Voltage-sensitive Ion Channels Research Articles

Antiarrhythmic Effects of Polyunsaturated Fatty Acids

The epidemiological studies of Kromann and Green1 on the low mortality rate of Greenland Inuits from ischemic heart disease led to the suggestion by Bang et al2 that despite the high total fat intake of the Eskimos, this low mortality rate was due to the abundance of n-3 fatty acids from seafood in their diet. This hypothesis initiated research by many investigators into possible antiatherogenic effects of n-3 PUFAs. Much has been learned regarding physiological and biochemical changes induced by this class of essential fatty acids that could have potential antiatherogenic effects; nevertheless, controversy persists in the current literature regarding the clinical evidence for beneficial effects from fish ingestion (the major dietary source of n-3 fatty acids) on the development of coronary heart disease.3 4 Studies seeking a resolution to the potential antiatherogenic effects of n-3 PUFA will undoubtedly continue.5 Meanwhile, on the basis of earlier sporadic suggestions that n-3 PUFA might possess antiarrhythmic effects,6 7 McLennan and coworkers8 9 pursued this possibility. They showed in feeding studies in rats that when saturated fats or olive oil is the major dietary fat, a high incidence of fatal, irreversible VF occurs from experimental coronary artery ligation, which was significantly reduced when the dietary fat was vegetable oil but was essentially abolished by fish oil. They have confirmed their basic finding in marmosets.10 It is not our purpose in this brief review to discuss possible antiatherogenic effects of n-3 PUFA or the role of coronary heart disease in causing malignant ventricular arrhythmias. Clearly, coronary heart disease is the major clinical setting in which malignant ventricular arrhythmias occur today, and myocardial ischemia is the most common trigger eliciting such arrhythmias. Rather, we will focus on our recent studies on the mechanism of the apparent antiarrhythmic actions …

Okadaic acid-like toxin in systemic lupus erythematosus patients: Hypothesis for toxin-induced pathology, immune dysregulation, and transactivation of herpesviruses

Preliminary evidence suggests there is a toxin in the sera of systemic lupus erythematosus patients which reacts with a commercial enzyme-linked immunosorbent assay kit for the detection of the marine toxin, okadaic acid. Data is presented which supports the hypothesis that an okadaic acid-like toxin may be the principle agent of lymphocyte dysregulation in systemic lupus erythematosus and other immune-dysreguaated states. The okadaic acid-like toxin can produce the specific abnormalities in T-lymphocyte phenotype and function typical of systemic lupus erythematosus, principally through its ability to inhibit serine/threonine phosphatases necessary for secondary signalling processes, and through its ability to inhibit calcium which is crucial to protein kinase C-mediated signalling of T-lymphocytes. The disruption probably occurs through the protein tyrosine kinase p56 lck pathway crucial for IL-2. Additionally, the toxin's ability to disrupt voltage-sensitive ion channels in cell membranes may be responsible for the multi-organ pathology observed in systemic lupus erythematosus patients, particularly neurological, cardiac and nephritic. Data from a different study conducted by the author suggests that latent and persistent viruses are reactivated in active lupus. This activation could be the result of the toxin's ability to act as an immune modulator, or its ability to act as a transactivating factor.

Nongenomic effects of progesterone on human intestinal smooth muscle cells.

Previous experiments demonstrated that progesterone affects intestinal smooth muscle cells through genomic and nongenomic pathways. We hypothesized that the nongenomic effect was mediated by changes in membrane excitability. We studied the effects of progesterone and other steroid hormones on a human intestinal smooth muscle cell line, using the whole cell patch-clamp technique. Ionic currents were elicited through steps from -70 mV to various test potentials. Progesterone dose-dependently reduced calcium currents. The decrease in inward current was partly due to a shift in the steady-state inactivation to more hyperpolarized potentials. This effect did not involve gene transcription, since it was not blocked by the progesterone antagonist ZK-98-299. The progesterone analogue 5-beta-dihydroprogesterone also decreased calcium currents, whereas its stereoisomer, 5-alpha- dihydroprogesterone, did not affect the properties of voltage-sensitive ion channels. Similarly, estradiol and dexamethasone did not alter inward currents. We conclude that progestins exert their nongenomic effects on intestinal smooth muscle cells by decreasing calcium currents. The change in the calcium signal may contribute to the reduction in muscle contraction observed after progesterone.

