Voltage-sensitive Ion Channels Research Articles

Nano to Micro — Fluorescence Measurements of Electric Fields in Molecules and Genetically Specified Neurons

Our central nervous system is based on the generation and propagation of electrical signals along the neuronal pathways. These variations of the membrane potential are arranged by the concerted action of ion channels in the neuronal membrane. Therefore, the exact measurement of the electric field in the central nervous system is the focus of intensive investigation. While electrophysiological methods provide exact measurements on the single-cell or single-molecule level with high temporal resolution, they are limited in their spatial resolution ranging from a few single cells to a single molecule. To thoroughly understand how the voltage-dependent ion channels sense the membrane potential and are precisely gated by it, the electric field within the protein has to be investigated. Likewise, the propagation of electrical impulses in a network of neurons involves a large number of cells, which have to be monitored simultaneously. For these endeavors, optical methods have proven to be useful due to their scalability, temporal and spatial resolution. Here, we will summarize the properties of the optical probes that we used to determine the electrical field strength within voltage-sensitive ion channels and discuss the hybrid approach to detect membrane potential changes in genetically specified neurons in terms of design, limitations and future developments.

Interface connections of a transmembrane voltage sensor

Voltage-sensitive ion channels open and close in response to changes in transmembrane (TM) potential caused by the motion of the S4 voltage sensors. These sensors are alpha-helices that include four or more positively charged amino acids, most commonly arginine. The so-called paddle model, based on the high-resolution structure of the KvAP K+ channel [Jiang, et al. (2003) Nature 423, 33-41], posits that the S4 sensors move within the membrane bilayer in response to TM voltage changes. Direct exposure of S4 sensors to lipid is contrary to the classical expectation that the dielectric contrast between the membrane hydrocarbon core and water presents an insurmountable energetic penalty to burial of electric charges. Nevertheless, recent experiments have shown that a helix with the sequence of KvAP S4 can be inserted across the endoplasmic reticulum membrane. To reconcile this result with the classical energetics argument, we have carried out a molecular dynamics simulation of an isolated TM S4 helix in a lipid bilayer. The simulation reveals a stabilizing hydrogen-bonded network of water and lipid phosphates around the arginines that reduces the effective thickness of the bilayer hydrocarbon core to approximately 10 A in the vicinity of the helix. It suggests that bilayer phospholipids can adapt locally to strongly perturbing protein elements, causing the phospholipids to become a structural extension of the protein.

Dendritic Excitability of Mouse Frontal Cortex Pyramidal Neurons Is Shaped by the Interaction among HCN, Kir2, and KleakChannels

Dendritically placed, voltage-sensitive ion channels are key regulators of neuronal synaptic integration. In several cell types, hyperpolarization/cyclic nucleotide gated (HCN) cation channels figure prominently in dendritic mechanisms controlling the temporal summation of excitatory synaptic events. In prefrontal cortex, the sustained activity of pyramidal neurons in working memory tasks is thought to depend on the temporal summation of dendritic excitatory inputs. Yet we know little about how this is accomplished in these neurons and whether HCN channels play a role. To gain a better understanding of this process, layer V-VI pyramidal neurons in slices of mouse prelimbic and infralimbic cortex were studied. Somatic voltage-clamp experiments revealed the presence of rapidly activating and deactivating cationic currents attributable to HCN1/HCN2 channels. These channels were open at the resting membrane potential and had an apparent half-activation voltage near -90 mV. In the same voltage range, K+ currents attributable to Kir2.2/2.3 and K+-selective leak (Kleak) channels were prominent. Computer simulations grounded in the biophysical measurements suggested a dynamic interaction among Kir2, Kleak, and HCN channel currents in shaping membrane potential and the temporal integration of synaptic potentials. This inference was corroborated by experiment. Blockade of Kir2/Kleak channels caused neurons to depolarize, leading to the deactivation of HCN channels, the initiation of regular spiking (4-5 Hz), and enhanced temporal summation of EPSPs. These studies show that HCN channels are key regulators of synaptic integration in prefrontal pyramidal neurons but that their functional contribution is dependent on a partnership with Kir2 and Kleak channels.

