, and studied in their native "blackwater" which is acidic (pH 4.5), ion-poor (Na(+), Cl(-) ~20µM), and rich in dissolved organic matter (DOM11.5mg Cl(-1)). J in (Na) , J in (Cl) , and J Amm were higher than in previous reports on tetras obtained from the North America aquarium trade and/or studied in low DOM water. In all three species, J in (Na) was unaffected by amiloride (10(-4)M, NHE and Na(+) channel blocker), but both J in (Na) and J in (Cl) were virtually eliminated (85-99% blockade) by AgNO3 (10(-7)M). A time course study on cardinal tetras demonstrated that J in (Na) blockade by AgNO3 was very rapid (<5min), suggesting inhibition of branchial carbonic anhydrase (CA), and exposure to the CA-blocker acetazolamide (10(-4)M) caused a 50% reduction in J in (Na) .. Additionally, J in (Na) was unaffected by phenamil (10(-5)M, Na(+) channel blocker), bumetanide (10(-4)M, NKCC blocker), hydrochlorothiazide (5×10(-3)M, NCC blocker), and exposure to an acute 3 unit increase in water pH. None of these treatments, including partial or complete elimination of J in (Na) (by acetazolamide and AgNO3 respectively), had any inhibitory effect on J Amm. Therefore, Na(+) uptake in Rio Negro tetras depends on an internal supply of H(+) from CA, but does not fit any of the currently accepted H(+)-dependent models (NHE, Na(+) channel/V-type H(+)-ATPase), or co-transport schemes (NCC, NKCC), and ammonia excretion does not fit the current "Na(+)/NH4 (+) exchange metabolon" paradigm. Na(+), K(+)-ATPase and V-type H(+)-ATPase activities were present at similar levels in gill homogenates, Acute exposure to high environmental ammonia (NH4Cl, 10(-3)M) significantly increased J in (Na) , and NH4 (+) was equally or more effective than K(+) in activating branchial Na(+),(K(+)) ATPase activity in vitro. We propose that ammonia excretion does not depend on Na(+) uptake, but that Na(+) uptake (by an as yet unknown H(+)-dependent apical mechanism) depends on ammonia excretion, driven by active NH4 (+) entry via basolateral Na(+),(K(+))-ATPase.

Mechanisms of action of general anaesthetic drugs

Based on the diverse array of anaesthetic structures, a single anaesthetic target site seems unlikely. With the knowledge that anaesthesia likely results from central nervous system depression, it can be hypothesized that anaesthesia results from either enhanced inhibitory transmission or reduced excitatory transmission. Two main targets have been extensively described: GABAA receptors and N-methyl-d-aspartate (NMDA) glutamate receptors. On γ-aminobutyric acid (GABA) binding to GABAA receptors, an influx of Cl− results to produce hyperpolarization. With the exception of ketamine, xenon and nitrous oxide, all anaesthetic agents potentiate GABA-mediated conductance. On binding of the main excitatory transmitter glutamate, NMDA receptors gate an influx of Ca2+ and Na+. Ketamine, xenon and nitrous oxide inhibit this ion movement to depress excitatory transmission.

Ginseng ginsenoside pharmacology in the nervous system: involvement in the regulation of ion channels and receptors

Ginseng, the root of Panax ginseng C.A. Meyer, is one of the oldest traditional medicines and is thought to be a tonic. It has been claimed that ginseng may improve vitality and health. Recent studies have advanced ginseng pharmacology and shown that ginseng has various pharmacological effects in the nervous system. Ginsenosides, steroid glycosides extracted from ginseng, were one of the first class of biologically active plant glycosides identified. The diverse pharmacological effects of ginsenosides have been investigated through the regulation of various types of ion channels and receptors in neuronal cells and heterologous expression systems. Ginsenoside Rg3 regulates voltage-gated ion channels such as Ca2+, K+, and Na+ channels, and ligand-gated ion channels such as GABAA, 5-HT3, nicotinic acetylcholine, and N-methyl-D-aspartate (NMDA) receptors through interactions with various sites including channel blocker binding sites, toxin-binding sites, channel gating regions, and allosteric channel regulator binding sites when the respective ion channels or receptors are stimulated with depolarization or ligand treatment. Treatment with ginsenoside Rg3 has been found to stabilize excitable cells by blocking influxes of cations such as Ca2+ and Na+, or by enhancing Cl− influx. The aim of this review is to present recent findings on the pharmacological functions of the ginsenosides through the interactions with ion channels and receptors. This review will detail the pharmacological applications of ginsenosides as neuroprotective drugs that target ion channels and ligand-gated ion channels.