A Novel Chloride Channel Localizes toCaenorhabditis elegansSpermatids and Chloride Channel Blockers Induce Spermatid Differentiation

Caenorhabditis elegansspermatogenesis is especially suited for studies of nonrandom cytoplasmic segregation during cellular differentiation. Spermatocytes separate from an anuclear cytoplasmic core and undergo two sequential divisions. During the second division, intracellular organelles segregate specifically to spermatids as they bud from an anuclear residual body. We have applied patch-clamp techniques in order to investigate membrane protein distribution during these asymmetric divisions. We show that membrane components, as assayed by voltage-dependent ion channel activity, follow a specific distribution pattern during sperm development. Several voltage-sensitive ion channel activities are observed in spermatocytes and residual bodies, but only a single-channel type can be detected in spermatids, indicating that other channel activities are excluded from or inactivated within these cells as they form. The channel that is observed in spermatids is an inward-rectifying chloride channel (Clir), as indicated by its sensitivity to chloride channel inhibitors and Cl-dependent shifts in its conductance. Treatment of spermatids with Cl channel blockers induce their differentiation into spermatozoa, suggesting that Clir plays a role during this developmental step. These studies are the first application of patch-clamp electrophysiology toC. elegansdevelopment.

Control of voltage-dependent biomolecules via nonequilibrium kinetic focusing.

We show that voltage-sensitive macromolecules can, by the application of the right type of external voltage fluctuations, be focused with great probability into a desired kinetic substate. As an illustration we consider an eight-state model of the Shaker K channel driven by dichotomous voltage fluctuations. This technique may provide a powerful new tool for the study of the kinetics of voltage sensitive ion channels.

Properties of voltage-gated ion channels formed by syringomycin E in planar lipid bilayers.

Using the planar lipid bilayer technique we demonstrate that the lipodepsipeptide antibiotic, syringomycin E, forms voltage-sensitive ion channels of weak anion selectivity. The formation of channels in bilayers made from dioleoylglycerophosphatidylserine doped with syringomycin E at one side (1-40 micrograms/ml) was greatly affected by cis-positive voltage. A change of voltage from a positive to a negative value resulted in (i) an abrupt increase in the single channel conductance (the rate of increase was voltage dependent) simultaneous with (ii) a closing of these channels and an exponential decrease in macroscopic conductance over time. The strong voltage dependence of multichannel steady state conductance, the single channel conductance, the rate of opening of channels at positive voltages and closing them at negative voltages, as well as the observed abrupt increase of single channel conductance after voltage sign reversal suggest that the change of the transmembrane field induces a significant rearrangement of syringomycin E channels, including a change in the spacing of charged groups that function as voltage sensors. The conductance induced by syringomycin E increased with the sixth power of syringomycin E concentration suggesting that at least six monomers are required for channel formation.

Voltage-sensitive Adenylyl Cyclase Activity in Cultured Neurons. A CALCIUM-INDEPENDENT PHENOMENON

Catalytic subunits of mammalian adenylyl cyclases have been proposed to contain 12 transmembrane domains, a property shared with some voltage-sensitive ion channels. Here we report that adenylyl cyclase activity in cerebellar neurons is synergistically stimulated by depolarizing agents and beta-adrenergic receptor activation. This phenomenon is Ca(2+)-independent and not attributable to Ca(2+)-stimulated adenylyl cyclase activity. Cholera toxin and forskolin also synergistically stimulate adenylyl cyclase activity in combination with depolarizing agents. We hypothesize that conformational changes in the catalytic subunit of the enzymes caused by changes in the membrane potential may enhance stimulation of adenylyl cyclases by the guanylyl nucleotide stimulatory protein. This novel mechanism for regulation of adenylyl cyclases generates robust cAMP signals that may contribute to various neuromodulatory events including some forms of neuroplasticity.

Spatial integration of local transmitter responses in motoneurones of the turtle spinal cord in vitro.