Molecular Pharmacology of Topiramate: Managing Seizures and Preventing Migraine

Topiramate is a neuromodulatory compound with stabilizing properties that was initially introduced for the management of partial seizures. Topiramate has been demonstrated to modify several receptor-gated and voltage-sensitive ion channels, including voltage-activated Na+ and Ca2+ channels and non-NMDA receptors. These receptors have been implicated in the pathophysiology of both epilepsy and migraine. The pharmacological mechanisms of action for topiramate that may explain its antiepileptic and migraine preventive activities will be discussed in this review. In addition, the potential relationship between the molecular activities of topiramate and its efficacy in epilepsy and migraine prevention will be emphasized.

Active dendritic properties constrain input-output relationships in neurons of the central olfactory pathway in the crayfish forebrain.

Parasol cells are multimodal sensory interneurons of the hemi-ellipsoid body in the decapod forebrain. In reptant crustaceans, the hemi-ellipsoid body resides in the base of the eyecup, as an appendage to the terminal medulla. Parasol cells exhibit periodic depolarizations at a frequency of 0.5-1.0 Hz. I have investigated the role of these periodic depolarizations and their superimposed impulse bursts in affecting the input/output properties of these neurons. Parasol cells receive input from photic, olfactory, and mechanosensory pathways. Strong stimulation over any one of these pathways can lead to the generation of one or more impulse bursts in a subset of parasol cells, timed to occur at the peak of successive periodic depolarizations. A role for the periodic depolarizations in the function of the parasol cells has yet to be established. I suggest the possibility that they may act as a nonlinear amplifier that boosts spatially-summated excitatory synaptic potentials from strong or appropriate stimuli above threshold for burst generation. Another possibility includes modification of voltage-sensitive ion channels in the dendritic membrane, permitting a more effective spread of excitatory synaptic currents to impulse or burst initiating zones. Impulse bursts may be a highly effective mode of output for these neurons, especially so as they occur synchronously in a subset of cells in response to strong sensory input. Furthermore, backfiring of bursts into the dendritic tree has a brief (2-3 second) but effective suppressive action upon weak sensory input, which can thereby be masked by stronger, burst-generating input. This masking phenomenon is seen in other arthropod sensory interneurons, where its physiological basis appears to be a transient accumulation of intracellular Ca(++) ions that open calcium-sensitive potassium channels.

The physical nature of life

Life evolved from the primeval world of physics. Sensory systems inform animals of the natural environment, enabling them to conduct responsively. The discovery of weak, DC bioelectric fields in the vicinity of aquatic organisms and the role they play in guiding sharks and rays to their prey have led to the recognition of fundamental, hitherto less well known, physical aspects of sensory biology. The inferred cybernetic algorithm of electric-field orientation in sharks and rays is highly effective and extremely robust. In orienting to the weak DC electric fields of ocean currents and to the earth's magnetic field, sharks and rays unwittingly practise the motional-electric principles that Einstein had in mind when he introduced the special theory of relativity. At the sense-organ, receptor-membrane, and ion-channel levels, the elasmobranch ampullae of Lorenzini operate on the basis of graded positive feedback driven by negative conductance, supposedly employing voltage-sensitive ion channels as the active, excitable elements. The electric sense of sharks and rays presents an exquisite implementation of the very biophysical principles that also govern the graded, much richer than all-or-none, integrative brain processes of animal and man.

New expression profiles of voltage-gated ion channels in arteries exposed to high blood pressure.

The diameters of small arteries and arterioles are tightly regulated by the dynamic interaction between Ca(2+) and K(+) channels in the vascular smooth muscle cells. Calcium influx through voltage-gated Ca(2+) channels induces vasoconstriction, whereas the opening of K(+) channels mediates hyperpolarization, inactivation of voltage-gated Ca(2+) channels, and vasodilation. Three types of voltage-sensitive ion channels have been highly implicated in the regulation of resting vascular tone. These include the L-type Ca(2+) (Ca(L)) channels, voltage-gated K(+) (K(V)) channels, and high-conductance voltage- and Ca(2+)-sensitive K(+) (BK(Ca)) channels. Recently, abnormal expression profiles of these ion channels have been identified as part of the pathogenesis of arterial hypertension and other vasospastic diseases. An increasing number of studies suggest that high blood pressure may trigger cellular signaling cascades that dynamically alter the expression profile of arterial ion channels to further modify vascular tone. This article will briefly review the properties of Ca(L), K(V), and BK(Ca) channels, present evidence that their expression profile is altered during systemic hypertension, and suggest potential mechanisms by which the signal of elevated blood pressure may result in altered ion channel expression. A final section will discuss emerging concepts and opportunities for the development of new vasoactive drugs, which may rely on targeting disease-specific changes in ion channel expression as a mechanism to lower vascular tone during hypertensive diseases.

