Concentrations Of Tetraethylammonium Research Articles

Transport of tetraethylammonium by the Malpighian tubules of Trichoplusia ni: Regional specialization and the influence of diet

Insect Malpighian tubules (MTs) play a major role in elimination of many potentially toxic compounds, including the organic cation tetraethylammonium (TEA). This paper examines transport of TEA by different segments of the MTs of the cabbage looper, Trichoplusia ni. The results show that the proximal ileac plexus (PIP) region of the MTs plays a dominant role in secretion of the organic cation TEA and that the rate of secretion is altered by feeding; principal cells of the proximal ileac plexus in tubules from larvae with full guts secreted TEA at higher rates than did the same cells in tubules of larvae in which the gut was empty. Michaelis-Menten analysis revealed that TEA secretion by the PIP was saturable and was blocked in a concentration-dependent manner by the organic cation cimetidine. For larvae reared from eggs on TEA-rich diet, higher concentrations of TEA in fluid secreted by the ileac plexus of tubules, and lower concentrations of TEA in the hemolymph, relative to larvae reared on control diet, is consistent with an upregulation of TEA transport in response to higher levels of dietary intake of an exogenous organic cation. The distal and proximal regions of the ileac plexus were also differentiated on the basis of transepithelial and basolateral membrane potentials and the influence of these electrical potentials on organic cation transport are discussed.

Reduction of Cd(II) Ions in the Presence of Tetraethylammonium Cations. Adsorption Effect on the Electrode Process

The effect of the adsorption of tetraethylammonium (TEA) cations, which present both ionic and organic characteristics, on the reduction of Cd(II) ions have been studied from dc and ac measurements at the dropping mercury electrode. The resistance to the charge transfer (Rct) and Warburg coefficient (σ) parameters have been determined through impedance measurements. Thus, the global velocity constant has been obtained. The reduction process of Cd(II) in perchloric media is reversible and is affected by the adsorption of TEA cations, especially at high TEA concentrations. Values of E1/2, half wave potential, and DO, diffusion coefficient, obtained from both dc and ac measurements agree. The velocity constants show a decrease as TEA concentration increases, with values ranging from 0.6 to 0.01 cm·s−1. The inhibitory effect of TEA adsorption on the electrode process and the relationship between electrode coverage, θ, and velocity constants, K, using several isotherm equations, have been discussed. The best fit was obtained with the equation K = 0K(1 − θ)a with an a value close to three, indicating a blocking effect and electrostatic repulsion due to TEA.

Modulatory Effects of Perineuronal Oligodendrocytes on Neuronal Activity in the Rat Hippocampus.

Action potentials are fundamental to relaying information from region to region in the nervous system. Changes in action potential firing patterns in neural circuits influence how the brain processes information. In our previous study, we focused on interneuron/perineuronal astrocyte pairs in the hippocampal CA1 region and reported that direct depolarization of perineuronal astrocytes modulated the firing pattern of interneurons. In the current study, we investigated the morphological and electrophysiological properties of perineuronal oligodendrocytes, and examined their modulatory effects on interneuronal firing in the CA1 region. Perineuronal oligodendrocytes only had a few processes, which were crooked, intricately twisted, and twined around the soma and proximal region of the main processes of adjacent interneurons. Whole-cell current patterns of perineuronal oligodendrocytes were homogenous and the current-voltage relationship showed remarkable outward rectification. Although the K+ channel blockers, tetraethylammonium and 4-aminopyridine, clearly blocked outward currents, Ba2+ did not significantly alter whole-cell currents. Unlike perineuronal astrocytes, the depolarization of perineuronal oligodendrocytes had no effect on interneuronal firing; however, when the interneurons were firing at a higher frequency, the hyperpolarization of perineuronal oligodendrocytes suppressed their action potentials. The suppressive effects of perineuronal oligodendrocytes were inhibited in the presence of a low concentration of tetraethylammonium, which selectively blocked deep and fast afterhyperpolarization. These results suggest that perineuronal oligodendrocytes suppress interneuronal firing through their influence on K+ channels, which are responsible for deep and fast afterhyperpolarization.

Sensory transduction at the frog semicircular canal: how hair cell membrane potential controls junctional transmission.