1. Integration of responses to local activation of transmitter receptors in the dendrites of motoneurones was investigated in a slice preparation of the turtle spinal cord. Membrane-active substances were applied from up to three independent iontophoresis electrodes during intracellular recording from the cell body. 2. Responses to glutamate could be evoked from dendrites closer than 20 microns from the tip of the glutamate electrode. The effects of other substances were more widespread. 3. In normal medium the configuration of a glutamate response was affected by time-dependent anomalous rectification. In the presence of muscarine the sum of glutamate responses from two different dendrites recruited a voltage-sensitive plateau potential. 4. The response to glutamate from one dendrite could be attenuated by local application of gamma-aminobutyric acid (GABA) without effects on soma conductance or glutamate responses from other dendrites. 5. The response to glutamate from one dendrite could be selectively enhanced by local application of tetraethylammonium (TEA) or N-methyl-D-aspartate (NMDA) without effects on soma conductance or glutamate responses from other dendrites. 6. NMDA could convert a tonic glutamate response from one dendrite into a phasic response without affecting the configuration of glutamate responses from other dendrites. 7. The effects of TEA and NMDA were facilitated by depolarization and reduced by hyperpolarization. 8. We conclude that the cable structure of motoneurones and the distribution of synapses and voltage-sensitive ion channels provide relative autonomy to non-linear synaptic processing and modulation in confined dendritic regions.

Axolemmal and septal conduction in the impedance of the earthworm medial giant nerve fiber

Ionic conduction in the axolemmal and septal membranes of the medial giant fiber (MGF) of the earthworm (EW) Lumbricus terrestris was assessed by impedance spectroscopy in the frequency range 2.5-1000 Hz. Impedance loci in the complex plane were described by two semi-circular arcs, one at a lower characteristic frequency (100 Hz) and the other at a higher frequency (500 Hz). The lower frequency arc had a chord resistance of 53 k omega and was not affected by membrane potential changes or ion channel blockers [tetrodotoxin (TTX), 3,4-diaminopyridine (3,4-DAP), 4-aminopyridine (4-AP), and tetraethylammonium (TEA)]. The higher frequency arc had a chord resistance of 274 k omega at resting potential, was voltage-dependent, and was affected by the addition of TTX, 3,4-DAP, 4-AP, and TEA to the physiological EW salines. When all four blockers were added to the bathing solution, the impedance locus was described by two voltage-independent arcs. Considering the effects of these and other (i.e., Cd and Ni) ion channel blockers, we conclude that: 1) the higher frequency locus reflects conduction by voltage-sensitive ion channels in the axolemmal membrane, which contains at least four ion channels selective for sodium, calcium, and potassium (delayed rectifier and calcium-dependent), and 2) the lower frequency locus reflects voltage-insensitive channels in the septal membrane, which separates adjacent MGFs.

Endothelin-A receptor mediates cardiac inhibition by regulating calcium and potassium currents.

Voltage-sensitive ion channels play fundamental roles in the regulation of cardiac function by various neurotransmitters. Endothelins have strong positive inotropic and chronotropic effects, for which recent studies have implicated various intracellular mechanisms. However, very little is known about the underlying ion-channel regulation by the peptide. We report here that endothelin-1 consistently hyperpolarizes the membrane and shortens the duration of the action potential in mammalian atrial myocytes, leading to suppression of electrical excitability of the heart. Endothelin-1, but not endothelin-3, inhibited the L-type calcium current by decreasing cyclic AMP accumulation and activated the muscarinic potassium current by stimulating a pertussis toxin-sensitive GTP-binding protein. Consistent with these results, endothelin-1 strongly reduced the heart rate when it was increased by beta-adrenoceptor stimulation. These effects were blocked by an ETA (endothelin-1-selective) receptor-selective antagonist, BQ123 (refs 8-11). The ETA receptor-mediated regulation of cardiac ion channels gives new insight into our understanding of the physiological and pathophysiological roles of endothelins in the control of cardiac function.

Gating current noise produced by elementary transitions in Shaker potassium channels.