Do preclinical seizure models preselect certain adverse effects of antiepileptic drugs

Classical screening tests (maximal electroshock, MES, and threshold pentylenetetrazol, PTZ) employ non-epileptic rodents and identify antiepileptic drugs (AEDs) with mechanisms of action associated with significant CNS side effects. Thus MES identifies drugs acting on Na + channels that produce cerebellar toxicity. It may be possible to produce novel AEDs more selectively targeted at voltage-sensitive (VS) ion channels. There is little specific evidence for the likely success of this strategy with subunit selective agents targeted at the different VS Na + channels. Drugs targeted at specific VS Ca ++ channels (T, N, P/Q types) may be useful in generalised seizures. There are many as yet unexplored possibilities relating to K+ channels. GABA related drugs acting on PTZ clonic seizures tend to induce sedation and muscle hypotonia. Studies in mice, particularly with knock-in mutations, but also with subunit selective agents acting via the GABA A benzodiazepine site, suggest that it is possible to produce agents which do or do not induce particular side effects (sedative, hypnotic, anxiolytic, muscle relaxant, amnesia, anaesthesia). Whether these findings transfer to man has yet to be established. Acquired epilepsy in rodents (e.g. kindling or spontaneous seizures following chemically- or electrically-induced status epilepticus) or acquired epilepsy in man (following prolonged febrile seizures or traumatic brain injury) is associated with multiple changes in the function and subunit composition of ion channels and receptor molecules. Optimal screening of novel AEDs, both for efficacy and side effects, requires models with receptor and ion channel changes similar to those in the target human syndrome.

Post-insult exposure to (±) kavain potentiates N-methyl- d-aspartate toxicity in the developing hippocampus

Kavapyrone extracts of the pepper plant Piper methysticum Forst. have been reported to be pharmacologically active in the brain by modulating the function of several ionotropic receptor systems and voltage-sensitive ion channels. While kavapyrones have previously demonstrated neuroprotective effects against several forms of neurotoxicity, the possibility remains that perturbed function of neuronal ion transport may prove to be neurotoxic in some instances. The present studies were designed to examine the effects of the kavapyrone, (±) kavain, on viability of organotypic hippocampal explants exposed to the excitotoxin N-methyl- d-aspartate (NMDA). Exposure to (±) kavain (1–600 μM) for 24 h did not alter neuronal viability in the CA1, CA3, or dentate gyrus regions of hippocampal explants. However, higher concentrations of (±) kavain (≥300 μM) produced marked neurotoxicity in the lacunosum moleculare layer of the hippocampus. One hour of exposure to NMDA (20 μM) produced significant neuronal death in both the CA3 and CA1 pyramidal cell regions, effects prevented by co-exposure to MK-801 (30 μM). Co-exposure of explants to (±) kavain (1–100 μM) with NMDA did not alter the severity of NMDA-induced neurotoxicity. However, exposure of NMDA-treated explants to (±) kavain (≥10 μM) for 24 h after insult produced significant increases in neurotoxicity in the CA1 and dentate gyrus regions of explants. In conclusion, while the kavapyrone (±) kavain is neurotoxic only at high concentrations when exposed alone to the developing hippocampus, it appears to adversely affect neuronal recovery following excitotoxic insults.

Three-dimensional reconstruction of single particle electron microscopy: the voltage sensitive sodium channel structure.

Single particle analysis in electron microscopy allows direct observation of the reconstructed three-dimensional structures of protein molecules. This method enables a more comprehensive study of membrane proteins which have been problematic in structural studies using X-ray crystallography. These membrane proteins include the voltage-sensitive ion channel proteins, which play an important rule in neural activities, and have great medical significance. The method described is supported by the development of cryo-electron microscopy and the angular reconstitution method. This review summarizes certain principles governing single particle analysis employing angular reconstitution. This method was applied to our study of the voltage-sensitive sodium channel, and the results are discussed. With improvements in resolutions and statistical analyses, the single particle technique is considered to be advantageous in studies of the structural changes and molecular interactions of protein molecules.

