Concentrations Of Veratridine Research Articles

Intrathecal Veratridine Administration Increases Minimum Alveolar Concentration in Rats

Results from several studies point to sodium channels as potential mediators of the immobility produced by inhaled anesthetics. We hypothesized that the intrathecal administration of veratridine, a drug that enhances the activity or effect of sodium channels, should increase MAC. We measured the change in isoflurane MAC caused by intrathecal infusion of various concentrations of veratridine into the lumbothoracic subarachnoid space of rats. We compared these result with those obtained from intracerebroventricular infusion. As predicted, intrathecal infusion of veratridine increased MAC. The greatest infused concentration (25 microM) also produced neuronal injury in the hindlimbs of two rats and decreased the peak effect on MAC. A concentration of 1.6 microM produced the largest (21%) increase in MAC. Intraventricular infusion of 1.6 and 6.4 microM veratridine did not alter MAC. Rats given 25 microM died. Intrathecal administration of veratradine increases MAC of isoflurane, a finding consistent with a role for sodium channels as potential mediators of the immobility produced by inhaled anesthetics.

Paradoxical nifedipine facilitation of 45Ca uptake into rat hippocampal synaptosomes

Nifedipine has a high incidence of neurologic adverse reactions as compared with other dihydropyridine Ca v1 (L-type) channel blockers used for treating cardiovascular diseases. The mechanism mediating neuronal excitation by nifedipine is still in debate. Nifedipine caused a dual role on veratridine-induced 45Ca uptake by rat hippocampal synaptosomes. In the nanomolar range (0.001–0.3 μM), nifedipine decreased 45Ca uptake in a cadmium-sensitive manner. In contrast with nitrendipine (0.001–10 μM), nifedipine consistently facilitated 45Ca accumulation when used in low micromolar concentrations (0.3–10 μM). The cadmium-insensitive nifedipine facilitation became less evident upon increasing veratridine concentration from 5 to 20 μM and was not detected when the synaptosomes where depolarised with 30 mM KCl. Na + substitution by N-methyl- d-glucamine (132 mM) or blockade of Na + currents with tetrodotoxin (1 μM) both prevented nifedipine excitation. The Na +/Ca 2+-exchanger inhibitor, KB-R7943 (3–50 μM), did not reproduce nifedipine actions. Data suggest that tetrodotoxin-sensitive Na + channels may operate paradoxical nifedipine facilitation of 45Ca uptake by rat hippocampal synaptosomes.

Natural products from ginseng inhibit [ 3H]batrachotoxinin A 20-α-benzoate binding to Na + channels in mammalian brain

A [ 3H]batrachotoxinin A-20α-benzoate ([ 3H]BTX-B) binding assay was used to investigate the interaction of two ginseng aglycones (20( S)protopanaxadiol and 20( S)protopanaxatriol) and Rh 2 (a monoglucoside of 20( S)protopanaxadiol) with voltage-gated sodium channels in mouse brain. All compounds inhibited the binding of [ 3H]BTX-B and IC 50s were established at 42 μM (20( S)protopanaxadiol), 79 μM (20( S)protopanaxatriol) and 162 μM (Rh 2). Scatchard analysis confirmed that 20( S)protopanaxadiol and Rh-2 reduced the B max of [ 3H]BTX-B binding while Rh 2 also increased the K d. At IC 50 concentrations and above, 20( S)protopanaxadiol and Rh 2 increased the dissociation of the [ 3H]BTX-B:sodium channel complex above that produced by a saturating concentration of veratridine, but failed to reduce the rate of association of [ 3H]BTX-B with sodium channels. Reversal of the inhibition of [ 3H]BTX-B binding by 20( S)protopanaxadiol and Rh 2 occurred slowly. We conclude that the 20( S)protopanaxadiol and the less potent inhibitor Rh 2 destabilize BTX-B-activated sodium channels through non-covalent allosteric modification of neurotoxin binding site 2.