At the frog semicircular canals, the afferent fibers display high spontaneous activity (mEPSPs), due to transmitter release from hair cells. mEPSP and spike frequencies are modulated by stimulation that activates the hair cell receptor conductance. The relation between receptor current and transmitter release cannot be studied at the intact semicircular canal. To circumvent the problem, we combined patch-clamp recordings at the isolated hair cell and electrophysiological recordings at the cytoneural junction in the intact preparation. At isolated hair cells, the K channel blocker tetraethylammonium (TEA) is shown to block a fraction of total voltage-dependent K-conductance (IKD) that depends on TEA concentration but not on membrane potential (Vm). Considering the bioelectric properties of the hair cell, as previously characterized by this lab, a fixed fractional block of IKD is shown to induce a relatively fixed shift in Vm, provided it lies in the range −30 to −10 mV. The same concentrations of TEA were applied to the intact labyrinth while recording from single afferent fibers of the posterior canal, at rest and during mechanical stimulation. At the peak of stimulation, TEA produced increases in mEPSP rate that were linearly related to the shifts produced by the same TEA concentrations (0.1–3 mM) in hair cell Vm (0.7–5 mV), with a slope of 29.8 Hz/mV. The membrane potential of the hair cell is not linearly related to receptor conductance, so that the slope of quantal release vs. receptor conductance depends on the prevailing Vm (19.8 Hz/nS at −20 mV; 11 Hz/nS at −10 mV). Changes in mEPSP peak size were negligible at rest as well as during stimulation. Since ample spatial summation of mEPSPs occurs at the afferent terminal and threshold-governed spike firing is intrinsically nonlinear, the observed increases in mEPSP frequency, though not very large, may suffice to trigger afferent spike discharge.

A Novel Modulator of Kv3 Potassium Channels Regulates the Firing of Parvalbumin-Positive Cortical Interneurons.

Kv3.1 and Kv3.2 high voltage-activated potassium channels, which display fast activation and deactivation kinetics, are known to make a crucial contribution to the fast-spiking phenotype of certain neurons. Pharmacological experiments show that the blockade of native Kv3 currents with low concentrations of tetraethylammonium or 4-aminopyridine impairs the expression of this firing phenotype. In particular, Kv3 channels are highly expressed by fast-spiking, parvalbumin-positive interneurons in corticolimbic brain circuits, which modulate the synchronization of cortical circuits and the generation of brain rhythms. Here, we describe a novel small molecule, (5R)-5-ethyl-3-(6-{[4-methyl-3-(methyloxy)phenyl]oxy}-3-pyridinyl)-2,4-imidazolidinedione (AUT1), which modulates Kv3.1 and Kv3.2 channels in human recombinant and rodent native neurons. AUT1 increased whole currents mediated by human Kv3.1b and Kv3.2a channels, with a concomitant leftward shift in the voltage dependence of activation. A less potent effect was observed on hKv3.3 currents. In mouse somatosensory cortex slices in vitro, AUT1 rescued the fast-spiking phenotype of parvalbumin-positive-fast-spiking interneurons following an impairment of their firing capacity by blocking a proportion of Kv3 channels with a low concentration of tetraethylammonium. Notably, AUT1 had no effect on interneuron firing when applied alone. Together, these data confirm the role played by Kv3 channels in the regulation of the firing phenotype of somatosensory interneurons and suggest that AUT1 and other Kv3 modulators could represent a new and promising therapeutic approach to the treatment of disorders associated with dysfunction of inhibitory feedback in corticolimbic circuits, such as schizophrenia.

L-Type Ca(2+) Channels and SK Channels in Mouse Embryonic Stem Cells and Their Contribution to Cell Proliferation.

Mouse embryonic stem cells (mESCs) are capable of both self-renewal and multilineage differentiation; thus, they can be expanded in vivo or in vitro and differentiated to produce different cell types. Despite their biological and medical interest, many physiological properties of undifferentiated mESCs, such as ion channel function, are not fully understood. Ion channels are thought to be involved in cell proliferation and differentiation. The aim of this study was to characterize functional ion channels in cultured undifferentiated mESCs and their role in cell proliferation. L-type voltage-activated Ca(2+) channels sensitive to nifedipine and small-conductance Ca(2+)-activated K(+) (SK) channels sensitive to apamin were identified. Ca(2+)-activated K(+) currents were blocked by millimolar concentrations of tetraethylammonium. The effects of Ca(2+) channel and Ca(2+)-activated K(+) channel blockers on the proliferation of undifferentiated mESCs were investigated by bromodeoxyuridine (BrdU) incorporation. Dihydropyridine derivatives, such as nifedipine, inhibited cell growth and BrdU incorporation into the cells, whereas apamin, which selectively blocks SK channels, had no effect on cell growth. These results demonstrate that functional voltage-operated Ca(2+) channels and Ca(2+)-activated K(+) channels are present in undifferentiated mESCs. Moreover, voltage-gated L-type Ca(2+) channels, but not SK channels, might be necessary for proliferation of undifferentiated mESCs.