Calcium transient and its propagation by poking a single human microvascular endothelial cell with a heat-blunt-ended micropipette

In order to elucidate signal transduction mechanism on how cells sense mechanical deformation, cytosolic Ca transients, by poking with a heat-blunted glass micropipette, were directly observed in human microvascular endothelial cell (HMEC). Cytosolic Ca transients were observed in almost all the HMECs poked with a micropipette. The transients started at the point of poking, and propagated at the peripheral regions of the poked cell. Those transients were also observed to propagate to the adjacent cells. Neither transient of poked cells nor propagation to adjacent cells was detected under the condition of depletion of extracellular Ca or intracellular Ca stores. Either stretch-activated channel or voltage-sensitive ion channel blockage inhibited significantly Ca transients. Phospholipase C inhibition, resulting in inhibition of inositol 1,4,5-triphosphate (IP 3) production, caused no Ca transient by poking. The results suggested that Ca influx might be a trigger for Ca release from intracellular Ca stores by IP 3 in the case of HMEC poking experiments.

Effect of ginseng total saponin on extracellular dopamine release elicited by local infusion of nicotine into the striatum of freely moving rats.

We investigated the effect of ginseng total saponin (GTS) on nicotine-induced dopamine (DA) release in the striatum of freely moving rats using an in vivo microdialysis technique. In order to further characterize the mechanism by which GTS affects DA release, the effect of GTS on K(+)-induced DA release was also examined. Local infusion of nicotine (1, 5, and 10 mM) into the striatum produced a dose-dependent increase in extracellular DA in dialysate samples (maximal response = 154.0 +/- 10.8%, 308.1 +/- 55.7%, and 499.9 +/- 77.9% over basal levels, respectively). GTS (100 mg/kg i.p.) had no effect on basal levels of extracellular DA. However, GTS inhibited maximal DA release induced by intrastriatal infusion of nicotine (1, 5, and 10 mM) by 35.3%, 36.6%, and 58.5%, respectively. Intra-striatal infusion of high K+ solution (100 mM) produced an increase in extracellular DA in the striatum (maximal response = 796.6 +/- 98.8% over basal levels). However, GTS had no effect on the K(+)-induced increase in extracellular DA. The present study demonstrated that GTS inhibited striatal DA release stimulated by local infusion of nicotine. This may reflect the blocking effect of GTS on the striatum-related behavior induced by nicotine as well as other psychostimulants. The results also suggest that GTS may act on presynaptic nicotinic acetylcholine receptors or receptor-operated Na+ channels in dopaminergic nerve terminals, but not on voltage-sensitive ion channels.

Scorpion neurotoxins: structure/function relationships and application in agriculture

Continued use of non-specific chemical insecticides poses potential risks to the environment and to human health resulting from non-target toxicity and increased insect resistance to these agents. Scorpions produce anti-insect selective polypeptide toxins that bind to and modulate voltage-sensitive ion channels in excitable tissues, thus offering alternative, environmentally safer means for insect pest control. Despite this potential, little is known about their structural elements dictating anti-insect preference, which may be useful for the design of selective insecticides. We used a bacterial system for expression and genetic dissection of two pharmacologically distinct scorpion toxins: alpha and excitatory. By exploiting a multi-disciplinary approach consisting of mutagenesis, protein chemistry, electrophysiology, binding and toxicity assays, and structural studies, we elucidated the bioactive surface of two anti-insect toxins, LqhαIT and Bj-xtrIT. In both polypeptides the bioactive surface is composed of residues surrounding the C-terminal region. In addition, a direct, immediate approach in using the toxin genes was demonstrated by engineering baculoviruses with cDNAs encoding LqhIT2 (depressant toxin), and LqhIT1 (excitatory toxin) resulting in viral vectors with significantly improved insecticidal efficacy. © 2000 Society of Chemical Industry

Mechanical sensitivity of muscle afferents in a nerve treated with colchicine.