The binding of BmK abT, a unique neurotoxin, to mammal brain and insect Na + channels using biosensor

The binding properties of BmK abT (a novel neurotoxic polypeptide abT from Chinese scorpion Buthus martensi Karsch), a unique neurotoxin from Chinese scorpion, on mammal brain and insect sodium channels were investigated using the BIAcore assay. Results showed that BmK abT could bind to rat brain synaptosomes with an association rate constant of about 2.49×10 6 M −1 s −1 and a dissociation rate constant of about 1.57×10 −4 s −1, and to Heliothis nerve cord synaptosomes with an association rate constant of about 1.21×10 7 M −1 s −1 and a dissociation rate constant of about 0.99×10 −3 s −1. The binding of BmK abT to rat brain synaptosomes could be partially inhibited by increasing the membrane potential, but not by BmK AS (a novel active polypeptide AS from B. martensi Karsch), BmK IT2 (a depressant insect-selective toxin IT2 from B. martensi Karsch), and BmK I (an α-like anti-mammal toxin I from B. martensi Karsch). Binding was not modulated by veratridine. In addition, the binding of BmK abT to Heliothis nerve cord synaptosomes was significantly enhanced by increasing the membrane potential and veratridine concentration and was inhibited by BmK IT2, but not by BmK AS or BmK I. The results suggest that BmK abT binds to a distinct receptor site on mammal brain Na + channels and associates with a related site for depressant insect-selective toxins on insect sodium channels.

The effect of boromycin on the Ca2+ homeostasis.

A boron-containing antibiotic, boromycin (BM), was found to influence the Ca2+ homeostasis in both excitable and non-excitable cells. In non-excitable cells (human erythrocytes and leucocytes) it inhibited the resting passive 45Ca2+ transport in 10(-6)-10(-5) mol/L concentrations. In human erythrocytes, the passive 15Ca2+ transport induced by the presence of 1 mmol/L NaVO3 was inhibited by boromycin (90% inhibition) as well. The inhibitory effect of BM on the NaVO3-induced passive 45Ca2+ transport was diminished in the presence of inhibitory concentrations of nifedipine (10 micromol/L -60% inhibition) or of those of K+o (75 mmol/L -20% inhibition). On the other hand, in rat brain synaptosomes, and rat cardiomyocytes, BM stimulated the passive 45Ca2+ transport in 'resting' cells at similar concentrations. In rat cardiomyocytes the stimulation was transient. The stimulatory effect on the passive 45Ca2+ transport in rat brain synaptosomes was accompanied with the increase of cytoplasmic Ca2+ concentration measured by means of the entrapped fluorescent Ca2+ chelator fura-2. The stimulatory effect of BM was diminished when synaptosomes were pre-treated with veratridine (10 micromol/L) which itself stimulated the passive 45Ca2+ transport. At saturating concentrations of veratridine, no stimulatory effect of BM was observed. These results could be explained by the indirect interaction of BM with both Ca2+ and Na+ transport systems via transmembrane ionic gradients of monovalent cations and could be useful in determining whether the cells belong to excitable, or non-excitable cells.

Short- and long-term differential effects of neuroprotective drug NS-7 on voltage-dependent sodium channels in adrenal chromaffin cells.

In cultured bovine adrenal chromaffin cells, NS-7 [4-(4-fluorophenyl)-2-methyl-6-(5-piperidinopentyloxy) pyrimidine hydrochloride], a newly-synthesized neuroprotective drug, inhibited veratridine-induced (22)Na(+) influx via voltage-dependent Na(+) channels (IC(50)=11.4 microM). The inhibition by NS-7 occurred in the presence of ouabain, an inhibitor of Na(+),K(+) ATPase, but disappeared at higher concentration of veratridine, and upon the washout of NS-7. NS-7 attenuated veratridine-induced (45)Ca(2+) influx via voltage-dependent Ca(2+) channels (IC(50)=20.0 microM) and catecholamine secretion (IC(50)=25.8 microM). Chronic (>/=12 h) treatment of cells with NS-7 increased cell surface [(3)H]-STX binding by 86% (EC(50)=10.5 microM; t(1/2)=27 h), but did not alter the K(D) value; it was prevented by cycloheximide, an inhibitor of protein synthesis, or brefeldin A, an inhibitor of vesicular transport from the trans-Golgi network, but was not associated with increased levels of Na(+) channel alpha- and beta(1)-subunit mRNAs. In cells subjected to chronic NS-7 treatment, (22)Na(+) influx caused by veratridine (site 2 toxin), alpha-scorpion venom (site 3 toxin) or beta-scorpion venom (site 4 toxin) was suppressed even after the extensive washout of NS-7, and veratridine-induced (22)Na(+) influx remained depressed even at higher concentration of veratridine; however, either alpha- or beta-scorpion venom, or Ptychodiscus brevis toxin-3 (site 5 toxin) enhanced veratridine-induced (22)Na(+) influx as in nontreated cells. These results suggest that in the acute treatment, NS-7 binds to the site 2 and reversibly inhibits Na(+) channels, thereby reducing Ca(2+) channel gating and catecholamine secretion. Chronic treatment with NS-7 up-regulates cell surface Na(+) channels via translational and externalization events, but persistently inhibits Na(+) channel gating without impairing the cooperative interaction between the functional domains of Na(+) channels.