Potassium-induced bifurcations and chaos of firing patterns observed from biological experiment on a neural pacemaker

Changes of neural firing patterns and transitions between firing patterns induced by the introduction of external stimulation or adjustment of biological parameter have been demonstrated to play key roles in information coding. In this paper, bifurcation processes of bursting patterns were observed from an experimental neural pacemaker, through the adjustment of potassium parameter including ion concentration and calcium-dependent channel conductance. The adjustment of calcium-dependent potassium channel conductance was achieved by changing the extracellular tetraethylammonium concentration. The deterministic dynamics of chaotic bursting patterns induced by period-doubling bifurcation and intermittency, and lying between two periodic bursting patterns in a period-adding bifurcation process was investigated with a nonlinear prediction method. The bifurcations included period-doubling and period-adding bifurcations of bursting patterns. The experimental bifurcations and chaos closely matched those previously simulated in the theoretical neuronal model by adjusting potassium parameter, which demonstrated the simulation results of the theoretical model. The experimental results indicate that the potassium concentration and conductance of calcium-dependent potassium channel can induce bifurcations of the neural firing patterns. The potential role of these bifurcation structures in neural information coding mechanism is discussed.

P2Y1 receptor modulation of Ca2+-activated K+ currents in medium-sized neurons from neonatal rat striatal slices

ATP signaling to neurons and glia in the nervous system occurs via activation of both P2Y and P2X receptors. Here, we investigated the effects of P2Y1 receptor stimulation in developing striatal medium-sized neurons using patch-clamp recordings from acute brain slices of 7- and 28-day-old rats. Application of the selective P2Y1 receptor agonist 2-(Methylthio) ADP trisodium salt (2-MeSADP; 250 nM) increased outward K+ currents evoked by a ramp depolarization protocol in voltage-clamp recordings. This effect was observed in 59 out of 82 cells (72%) and was blocked completely by the P2Y1 antagonist, 2′-deoxy-N6-methyl adenosine 3′,5′-diphosphate. The averaged 2-MeSADP-sensitive conductance was fitted by the sum of a linear conductance and a Boltzmann relation, giving one-half activation voltage of −14.2 mV and an equivalent charge of 2.91. The 2MeSADP-mediated effect was sensitive to submillimolar concentrations of tetraethylammonium (TEA; 200 μM), to 200 nM iberiotoxin and to 100 nM apamin, suggesting the involvement of both big and small potassium (BK and SK, respectively) calcium-activated channels. In current-clamp experiments, 2-MeSADP decreased depolarization-evoked action potential (AP) firing in all 26 cells investigated, and this effect was reversed by TEA and by apamin but not by iberiotoxin. We conclude that the stimulation of P2Y1 receptors in developing striatal neurons leads to activation of calcium-activated potassium channels [IK(Ca)] of both BK and SK subtypes, the latter responsible for decreasing the frequency of AP firing in response to current injection. Therefore, P2Y1 signaling leading to activation of IK(Ca) may be important in regulating the activity of medium-sized neurons in the striatum.

Two‐pore domain K+ channels regulate membrane potential of isolated human articular chondrocytes