Gating currents provide a direct record of the spatial rearrangement of charges occurring within the protein of voltage-sensitive ion channels. If the elementary charges move as very brief discrete pulses of current, they will produce fluctuations in the macroscopic gating current. The variance of such fluctuations in gating currents was measured in Shaker potassium channels expressed in Xenopus oocytes with a sufficiently high recording bandwidth to estimate the magnitude and time distribution of the elementary transition charge movements. Channel activation occurred in two sequential stages. The first stage consisted of numerous, fast transitions, each moving small amounts of charge that contributed little to the fluctuation in gating current, whereas the second stage, which contributed the bulk of the fluctuation, was represented by a number of discrete, correlated transitions, one or more of which carried a charge of at least 2.4 elementary charges across the membrane field.

Inhibition of serine/threonine protein phosphatases promotes opening of voltage-activated L-type Ca2+ channels in insulin-secreting cells

The biological activity of many proteins, including voltage-sensitive ion channels, is controlled by their state of phosphorylation. Ca2+ influx through voltage-activated L-type Ca2+ channels serves as the major stimulatory signal in insulin-secreting cells. We have now investigated the extent to which Ca2+ handling in clonal insulin-secreting RiNm5F cells was affected by okadaic acid, an inhibitor of various serine/threonine protein phosphatases. Whole-cell patch-clamp experiments showed that okadaic acid generated an increase in membrane current, suggesting that it promotes Ca2+ influx through L-type voltage-gated Ca2+ channels probably by modifying their phosphorylation state. Okadaic acid was found to provoke a transient rise in the cytoplasmic free Ca2+ concentration ([Ca2+]i) but had no further effect on the K(+)-induced increase. The Ca2+ transient induced by okadaic acid was dependent on the presence of extracellular Ca2+ and was abolished by D600, a blocker of voltage-activated L-type Ca2+ channels. Concomitant with the rise in [Ca2+]i, okadaic acid induced insulin secretion, a phenomenon that was also dependent on extracellular Ca2+. It is proposed that hyperphosphorylation of voltage-activated L-type Ca2+ channels in insulin-secreting cells lowers the threshold potential for their activation.

Hippocampal noradrenaline release is modulated by γ- and δ-hexachlorocyclohexane isomers: which mechanisms are involved?

The differential effects of γ- and δ-hexachlorocyclohexane isomers on 25 mM K +-evoked release of [ 3H]noradrenaline were studied in hippocampal slices treated with selected agents to activate or block L- and N-type Ca 2+ and Na + voltage-sensitive ion channels, Cl − transport and Ca 2+-dependent protein activity. At maximally effective concentrations, the L- and N-type Ca 2+ channel blockers nifedipine and ω-conotoxin, respectively, and the Na + channel antagonist tetrodotoxin did not modify the enhancement of K +-evoked [ 3H]noradrenaline release induced by γ-hexachlorocyclohexane. Likewise, under activation of protein kinase C by phorbol 12,13-dibutyrate (PDB) or inhibition of calmodulin by N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide (W-7), the stimulatory effect of γ-hexachlorocyclohexane remained almost unchanged. The Cl − transport blocker 4,4-diisothiocyanato-stilbene-2,2′-disulfonic acid (DIDS) significantly reduced the effect of γ-hexachlorocyclohexane on [ 3H]noradrenaline release. The enhanced release in the presence of Bay K 8644, the L-type Ca 2+ channel activator, was significantly inhibited by nifedipine but not by δ-hexachlorocyclohexane. The combination of ω-conotoxin and tetrodotoxin with δ-hexachlorocyclohexane did not alter the [ 3H]noradrenaline release effects of each agent alone. Activation of protein kinase C in the presence of δ-hexachlorocyclohexane resulted in a reduction of the δ isomer effect and in a potentiation of the PDB effect. W-7 did not further facilitate the inhibition induced by δ-hexachlorocyclohexane alone. These data suggest that hexachlorocyclohexane isomers may modify K +-evoked [ 3H]noradrenaline release by interacting with presynaptic molecular processes involving changes in Cl − membrane permeability and intracellular Ca 2+ homeostasis.

Convergence of auditory nerve fibers onto bushy cells in the ventral cochlear nucleus: implications of a computational model.