Sodium overload through voltage-dependent Na + channels induces necrosis and apoptosis of rat superior cervical ganglion cells in vitro

Using the failure to exclude trypan blue as a criterion for cell death, we found that veratridine, the voltage-dependent Na + channel activator, exerted its toxicity to cultured sympathetic neurons in a dose-dependent manner (half-maximal toxicity occurred at 2 μM). The co-presence of tetrodotoxin completely reversed the toxicity only at concentrations of veratridine <20μM. Veratridine neurotoxicity was due to the influx of Na +; a medium low in Na + (36 mM) completely abolished its neurotoxicity, whereas a Ca 2+-free medium did not attenuate its neurotoxicity. Furthermore, the buffering action of 1,2-Bis-(2-aminophenoxy)ethane-N,N,N′,N′,-tetraacetate (BAPTA) on veratridine-induced increase in intracellular Ca 2+ levels neither blocked veratridine neurotoxicity in normal medium, nor attenuated the low Na + effect. Elevated K + effectively blocked veratridine neurotoxicity in a Ca 2+-dependent manner. Cytoplasmic pH measurements using a fluorescent pH indicator demonstrated that cellular acidification (from pH 7.0 to pH 6.5) occurred upon treatment with veratridine. Both veratridine-induced acidification and cell death were ameliorated by 5-(N-ethyl-N-isopropyl)amiloride, the specific inhibitor of the Na +/H + exchanger (IC 50 = 0.5 μM). Finally, necrosis occurred predominantly in veratridine neurotoxicity, but both staining with bis-benzimide and TUNEL analysis showed nuclear features of apoptosis in sympathetic neurons undergoing cell death.

Selective inhibition by riluzole of voltage-dependent sodium channels and catecholamine secretion in adrenal chromaffin cells.

We examined the effects of riluzole, a neuroprotective drug, on voltage-dependent Na channels, nicotinic receptors, and voltage-dependent Ca channels, as well as catecholamine secretion, in comparison with those of verapamil and nicardipine, in primary cultures of bovine adrenal chromaffin cells. Riluzole inhibited veratridine-induced 22Na influx via voltage-dependent Na channels even in the presence of ouabain, an inhibitor of Na,K-ATPase. Blockade of Na channels by riluzole was concentration-dependent with an IC50 of 5.3 microM. It was associated with a similar concentration-related reduction of veratridine-induced 45Ca influx via voltage-dependent Ca channels, and of catecholamine secretion. Riluzole had no effect on 45Ca influx caused by high K, which directly activates voltage-dependent Ca channels, and on nicotine-induced 22Na influx, which passes through the nicotinic receptors. Verapamil and nicardipine attenuated 22Na influx caused by veratridine or nicotine at the same concentrations as they suppressed high K-induced 45Ca influx. The inhibitory effect of riluzole on veratridine-induced 22Na influx disappeared at high concentrations of veratridine. A potentiation of veratridine (site 2 toxin)-induced 22Na influx caused by alpha-scorpion venom (site 3 toxin), beta-scorpion venom (site 4 toxin), or brevetoxin PbTx-3 (site 5 toxin), occurred in the presence of riluzole in the same manner as in control cells. These results suggest that riluzole binds to the veratridine site in voltage-dependent Na channels. It does not impair the cooperative interaction between the functional peptide segments of Na channels, but selectively inhibits gating of Na channels, thereby reducing Ca influx via Ca channels and catecholamine secretion. In contrast, verapamil and nicardipine suppress Na influx both Na channels and nicotinic receptors.