Potassium channels that regulate resting membrane potential (RMP) of human articular chondrocytes (HACs) of the tibial joint maintained in short-term (0-3 days) non-confluent cell culture were studied using patch-clamp techniques. Quantitative PCR showed that transcripts of genes for two-pore domain K(+) channels (KCNK1, KCNK5 and KCNK6), and 'BK' Ca(2+)-activated K(+) channels (KCNMA1) were abundantly expressed. Immunocytological methods detected α-subunits for BK and K(2p)5.1 (TASK-2) K(+) channels. Electrophysiological recordings identified three distinct K(+) currents in isolated HACs: (i) a voltage- and time-dependent 'delayed rectifier', blocked by 100 nM α-dendrotoxin, (ii) a large 'noisy' voltage-dependent current that was blocked by low concentrations of tetraethylammonium (TEA; 50% blocking dose = 0.15 mM) and iberiotoxin (52% block, 100 nM) and (iii) a voltage-independent 'background' K(+) current that was blocked by acidic pH (5.5-6), was increased by alkaline pH (8.5), and was not blocked by TEA, but was blocked by the local anaesthetic bupivacaine (0.25 mM). The RMP of isolated HACs was very slightly affected by 5 mM TEA, which was sufficient to block both voltage-dependent K(+) currents, suggesting that these currents probably contributed little to maintaining RMP under 'resting' conditions (i.e. low internal [Ca(2+)]). Increases in external K(+) concentration depolarized HACs by 30 mV in response to a 10-fold increase in [K(+)], indicating a significant but not exclusive role for K(+) current in determining RMP. Increases in external [K(+)] in voltage-clamped HACs revealed a voltage-independent K(+) current whose inward current magnitude increased with external [K(+)]. Block of this current by bupivacaine (0.25-1 mM) in 5 and 25 mM external [K(+)] resulted in a large (8-25 mV) depolarization of RMP. The biophysical and pharmacological properties of the background K(+) current, together with expression of mRNA and α-subunit protein for TASK-2, strongly suggest that these two-pore domain K(+) channels contribute significantly to stabilizing the RMP of HACs.

Voltage-gated potassium channel in a human skin squamous cell carcinoma cell line A431

Objective To investigate the role of voltage-gated potassium channel in the human skin squamous cell carcinoma A431 cells and human keratinocyte HaCaT cells. Methods MTT assay was performed to detect the effect of different concentrations of tetraethylammonium (TEA) on the proliferation of cultured A431 cells and HaCaT cells. Besides, enzyme-linked immunosorbent assay (ELISA) was conducted to detect the expression of HERG channel protein in A431 cells, HaCaT cells and normal human skin tissue.Results TEA inhibited the proliferation of A431 cells and HaCaT cells in a dose- and time-dependent manner.After treated with TEA (≥ 10 mmol/L) for 24 or more hours, the proliferation of A431 cells and HaCaT cells was obviously suppressed. A significant difference was observed in the average concentration of HERG channel protein between A431 cells, HaCaT cells and normal human skin tissue (49.7114 ± 3.55696 pg/ml, 35.7471 ±4.14696 pg/ml, 36.8857 ± 3.47810 pg/ml, all P < 0.05). Conclusions The block of voltage-gated potassium channel could inhibit the proliferation of A431 cells and HaCaT cells, and the expression of voltage-gated potassium channel seems to be higher in human skin squamous cell carcinoma cells. Key words: Neoplasms, squamous cell; Potassium channels, voltage-gated; Keratinocytes

Role of Calcium and Potassium Channels in Effects of Hydrogen Sulfide on Frog Myocardial Contractility

The effects of sodium hydrosulfide NaHS, a donor of hydrogen sulfide H2S, on the force of muscle contraction were examined on isolated myocardial strips from frog ventricles. NaHS decreased the amplitude of muscle contractions in a dose-dependent manner under normal conditions and during inhibition of Ca channels with nifedipine. In contrast, under conditions of blockade of ATP-dependent potassium channels with glibenclamide, NaHS exerted a positive inotropic effect from the first minute of application. Neither blockade, nor activation of ATP-dependent K-channels with glibenclamide modulated the negative inotropic effect of NaHS. Inhibition of K-channels with tetraethylammonium (TEA) (3, 5, 10 mM) or 4-aminopyridine increased the amplitude of myocardial contractions. Preliminary application of 4-aminopyridine or TEA (3 mM) did not eliminate NaHS-induced negative inotropic effect, although higher TEA concentrations (5 or 10 mM) prevented it. The data indicate that the targets of H(2)S in frog myocardium are ATP-dependent, Ca-activated, and voltage-dependent K-channels.