The experiments reported here demonstrate that the mechanical sensitivity of peripheral nerve fibres typically seen after injury can be induced without overtly injuring the nerve, but by simply applying colchicine topically to the nerve. In cats anaesthetised with pentobarbitone sodium, the medial gastrocnemius nerve was exposed and 10 mM colchicine applied topically for 15 min. The animals recovered from the operation normally and showed no subsequent motor deficit. Six days later animals were re-anaesthetised, a laminectomy carried out and responses recorded in single afferents at the level of the dorsal root. It was found that many afferents, particularly those with conduction velocities in the group II-III range, had become sensitive to local mechanical stimulation of the nerve in the region treated with colchicine and showed slowly adapting responses to stretch of the nerve. Many of the smaller fibres exhibited spontaneous activity. Mechanically sensitive afferents exhibited impulse conduction blocks at the colchicine-treated site. Some afferents, which appeared to conduct impulses normally through the treated region, were associated with muscle receptors having normal response properties. However, other muscle receptors were clearly abnormal and were insensitive to muscle stretch or contraction or exhibited only phasic responses. When the nerve was cut proximal to the colchicine-treated site, some, but not all, spontaneous activity was abolished. It was subsequently shown using a collision technique that the activity in some axons had its origin in the cell body in the dorsal root ganglion. In one experiment, it was shown that after nerve section proximal to the colchicine-treated region three of five axons switched their activity from a peripheral to a central origin. It is postulated that colchicine disrupts fast axonal transport of mechanically sensitive or voltage-sensitive ion channels, from the cell body to the peripheral terminals of the axons, leading to an accumulation of these channels at the treated site. This induces mechanical sensitivity and spontaneous activity. It is postulated that interruption of a retrogradely transported signal induces the spontaneous activity in the cell body. These experiments suggest that an important influence is exerted by the cell body on the peripheral terminals of mechanoreceptors to confer on them their normal response properties.

Nonequilibrium Response Spectroscopy of Voltage-Sensitive Ion Channel Gating

We describe a new electrophysiological technique called nonequilibrium response spectroscopy, which involves application of rapidly fluctuating (as high as 14 kHz) large-amplitude voltage clamp waveforms to ion channels. As a consequence of the irreversible (in the sense of Carnot) exchange of energy between the fluctuating field and the channel protein, the gating response is exquisitely sensitive to features of the kinetics that are difficult or impossible to adequately resolve by means of traditional stepped potential protocols. Here we focus on the application of dichotomous (telegraph) noise voltage fluctuations, a broadband Markovian colored noise that fluctuates between two values. Because Markov kinetic models of channel gating can be embedded within higher-dimensional Markov models that take into account the effects of the voltage fluctuations, many features of the response of the channels can be calculated algebraically. This makes dichotomous noise and its generalizations uniquely suitable for model selection and kinetic analysis. Although we describe its application to macroscopic ionic current measurements, the nonequilibrium response method can also be applied to gating and single channel current recording techniques. We show how data from the human cardiac isoform (hH1a) of the Na + channel expressed in mammalian cells can be acquired and analyzed, and how these data reveal hidden aspects of the molecular kinetics that are not revealed by conventional methods.

Electrophysiology of embryonic, adult and aged rat hippocampal neurons in serum-free culture

Methods were recently developed for culturing neurons from adult rat hippocampus using the serum-free medium Neurobasal with B27 supplement. To determine whether adult cultured neurons have normal electrical properties, we studied cultures from rats of three age groups: (1) embryonic; (2) 10–11 months old and (3) 35–36 months old. Neurons had a polarized morphology with a large branching apical dendrite and small basal dendrites. Mean resting potentials were similar in the three age groups. All neurons had nonlinear current-voltage relationships, indicating the presence of voltage-sensitive ion channels. Most neurons had a voltage-sensitive inward current followed by a sustained voltage-sensitive outward current. Tetrodotoxin blocked the inward current, which is likely to be a sodium current. The sustained outward current, which is likely to be a potassium current, reversed at −71 mV. Most neurons exhibited anomalous rectification. Calcium currents were present in both embryonic and adult neurons. Embryonic neurons would sometimes fire multiple action potentials but adult neurons fired only single action potentials. Our results indicate that both embryonic and adult cultured neurons retain a clearly neuronal electrophysiological phenotype in Neurobasal/B27 serum-free medium.