The relaxing effect of BDF 9148 on the KCl-contracted aorta isolated from normo- and hyper-tensive rats

BDF 9148, a positive cardiac inotrope, relaxes the rat isolated portal vein and the KCl-contracted rat aorta. The aims of our study were to determine the mechanism of action of BDF 9148, and to ascertain whether the relaxing effect of BDF 9148 was maintained in the presence of the hypertrophy associated with hypertension, by investigating the effects of BDF 9148 on the contractility and electrophysiology of aortae of Wistar Kyoto normo-tensive rats (WKY) and Spontaneously Hypertensive Rats (SHRs). High concentrations of veratridine contracted the quiescent rat aorta. BDF 9148 had no effect on the quiescent, but relaxed the KCl-contracted WKY and SHR aorta by a tetrodotoxin insensitive mechanism, and these relaxations decreased with age but were not greatly altered by hypertrophy. The verapamil relaxations of the KCl-contracted aorta were not altered by age or hypertrophy. The ability of KCl to depolarise the aorta was reversed by verapamil, but not by BDF 9148. On the contracted rat aorta, the relaxant responses to acetylcholine were abolished by removal of the endothelium but potentiated by IBMX (10[-6] M), and the responses to isoprenaline were inhibited by propranolol (10[-6] M) but potentiated by forskolin (10[-7] M). The relaxation responses of the KCl-contracted aorta to BDF 9148 were not altered by removal of the endothelium, or by propranolol, forskolin and IBMX. In summary, the effects of verapamil and BDF 9148 on the aorta are different, and thus it is unlikely that the relaxant responses to BDF 9148 on the aorta are due to calcium channel blocking activity. The mechanism of the relaxant effect of BDF 9148 on the aorta remains unknown, but we have shown the response is endothelium-independent, and not mediated by sodium channel opening, hyperpolarization, beta-adrenoceptors, or by stimulating adenylate cyclase or guanylate cyclase.

Up‐Regulation of Sodium Channel Subunit mRNAs and Their Cell Surface Expression by Antiepileptic Valproic Acid: Activation of Calcium Channel and Catecholamine Secretion in Adrenal Chromaffin Cells

Treatment of cultured bovine adrenal chromaffin cells with a therapeutic concentration (0.6 mM) of valproic acid (VPA) for > 24 h caused a time-dependent (t1/2 = 74 h) increase in [3H]saxitoxin binding up to 1.4-fold without altering the KD value; it was prevented by the simultaneous treatment with cycloheximide (an inhibitor of protein synthesis). VPA also raised Na+ channel alpha- and beta 1-subunit mRNA levels 1.4- and 1.7-fold at 24 h, and 1.6- and 1.8-fold at 72 h, respectively. Chronic (but not acute) exposure to VPA enhanced 22Na+ influx caused by various concentrations of veratridine 1.4-2.1-fold, even when assayed in the presence of Na+,K(+)-ATPase inhibitor, but did not change the EC50 value of veratridine. Ptychodiscus brevis toxin-3 allosterically potentiated veratridine-induced 22Na+ influx by approximately 2-fold in VPA-treated cells as in nontreated cells. Long-term treatment with VPA augmented veratridine-induced 45Ca2+ influx via voltage-dependent Ca2+ channels and catecholamine secretion, but had no effect on 45Ca2+ influx and catecholamine secretion caused by high K+ (a direct activation of voltage-dependent Ca2+ channels). Chronic treatment with VPA also enhanced nicotine-induced 22Na+ influx via the nicotinic receptor-ion channel complex 1.2-1.4-fold with little change in the EC50 value of nicotine, thereby increasing the nicotine-induced 45Ca2+ influx via voltage-dependent Ca2+ channels and catecholamine secretion. These results suggest that chronic treatment with VPA up-regulates cell surface expression of Na+ channels via the transcription/translation-dependent mechanisms, and probably of nicotinic receptors, thereby resulting in the enhancement of Ca2+ channel gating and catecholamine secretion.

Evidence for the induction of repetitive action potentials in synaptosomes by K+-channel inhibitors: an analysis of plasma membrane ion fluxes.

The effects of four K+-channel inhibitors on synaptosomal free Ca2+ concentrations and 86Rb+ fluxes are analysed. 4-Aminopyridine, alpha-dendrotoxin, charybdotoxin, and tetraethylammonium all increase the free Ca2+ concentration, although their potencies differ widely. In each case, the elevation in free Ca2+ concentration is reversed by the subsequent addition of tetrodotoxin. The transient 86Rb+ efflux from preequilibrated synaptosomes induced with high concentrations of veratridine is partially inhibited by 4-aminopyridine and alpha-dendrotoxin. In contrast, when 4-aminopyridine or alpha-dendrotoxin is added to polarized synaptosomes, and enhanced 86Rb+ flux is seen, both for uptake and for efflux with no change in the total 86Rb+/K+ content of the synaptosomes and with only a slight time-averaged plasma membrane depolarization (6.4 and 3.3 mV, respectively). The enhancements of flux by 4-aminopyridine or alpha-dendrotoxin are sensitive to ouabain and/or to tetrodotoxin. Furthermore, these flux changes show the same concentration dependencies as the blocked component of veratridine-stimulated 86Rb+ efflux, the elevation of free Ca2+ concentration, and the facilitation of glutamate exocytosis that are elicited by 4-aminopyridine or alpha-dendrotoxin. It is concluded that these findings support the proposal of spontaneous, repetitive firing of synaptosomes evoked by K+-channel inhibitors and that the enhanced 86Rb+ flux is a consequence of the activity of 4-aminopyridine- and alpha-dendrotoxin-insensitive K+ channels during these action potentials.