Allosteric Block of KCa2 Channels by Apamin

Activation of small conductance calcium-activated potassium (K(Ca)2) channels can regulate neuronal firing and synaptic plasticity. They are characterized by their high sensitivity to the bee venom toxin apamin, but the mechanism of block is not understood. For example, apamin binds to both K(Ca)2.2 and K(Ca)2.3 with the same high affinity (K(D) approximately 5 pM for both subtypes) but requires significantly higher concentrations to block functional current (IC(50) values of approximately 100 pM and approximately 5 nM, respectively). This suggests that steps beyond binding are needed for channel block to occur. We have combined patch clamp and binding experiments on cell lines with molecular modeling and mutagenesis to gain more insight into the mechanism of action of the toxin. An outer pore histidine residue common to both subtypes was found to be critical for both binding and block by the toxin but not for block by tetraethylammonium (TEA) ions. These data indicated that apamin blocks K(Ca)2 channels by binding to a site distinct from that used by TEA, supported by a finding that the onset of block by apamin was not affected by the presence of TEA. Structural modeling of ligand-channel interaction indicated that TEA binds deep within the channel pore, which contrasted with apamin being modeled to interact with the channel outer pore by utilizing the outer pore histidine residue. This multidisciplinary approach suggested that apamin does not behave as a classical pore blocker but blocks using an allosteric mechanism that is consistent with observed differences between binding affinity and potency of block.

Functional expression of a large-conductance Ca 2+-activated K + channel in mouse substantia nigra pars compacta dopaminergic neurons

The existence of large-conductance Ca 2+-activated K + (BK) channels in substantia nigra pars compacta (SNc) has been a matter of debate. Using the patch-clamp technique in the inside-out configuration, we have recorded BK channel currents in SNc dopaminergic neurons. The channel has a conductance of 301 pS with a slight inward rectification and is both voltage- and calcium-dependent. Paxilline, a specific BK channel blocker, can completely block the channel, while tetraethylammonium (TEA), a nonspecific blocker of voltage-gated potassium channels, reduces its conductance and a high concentration of TEA (30 mM) inhibits its activity. ATP and GTP reduce the channel activity, while ADP is less potent, and AMP has no effect. The channel is also sensitive to changes in intracellular pH. Our results indicate that functional BK channels are expressed in SNc and suggest the possibility that the BK channel may be involved in the response of SNc dopaminergic neurons to metabolic stress.

Dual effect of exogenous hydrogen sulfide on the spontaneous contraction of gastric smooth muscle in guinea-pig

Hydrogen sulfide (H 2S) is produced endogenously in mammalian tissues and is important in both physiological and pathological processes. Despite its importance, little is known regarding the effect of H 2S on gastrointestinal motility. We evaluated the effect of H 2S on the spontaneous contraction of gastric antrum smooth muscle in the guinea pig ( Cavia porcellus) using a physiograph. In addition, we investigated whether the effect of H 2S was mediated by ionic channels by recording membrane currents in freshly dispersed gastric antrum myocytes using a whole-cell patch clamp. Sodium hydrogen sulfide (NaHS), an H 2S donor, had a dual effect on the spontaneous contraction of gastric antrum muscle strips. At high concentrations (0.3–1.0 mM), NaHS suppressed the amplitude of spontaneous contraction. At low concentrations (0.1–0.3 mM), NaHS enhanced the resting tension of muscle strips while slightly reducing the contractile amplitude. The excitatory effect on spontaneous contraction, caused by low concentrations of NaHS, was abolished when the muscle strips were pretreated with 10 mM tetraethylammonium (TEA), a nonselective potassium channel blocker, or 0.5 mM 4-Aminopyridine (4-AP), a voltage-gated K + channel blocker. However, the excitatory effect of NaHS was not completely blocked by low concentrations of TEA (1 mM). Pretreatment with both TEA (1 mM) and 4-AP (0.5 mM) completely abolished the excitatory effect. The dose-response curve for the inhibitory effect of NaHS on the spontaneous contraction of gastric smooth muscle was shifted significantly to the left by TEA and 4-AP. Both Pinacidil, a K ATP channel opener, and NaHS significantly inhibited TEA-potentiated spontaneous contraction. Glibenclamide, a K ATP channel blocker, partially, but significantly, reversed the reduction in amplitude. NaHS enhanced the amplitude of the K ATP current, but inhibited the voltage-gated K + channel current (IK V) in a dose-dependent manner. NaHS had no effect on STOC at low concentrations (0.1–1.0 mM) but significantly inhibited STOC at high concentrations (4–10 mM). Our results suggest that H 2S has multiple actions during the regulation of gastric motility in the guinea-pig. An excitatory effect is mediated via inhibition of the voltage-gated K + channel and an inhibitory effect is mediated via activation of the K ATP channel.