Short circuit current in rat adrenal capsule/glomerulosa

Angiotensin II (AII) and potassium are the primary regulators of aldosterone secretion by adrenal zona glomerulosa cells. Electrophysiological studies using isolated adrenal tissues and dispersed zona glomerulosa cells show that stimulation by these secretagogues results in depolarisation of the plasma membrane and the opening of voltage-sensitive ion channels. The concept that these cells act together to create a polarity within the gland has not been examined.Whole adrenal capsule/glomerulosa preparations were studied in Ussings chambers. An increase in [KC1] to either side of the capsule resulted in a concentration-related change in short-circuit current (SCC). KC1 added externally caused an increase in SCC, indicating net inward movement of positive ions or net outward movement of negative ions. Internal KC1 had a smaller opposite effect. Use of the non-specific potassium channel blocker tetraethylammonium chloride (TEA) resulted in an increase in SCC regardless of which side the addition was made, although on occasions the responses to TEA and internal KC1 were unexpectedly reversed. The aldosterone antagonist spironolactone produced a variable change in SCC suggesting an autocrine/paracrine role for aldosterone on the adrenal cortex. No responses were observed with the addition of AII, ACTH or aldosterone, though these may be present in excess. The results suggest that ion gradients may be created by stimulation that conceivably have a role in cellular organisation.

RNA Editing in the Drosophila Dmcaia Calcium-Channel α1 Subunit Transcript

Messenger RNA editing of transcripts encoding voltage-sensitive ion channels has not been extensively analyzed - least of all in Drosophila, for which several channel-encoding genes are known. Previous sequence studies of D. melanogaster's cacophony gene, which encodes an α1 calcium-channel subunit called Dmca 1 A, suggested that several nucleotides within the ORF of the primary transcript are edited such that “A-to-G” substitutions occur (these two nucleotides being the adenine that is found at the relevant sites in the sense strand of genomic DNA or the primary transcript, compared to the substitution of gua-nine that is detected at the level of cDNA analysis). Such A-to-G changes are the same kind of post-transcriptional variations originally discovered - in a neurobiological context -for a ligand-sensitive channel in vertebrates. Here, we extracted RNA from adult flies and revealed, by RT-PCR and restriction-enzyme analyses, that transcript heterogeneity exists in vivo for three distinct edited sites within the cac-encoded RNA. Each such nucleotide would lead to channel variability at the level of the DmcalA polypeptide. Owing to cacophony being originally identified as a “behavioral gene,” the possible significance of DmcalA RNA editing for influencing the relevant neuro-functional phenotypes is discussed.

Recent advances in potassium channel modulation.

The opening and closing of potassium (K) channels is the basic mechanism which underlies the control of many cellular excitatory processes. Since the intracellular [K+] of approximately 150 mM is much greater than the extracellular value of 5 mM, the opening of a significant number of K channels results in K + efflux. This flow of ionic current dominates the influence of other ionic conductances, and the membrane potential of the cell moves towards the potassium equilibrium potential (EK; approximately -90 mV).The resulting hyperpolarisation reduces the open probability of voltage-sensitive ion channels which are permeable to Ca2+ and to Na+, resulting in inhibition of cellular functions.KeywordsChannel StructureCystic Fibrosis Transmembrane RegulatorNiflumic AcidBladder SelectivityKATP OpenerThese keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Update on the mechanism of action of antiepileptic drugs.

Novel antiepileptic drugs (AEDs) are thought to act on voltage-sensitive ion channels, on inhibitory neurotransmission or on excitatory neurotransmission. Two successful examples of rational AED design that potentiate GABA-mediated inhibition are vigabatrin (VGB) by irreversible inhibition of GABA-transaminase, and tiagabine (TGB) by blocking GABA uptake. Lamotrigine (LTG) prolongs inactivation of voltage-dependent sodium channels. The anticonvulsant action of remacemide (RCM) is probably largely due to blockade of NMDA receptors and prolonged inactivation of sodium channels induced by its desglycinated metabolite. Felbamate (FBM) apparently blocks NMDA receptors, potentiates GABA-mediated responses, blocks L-type calcium channels, and possibly also prolongs sodium channel inactivation. Similarly, topiramate (TPM) has multiple probable sites of action, including sodium channels, GABA receptors, and glutamate (AMPA) receptors. Gabapentin (GBP) apparently has a completely novel type of action, probably involving potentiation of GABA-mediated inhibition and possibly also inactivation of sodium channels. The therapeutic advantages of the novel AEDs are as yet only partially explained by our present understanding of their mechanisms of action.