Effects of veratridine on Na and Ca currents in frog skeletal muscle

1. 1. Voltage-clamp experiments were performed to determine the effects of veratridine on Na and Ca currents in frog skeletal muscle fibres. 2. 2. Veratridine (1 μM) did not affect the kinetics of the fast Na current but it did induce a slowly inactivating tetrodotoxin-sensitive inward current that was apparent after Na current inactivation. This slow current had a peak amplitude of 6.7 ± 0.7 μA/cm 2 at −20mV and decayed monoexponentially with a time constant of 606 ± 77 ms. 3. 3. The slow current had a voltage-dependence for activation that was similar to that of the fast Na current. Single depolarizing prepulses that induced complete inactivation of the fast Na channels, prevented development of the slow current. Trains of brief depolarizations at increasing frequencies increased the amplitude of the slow current. These results suggest that the slow current may be mediated by veratridine modified Na channels that must be in the open position. 4. 4. The low concentration of veratridine (1 μM) did not affect the Ca current, while 100 μM veratridine reversibly suppressed the Ca current and shifted its peak current-voltage relation towards more negative potentials. Thus, veratridine appears not to be a selective fast Na channel modifier as it may also alter Ca channel gating properties in skeletal muscle fibres.

Veratridine blocks voltage-gated potassium current in human T lymphocytes and in mouse neuroblastoma cells.

(i) Effects of veratridine on ionic conductances of human peripheral blood T lymphocytes have been investigated using the whole-cell patch-clamp technique. (ii) Veratridine reduces the net outward current evoked by membrane depolarizations. The reduction originates from block of a 4-aminopyridine-sensitive, voltage-gated K+ current. (iii) Human T lymphocytes do not appear to express voltage-gated Na+ channels, since inward currents are observed neither in control nor in veratridine- and bretylium-exposed lymphocytes. (iv) The effect of veratridine consists of an increase in the rate of decay of the voltage-gated K+ current and a reduction of the peak current amplitude. Both effects depend on veratridine concentration. Half-maximum block occurs at 97 microM and the time constant of decay is reduced by 50% at 54 microM of veratridine. (v) Possible mechanisms of veratridine action are discussed. The increased rate of K+ current decay is most likely due to open channel block. The decrease of current amplitude may involve an additional mechanism. (vi) In cultured mouse neuroblastoma N1E-115 cells, veratridine blocks a component of voltage-gated K+ current, in addition to its effect on voltage-gated Na+ current. This result shows that the novel effect of veratridine is not confined to lymphocytes.

Release of endogenous benzodiazepine receptor ligands (endozepines) from cultured neurons.

Endozepines are naturally occurring small organic molecules, devoid of peptidic bonds and halogens, that act as allosteric modulators of the GABAA receptor through their actions at the benzodiazepine binding site. Endozepines are present in physiologically significant amounts in the brain and can act as potent positive allosteric modulators of the GABAA receptor. In this study, 3 endozepines present in cultured cerebellar granule cells were found to be released from neurons in a potassium-stimulated, calcium-dependent fashion. This release could also be mimicked by increasing concentrations of veratridine. Although endozepines were also found in cultured astrocytes, they could not be released in significant amounts by potassium depolarization. Differential release under depolarizing conditions and granule cell content of the various endozepines suggested a possible metabolic relationship between these two processes.

Relation between veratridine reaction dynamics and macroscopic Na current in single cardiac cells.