Testosterone‐Independent Down‐Regulation of Oct2 in the Kidney Medulla from a Uranyl Nitrate‐Induced Rat Model of Acute Renal Failure: Effects on Distribution of a Model Organic Cation, Tetraethylammonium

Although acute renal failure (ARF) has been an area of extensive research in recent decades, our understanding of ARF is far from complete. Organic cations (OCs) are primarily excreted via vectorial transport by various renal organic cation transporters (OCTs). It is reasonable to assume that ARF may alter the expression profiles of these transporters. In a rat ARF model, induction of ARF by uranyl nitrate (UN) treatment significantly decreased the levels of Oct2 (slc22a2) mRNA and protein in the kidney medulla. mRNA expression of the other OCTs was not appreciably altered. The plasma level of testosterone, a well‐known regulator of Oct2, was not changed, suggesting that the Oct2 down‐regulation is testosterone‐independent. The effect of reduced Oct2 expression on the distribution of a model OC, tetraethylammonium (TEA), in various rat tissues including kidney cortex and kidney medulla was investigated during steady state plasma TEA concentrations. The steady state tissue‐to‐plasma (T/P) TEA ratio was decreased in the kidney medulla (approximately 15‐fold) during ARF. These results indicate that, in a rat model of ARF, reduced Oct2 expression in the kidney medulla results in decreased distribution of TEA to the kidney medulla, thereby reducing renal clearance of TEA in UN‐ARF rats. © 2008 Wiley‐Liss, Inc. and the American Pharmacists Association J Pharm Sci 98:739–747, 2009

Tetraethylammonium (TEA) increases the inactivation time constant of the transient K + current in suprachiasmatic nucleus neurons

Identifying the mechanisms that drive suprachiasmatic nucleus (SCN) neurons to fire action potentials with a higher frequency during the day than during the night is an important goal of circadian neurobiology. Selective chemical tools with defined mechanisms of action on specific ion channels are required for successful completion of these studies. The transient K + current ( I A) plays an active role in neuronal action potential firing and may contribute to modulating the circadian firing frequency. Tetraethylammonium (TEA) is frequently used to inhibit delayed rectifier K + currents ( I DR) during electrophysiological recordings of I A. Depolarizing voltage-clamped hamster SCN neurons from a hyperpolarized holding potential activated both I A and I DR. Holding the membrane potential at depolarized values inactivated I A and only the I DR was activated during a voltage step. The identity of I A was confirmed by applying 4-aminopyridine (5 mM), which significantly inhibited I A. Reducing the TEA concentration from 40 mM to 1 mM significantly decreased the I A inactivation time constant ( τ inact) from 9.8 ± 3.0 ms to 4.9 ± 1.2 ms. The changes in I A τ inact were unlikely to be due to a surface charge effect. The I A amplitude was not affected by TEA at any concentration or membrane potential. The isosmotic replacement of NaCl with choline chloride had no effect in I A kinetics demonstrating that the TEA effects were not due to a reduction of extracellular NaCl. We conclude that TEA modulates, in a concentration dependent manner, τ inact but not I A amplitude in hamster SCN neurons.

Brownian Dynamics Theory for Predicting Internal and External Blockages of Tetraethylammonium in the KcsA Potassium Channel

The theory of Brownian dynamics is used to model permeation and the blocking of KcsA potassium channels by tetraethylammonium (TEA). A novel Brownian dynamics simulation algorithm is implemented that comprises two free energy profiles; one profile is seen by the potassium ions and the other by the TEA molecules whose shape is approximated by a sphere. Our simulations reveal that internally applied TEA blocks the passage of K + ions by physically occluding the pore. A TEA molecule in the external reservoir encounters an attractive energy-well created by four tyrosine residues at position 82, in addition to all other attractive and repulsive forces impinging on it. Using Brownian dynamics, we investigate how deep the energy-well needs to be to reproduce the experimentally determined inhibitory constant k i for the TEA blockade of KcsA or the mutant Shaker T449Y. The one-dimensional free energy profile obtained from molecular dynamics is first converted into a one-dimensional potential energy profile, and is then transformed into a three-dimensional free energy profile in Brownian dynamics by adding the short-range potential from the channel walls. When converted, the free energy profile calculated from molecular dynamics gives a well-depth of ∼10 kT. We systematically alter the depths of the profiles, and then use Brownian dynamics simulations to numerically determine the current versus TEA-concentration curves. We show that the sequence of binding and unbinding events of the TEA molecule to the binding pocket can be modeled by a first-order Markov process. The Brownian dynamics simulations also reveal that the probability of a TEA molecule binding to the binding pocket in KcsA potassium channels increases exponentially with TEA concentration and depends also on the applied potential and the K + concentration in the simulation assembly.