1. Convergence of auditory nerve (AN) fibers onto bushy cells of the ventral cochlear nucleus (VCN) was investigated with a model that describes the electrical membrane properties of these cells. The model consists of a single compartment, representing the soma, and includes three voltage-sensitive ion channels (fast sodium, delayed-rectifier-like potassium, and low-threshold potassium). These three channels have characteristics derived from voltage clamp data of VCN bushy cells. The model also contains a leakage channel, membrane capacitance, and synaptic inputs. The model accurately reproduces the membrane rectification seen in current clamp studies of bushy cells, as well as their unique current clamp responses. 2. In this study, the number and synaptic strength of excitatory AN inputs to the model were varied to investigate the relationship between input convergence parameters and response characteristics. From 1 to 20 excitatory synaptic inputs were applied through channels in parallel with the voltage-gated channels. Each synapse was driven by an independent AN spike train; spike arrivals produced brief (approximately 0.5 ms) conductance increases. The number (NS) and conductance (AE) of these inputs were systematically varied. The input spike trains were generated as a renewal point process that accurately models characteristics of AN fibers (refractoriness, adaptation, onset latency, irregularity of discharge, and phase locking). Adaptation characteristics of both high and low spontaneous rate (SR) AN fibers were simulated. 3. As NS and AE vary over the ranges 1-20 and 3-80 nS, respectively, the full range of response types seen in VCN bushy cells are produced by the model, with AN inputs typical of high-SR AN fibers. These include primarylike (PL), primarylike-with-notch (Pri-N), and onset-L (On-L). In addition, Onset responses, whose association with bushy cells in uncertain, and "dip" responses, which are not seen in the VCN, are produced. Dip responses occur with large NS and/or AE, and are due to depolarization block. When the AN inputs have the adaptation characteristics of low-SR AN fibers, PL--but not Pri-N or On-L responses--are produced. This suggests that neurons showing Pri-N and On-L responses must receive high-SR AN inputs. 4. The regularity of discharge of the model is compared with that of VCN bushy cells, using a measure derived from the mean and standard deviation of interspike intervals. Regularity is an important constraint on the model because the regularity of VCN bushy cells is the same as that of their AN inputs.(ABSTRACT TRUNCATED AT 400 WORDS)

Voltage-Activated Ionic Currents in Goldfish Pituitary Cells

The release of gonadotropin and growth hormone from goldfish pituitary cells has been shown to be dependent on the entry of extracellular Ca2+ through voltage-sensitive Ca2+ channels by pharmacological studies. As a first step to further investigate the involvement of voltage-dependent ion channels in the regulation of anterior pituitary hormone release in the goldfish, cell excitability and voltage-dependent ion currents were characterized using tight-seal whole-cell recordings in dispersed goldfish pituitary cells. Cultured goldfish pituitary cells had an average membrane potential of -36 ± 3 mV. When held at membrane potentials more negative than -60 mV, these cells were excitable, responding to depolarizing current pulses or anode break with the firing of single action potentials. Results from total current voltage-clamp recordings suggested that all goldfish pituitary cells possess voltage-dependent Na+ , Ca2+, and K+ currents. These currents were further characterized independently under isolated current recording conditions. The rapid, transient Na+ current activated at voltages more positive than -40 mV and was sensitive to tetrodotoxin. The steady state inactivation of this Na+ current was also voltage-dependent; at the measured resting potential, ⩾50% of the Na+ current was not available for activation. The voltage-dependence and activation kinetics of the tetraethylammonium-sensitive K+ current resembled those of the delayed rectifier K+ current. The K+ current activated slowly at potentials more positive than -40 mV, and showed little inactivation over the duration of a 1-sec depolarizing pulse. Steady-state inactivation characteristics indicated that ≤50% of the K+ current was inactivated at resting potentials. Experiments with 4-aminopyridine indicated the presence of an early transient K+ current that activated in a similar voltage range as the delayed rectifier current. Using barium as the charge carrier to measure Ca2+ currents, a high-voltage activated, long-lasting Ca2+ current was revealed. This "L-type" Ca2+ current activated at potentials more positive than -30 mV and was inhibited by verapamil and nifedipine. This study indicates that goldfish pituitary cells possess the electrophysiological properties required for the participation of voltage-sensitive ion channels in the regulation of hormone release.