Veratridine modification of Na current was examined in single dissociated ventricular myocytes from late-fetal rats. Extracellularly applied veratridine reduced peak Na current and induced a noninactivating current during the depolarizing pulse and an inward tail current that decayed exponentially (tau = 226 ms) after repolarization. The effect was quantitated as tail current amplitude, Itail (measured 10 ms after repolarization), relative to the maximum amplitude induced by a combination of 100 microM veratridine and 1 microM BDF 9145 (which removes inactivation) in the same cell. Saturation curves for Itail were predicted on the assumption of reversible veratridine binding to open Na channels during the pulse with reaction rate constants determined previously in the same type of cell at single Na channels comodified with BDF 9145. Experimental relationships between veratridine concentration and Itail confirmed those predicted by showing (a) half-maximum effect near 60 microM veratridine and no saturation up to 300 microM in cells with normally inactivating Na channels, and (b) half-maximum effect near 3.5 microM and saturation at 30 microM in cells treated with BDF 9145. Due to its known suppressive effect on single channel conductance, veratridine induced a progressive, but partial reduction of noninactivating Na current during the 50-ms depolarizations in the presence of BDF 9145, the kinetics of which were consistent with veratridine association kinetics in showing a decrease in time constant from 57 to 22 and 11 ms, when veratridine concentration was raised from 3 to 10 and 30 microM, respectively. As predicted for a dissociation process, the tail current time constant was insensitive to veratridine concentration in the range from 1 to 300 microM. In conclusion, we have shown that macroscopic Na current of a veratridine-treated cardiomyocyte can be quantitatively predicted on the assumption of a direct relationship between veratridine binding dynamics and Na current and as such can be successfully used to analyze molecular properties of the veratridine receptor site at the cardiac Na channel.

Insecticidal dihydropyrazoles antagonize the depolarizing action of veratridine in mammalian synaptosomes as measured with a voltage-sensitive dye

The actions of two dihydropyrazoles, RH-3421 and RH-5529, on chemically evoked depolarization of the nerve terminal have been examined in a synaptosomal preparation isolated from rat brain. Both compounds antagonized the change in membrane potential produced by veratridine in a dose-dependent manner. In this respect, RH-3421 was a more effective blocker than RH-5529, and inhibition with each dihydropyrazole, on a percentage basis, was greater at lower concentrations of veratridine. At high concentrations, dihydropyrazoles were unable to influence the depolarization of synaptosomes brought about by elevated potassium. In addition, no effect of dihydropyrazoles on the membrane potential of resting synaptosomes was observed. RH-5529 is known to inhibit the potassium-induced release of [ 3H]GABA, while RH-3421 and RH-5529 reduce the resting release of transmitter. The inability of (i) RH-5529 to influence potassium-induced depolarization of synaptosomes and (ii) either analogue to affect the membrane potential of resting synaptosomes demonstrates that inhibition of transmitter release with dihydropyrazoles under these circumstances does not result from effects on membrane potential. The pattern of inhibition observed with RH-3421 on veratridine- and potassium-induced depolarization of nerve terminals accords with previous results which show that stimulation of presynaptic release of [ 3H]GABA by sodium channel-directed activators is selectively blocked by RH-3421.

A Preferential Inhibition of Impulses in C-fibers of the Rabbit Vagus Nerve by Veratridine, an Activator of Sodium Channels

In seeking a means to reverse local anesthetic block of peripheral nerve, we examined the actions of veratridine (VTD), an agent known to antagonize competitively the binding of local anesthetics to Na channels. The actions of VTD, a steroidal alkaloid "activator" of voltage-gated Na channels, were studied in the rabbit vagus nerve by two methods. In one, the effects of VTD on compound action potentials (APc) propagating through a "veratrinized" segment (11-mm) of nerve were measured by extracellular recording. Single volleys of impulses were unaffected by VTD, but trains of impulses, triggered by repetitive stimulation, were selectively diminished. This "use-dependent" reduction was greatest for the C-fiber component of the APc, less for B-fibers, and inconsequential for A-fibers. Use-dependent inhibition was enhanced by higher stimulation frequency and by increased VTD concentration, and reversed rapidly when stimulation ceased. If the nerve sheath remained intact, the rate of VTD action was far less than in desheathed nerves, but the effects were the same. In the other experimental system, membrane potentials were measured in the veratrinized region of the nerve by a sucrose-gap method. Repetitive stimulation, particularly of C-fibers, produced a cumulative VTD-induced depolarization (VID) that was sustained over several seconds and during which the C-fiber APc was selectively reduced. We propose that this local, use-dependent VID provides the means to inhibit impulses propagating through the veratrinized region. The preferential effect of VTD on C-fibers suggests its possibilities as a relatively selective agent for block of impulse trains in nociceptive afferents.