Effects of tetraalkylammonium compounds with different affinities for organic cation transporters on the pharmacokinetics of metformin

The study sought to investigate the effects of tetraalkylammonium (TAA), inhibitors of the organic cation transporters (OCTs) with different affinities, on the pharmacokinetics of metformin. The inhibitory potentials of TAAs on the uptake of metformin were evaluated by determining IC(50) values in MDCK cells over-expressing OCTs and, to assess in vivo drug interactions, metformin and TAAs were coadministered to rats. Uptake of metformin was facilitated by over-expression of hOCT1 and hOCT2 and showed saturable processes, indicating that metformin is a substrate of hOCT1 and hOCT2. The IC(50) values of TAAs for hOCT2 were lower than hOCT1 and decreased with increasing alkyl chain length, indicating that the inhibitory potential of TAAs on metformin uptake was greater in hOCT2 than in hOCT1 and increased with increasing alkyl chain length. The plasma concentration of metformin was elevated by the coadministration of tetrapropylammonium (TPrA) and tetrapentylammonium (TPeA), but not by tetramethylammonium (TMA) or tetraethylammonium (TEA). However, the plasma concentrations of TMA, TEA and TPrA were not changed by the coadministration of metformin. In conclusion, in vivo drug interactions between metformin and TAAs were caused only when metformin was coadministered with TAAs showing higher affinities for OCTs.

Tetraethylammonium and nicotine transport by the Malpighian tubules of insects

We examined transepithelial transport of the prototypical type I organic cation (OC) tetraethylammonium (TEA) and the plant alkaloid nicotine by the isolated Malpighian tubules (MTs) of nine insect species from six orders. Isolated tubules were exposed to radiolabelled forms of either TEA or nicotine in the bathing (basal) fluid. Luminal (apical) secreted fluid was collected and TEA or nicotine concentration was determined. Active net transport of nicotine from bath to lumen was observed by the MTs of all the insects studied. TEA was also transported from bath to lumen in MTs of all species except Rhodnius prolixus and Aedes aegypti. MTs of both of these blood feeders did not show active transport of TEA under normal physiological conditions. Transport of TEA but not nicotine increased during the moult in the MTs of Rhodnius, but the concentrations of TEA in the secreted fluid were still consistent with passive accumulation in response to the lumen-negative transepithelial potential. Nicotine transport by Rhodnius MTs was inhibited by the type II OC quinidine, a known p-glycoprotein inhibitor, but not by the type I OCs N-methylnicotinamide or cimetidine. Taken together, the results suggest that active transport of OCs by the MTs is common among species from different orders and that transepithelial TEA and nicotine transport occur through separate pathways.

Fictive locomotor patterns generated by tetraethylammonium application to the neonatal rat spinal cord in vitro

Intrinsic spinal networks generate a locomotor rhythm characterized by alternating electrical discharges from flexor and extensor motor pools. Because this process is preserved in the rat isolated spinal cord, this preparation in vitro may be a useful model to explore methods to reactivate locomotor networks damaged by spinal injury. The present electrophysiological investigation examined whether the broad spectrum potassium channel blocker tetraethylammonium could generate locomotor-like patterns. Low (50–500μM) concentrations of tetraethylammonium induced irregular, synchronous discharges incompatible with locomotion. Higher concentrations (1–10mM) evoked alternating discharges between flexor and extensor motor pools, plus large depolarization of motoneurons with spike broadening. The alternating discharges were superimposed on slow, shallow waves of synchronous depolarization. Rhythmic alternating patterns were suppressed by blockers of glutamate, GABA A and glycine receptors, disclosing a background of depolarizing bursts inhibited by antagonism of group I metabotropic glutamate receptors. Furthermore, tetraethylammonium also evoked irregular discharges on dorsal roots. Rhythmic alternating patterns elicited by tetraethylammonium on ventral roots were relatively stereotypic, had limited synergy with fictive locomotion induced by dorsal root stimuli, and were not accelerated by 4-aminopyridine. Horizontal section of the spinal cord preserved irregular ventral root discharges and dorsal root discharges, demonstrating that the action of tetraethylammonium on spinal networks was fundamentally different from that of 4-aminopyridine. These results show that a potassium channel blocker such as tetraethylammonium could activate fictive locomotion in the rat isolated spinal cord, although the pattern quality lacked certain features like frequency modulation and strong synergy with other inputs to locomotor networks.