Rapid communication between neurons and astrocytes in primary cortical cultures

The identification of neurotransmitter receptors and voltage-sensitive ion channels on astrocytes (reviewed by Barres, 1991) has renewed interest in how these cells respond to neuronal activity. To investigate the physiology of neuron astrocyte signaling, we have employed primary cortical cultures that contain both neuronal and glial cells. As the neurons in these cultures exhibit synchronous spontaneous synaptic activity, we have used both calcium imaging and whole-cell recording techniques to identify physiological activity in astrocytes related to neuronal activity. Whole-cell voltage-clamp records from astrocytes revealed rapid inward currents that coincide with bursts of electrical activity in neighboring neurons. Calcium imaging studies demonstrate that these currents in astrocytes are not always associated with slowly propagating calcium waves. Inclusion of the dye Lucifer yellow within patch pipettes confirmed that astrocytes are extensively coupled to each other but not to adjacent neurons, indicating that the currents observed are not due to gap junction connections between these cell types. These currents do not reflect widespread diffusion of glutamate or potassium released during neuronal activity since a population of small, round, multipolar presumed glial cells that are not dye coupled to adjacent cells did not display electrical currents coincident with neuronal firing, even though they respond to locally applied glutamate and potassium. These findings indicate that, in addition to the relatively slow signaling conveyed by calcium waves, astrocytes also display rapid electrical responses to neuronal activity.

Molecular evolution of voltage-sensitive ion channel genes: on the origins of electrical excitability.

We have analyzed nucleic acid and amino acid sequence alignments of a variety of voltage-sensitive ion channels, using several methods for phylogenetic tree reconstruction. Ancient duplications within this family gave rise to three distantly related groups, one consisting of the Na+ and Ca++ channels, another the K+ channels, and a third including the cyclic nucleotide-binding channels. A series of gene duplications produced at least seven mammalian homologues of the Drosophila Shaker K+ channel; clones of only three of these genes are available from all three mammalian species examined (mouse, rat, and human), pointing to specific genes that have yet to be recovered in one or another of these species. The Shaw-related K+ channels and the Na+ channel family have also undergone considerable expansion in mammals, relative to flies. These expansions presumably reflect the needs of the high degree of physiological and neuronal complexity of mammals. Analysis of the separate domains of the four-domain channels (Ca++ and Na+) supports their having evolved by two sequential gene duplications and implies the historical existence of a functional two-domain channel.

Specificity of Lambert-Eaton Myasthenic Syndrome immunoglobulin for nerve terminal calcium channels

Lambert-Eaton Myasthenic Syndrome (LEMS) is a presynaptic, neuromuscular disorder characterized by impaired nerve-evoked release of ACh. Repetitive nerve stimulation, which increases the probability of quantal release, improves the transmission defect. An autoantibody to Ca 2+ channels of presynaptic motor nerve terminals is thought to mediate the pathogenesis of this disease. The goal of the present study was to examine the specificity of LEMS autoantibodies for nerve terminal Ca 2+ channels as compared to other voltage-sensitive ion channels in nerve terminals, and to determine if non-specific membrane damage contributed to the pathogenesis of LEMS. The ion channel specificity of LEMS autoantibody was assessed by comparing the ability of acute application of IgG isolated from the plasma of a patient with LEMS to reduce depolarization-dependent uptake of 45Ca 2+ and 22Na + into or efflux of 86Rb + from rat forebrain synaptosomes. The clinical diagnosis of LEMS was confirmed electrophysiologically by treatment of mice for 30 days with plasma (1.5 ml/day) taken from this patient. Characteristic reduction of quantal content elicited at 1 Hz and facilitation at 20 Hz was observed in mice treated wtih LEMS plasma compared to those treated with control plasma. One s, K +-stimulated 45Ca 2+ uptake was inhibited36.5 ± 14.5%and44.5 ± 9.8% by acute application of 2 and 4 mg/ml LEMS IgG, respectively; IgG from patients with small cell carcinoma of the lung (SCC) had no effect on 45Ca 2+ entry. The same concentrations of LEMS IgG affected neither voltage-dependent uptake of 22Na + into veratridine-depolarized synaptosomes nor 86R + efflux from K +-depolarized synaptosomes. Lactate dehydrogenase release from synaptosomes incubated with varying concentrations of LEMS IgG was not increased over control suggesting that the reduction in 45Ca 2+ uptake was not a result of disruption of synaptosomal membrane integrity. Likewise, normal activation of Ca 2+ channels was not prevented by IgG-induced alterations in synaptosomal membrane potentials as the fluorescence of a membrane potential-sensitive carbocyanine dye (diS-C 2(5)) was unaltered following acute application of 2 mg/ml LEMS or control IgG. These results indicate that immunoglobulins isolated from a patient with LEMS alter presynaptic nerve terminals Ca 2+ channel activity. These deficits are not due to decreases in synaptosomal integrity nor to IgG-induced alterations in the synaptosomal membrane potential but appear to result from direct action on voltage-dependent Ca 2+ channels. The antibody from this patient appears to be specific for nerve terminal Ca 2+ channels as it did not affect ion flux through voltage-dependent Na + or K + channels.