Micromolar concentrations of veratridine activate sodium channels in embryonic cockroach neurones in culture

The mode of action of the alkaloid veratridine has been reinvestigated on cultured cockroach neurones, which are normally inexcitable and do not have a detectable fast sodium current. The whole-cell and cell-attached configurations of the patch-clamp technique were used to record the macroscopic and single channel currents, respectively. Concentrations of veratridine ranging from 10(-8) to 10(-5) M were found to induce a small tetrodotoxin (TTX)-sensitive inward current, which peaked around +10 mV and reversed around +55 mV. This current exhibited a pronounced plateau and was insensitive to changes in the holding potential. Bath application of veratridine induced typical TTX-sensitive inwardly-directed single-channel activity, falling into two (apparently coupled) categories of events: first, relatively large events (1 pA at a hyperpolarized potential of -125 mV relative to rest) of short duration and, second, small bursting events (0.4 pA under similar conditions) of slightly longer duration. Pipette application of similar concentrations of veratridine had similar effects in that two categories of events were observed: first, bursts of large events with multiple conductance states and, second, small events of very long duration. The current/voltage relationship of these events was linear for the voltage range studied and the (extrapolated) reversal potential approximated +110 mV. These results support the hypothesis that veratridine, in small concentrations, induces a slow voltage-dependent activation of TTX-sensitive sodium channels, independent of the fast activating and inactivating sodium channels involved in action potential generation.

Presynaptic Receptors and Autonomic Effectors

Agonist interactions with release processes do not validate autoreceptor operation. Instead, the data suggests that autoreceptors function as homoreceptors. Declining efficacy of agonist inhibition of transmitter release with increasing stimulation "intensity" is not assignable to competition with endogenous transmitter for a finite population of receptors. Heteroreceptor activation also reveals a pattern of declining efficacy with increasing intensity. Inhibition of stimulation-induced release by agonists also does not correlate inversely with that of antagonist effectiveness, as it should if both effects are linked to biophase levels of transmitter. Further, per pulse release of transmitter, in the absence of drugs, does not comply with expectations for autoinhibition. Experiments with tetrodotoxin, and study of the magnitudes of agonist and antagonist effects suggest that in the periphery putative autoreceptors may actually be homoreceptors, e.g., sympathetic nerve terminal receptors responsive to circulating catecholamines. In the central nervous system the paracrine secretion of transmitter may invoke homoreceptor activation rather than autoreceptors. Such activation may include, for example, the release of adrenaline from one set of fibers in the hypothalamus to act on dopaminergic or noradrenergic fibers or the release of noradrenaline from the terminals of some noradrenergic fibers in the cortex to activate alpha2 receptors on other cortical noradrenergic fibers, the latter with a somewhat different function in the same brain region. The action of antagonist drugs to enhance transmitter release may be direct on nerve membranes in particular on sodium channels, and often unrelated to feedback regulation. This possibility is discussed by me elsewhere in this volume. It is shown that yohimbine and a low concentration of veratridine have similar and nonadditive effects on transmitter release.

Veratridine-induced oscillations in membrane potential of cultured rat skeletal muscle: role of the Na-K pump.

1. The acute effects of veratridine on membrane potential (Em) and Na-K pump activity in cultured skeletal muscle were examined. 2. At a concentration of 10(-4) M, veratridine caused depolarization of Em and a decrease in Na-K pump activity. At concentrations of 10(-5) and 10(-6) M, veratridine caused oscillations of Em and an increase in Na-K pump activity compared to untreated, control cells. The oscillations consisted of depolarization to about -40 mV followed by hyperpolarization to about -90 mV; the level of hyperpolarization was higher at 37 than at 23 degrees C. 3. Veratridine-induced oscillations could be prevented by pretreatment with tetrodotoxin (10(-6) M) and blocked or prevented by ouabain, which depolarizes Em of cultured myotubes. In contrast, depolarization of Em to -60 mV by excess K+ did not alter the amplitude or frequency of the oscillations. 4. The results demonstrate that veratridine-induced increase in Na influx both depolarizes cultured myotubes and increases the activity of the Na-K pump, which repolarizes Em to levels higher than control. This sequence accounts for veratridine-induced oscillations in Em. High concentrations of veratridine cause only depolarization of Em and inhibition of Na-K pump activity.