Gating currents from a delayed rectifier K+ channel with altered pore structure and function

For voltage-sensitive ion channels, charged elementswithin the membrane electric field are thought togeneratedisplacementorgatingcurrents(1). Topograph­ical models (2-4)have assigned thecharged elements orvoltage sensor to a fourth transmembrane a-helix(S4)and the models have received supportfrom mutagenesisexperiments (5-6). The pore has been assigned to adifferent region, and in K+ channels, this appears to bethe highly-conserved S5-S6 linker (7-9). To test thehypothesis that the pore and the voltage sensor arestructurally distinct, we compared ion conduction andgating currents arising from two delayed rectifier K+channels, DRKI (10) and a chimera DRK/NGKL374V(11, 12), whose sequences even though identical in S4,the putative voltage sensor differed at eight of the 21residues in the S5-S6linker(6) as shown in the followingalignment.parent in single-channel recordingsfrom cell-attachedpatches of oocytes expressing currents from the twospecies of cRNA. The single channel conductance forK+ was -8and 4 pS for DRKI and DRK/NGKL374V,respectively (Fig. 1, B, D). Single-channelkinetics alsoshowed large differences: the long openings typical ofDRKI were converted into short openings in DRKINGK L374V (Fig. 1C). Furthermore, in DRKI cell­attached patcheswith Iso-Rb+in the pipette, the inwardRb+ conductance and theoutward K+ conductancewereidentical (gRb+IgK+ was -1;Fig. 1D, left), whereasDRK/NGKL374V showed a strong inward rectification(Fig. 1, D, right), and the gRb+IgK+ ratio rose to 2.4. Inaddition, the profile for blockade by external and inter­nal tetraethylammonium ions differed markedly be­tween the two channels (7,11,13).DRKI 357:

Tetraethylammonium, a K + channel blocker, inhibits interferon-γ-induced major histocompatibility class II antigen (Ia) expression and DNA synthesis in rat astrocytes

Astrocytes play an important role in antigen presentation to T lymphocytes by their ability to express major histocompatibility class II (Ia) antigen upon exposure to a number of agents, including interferon-γ (IFN-γ). Astrocytes have been shown to express a variety of voltage-sensitive ion channels including voltage-sensitive K + channels. The function(s) of these channels in astrocyte functions is not clearly understood. In this investigation, we examined: (1) the comparative effects of mouse, rat, and human recombinant IFN-γ (rIFN-γ) on the induction of Ia antigen and DNA synthesis in rat astrocytes; (2) the effect of tetraethylammonium (TEA), a K + channel blocker, on rat IFN-γ-induced Ia expression and DNA synthesis in rat astrocytes. Our data show that all INF-γ induce DNA synthesis in rat astrocytes but only rat and mouse and not the human IFN-γ will induce Ia expression. TEA in a dose-dependent manner inhibited both Ia expression and DNA synthesis in rat astrocytes. The concentration kinetics of TEA with regard to maximum inhibition of Ia and DNA synthesis were different. Furthermore, these inhibitory effects were not a result of toxic or nonspecific effect of TEA on astrocytes as demonstrated by viability data and lack of any effect of tetramethylammonium, an analogue of TEA, that does not block K + ion channels. These data suggest a role of K + channels in Ia expression and DNA synthesis, therefore in immunological functions of astrocytes.