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Biomedical subjects

G R Strichartz

Publications and source records attributed to G R Strichartz.

At least 73 records · Page 4Linked to original sources
PubMed1987

Effects of venom from Conus striatus on the delayed rectifier potassium current of molluscan neurons.

The effect of crude venom extracted from venom ducts of Conus striatus on the delayed rectifier potassium current (IK) of the marine mollusc Aplysia californica was studied using voltage clamp techniques. Initial experiments indicated that the venom had phospholipase activity which destroyed the cells. The use of phospholipase inhibitors prevented destruction of the cell and permitted long-term electrophysiological measurements to be made. Application of the venom to unclamped cells caused a dramatic increase in the frequency of action potentials associated with a depolarization of the membrane potential. A broadening of the action potential was also observed. Three separate effects of the venom were observed on IK in voltage clamped cells: an increase in peak current (effect I), a slowing of both the activation and inactivation kinetics (effect K) and a decrease in the peak current (effect D). All three effects were dose dependent and both effects on peak current were greater at more depolarized membrane potentials. The data suggest that the three effects on IK are caused by different components of the venom. Effect D appears to be caused by a heat-labile compound of molecular weight greater than 50,000, effect I by a heat-stable compound of less than 50,000 and effect K by a heat-stable compound of intermediate molecular size.

Action Potentials↗
PubMed1987

Limited nerve impulse blockade by "leashed" local anesthetics.

To measure the depth of the local anesthetic binding site within the neuronal membrane, biotin-containing polyethylene glycols having zero, three, and six ethylene glycol subunits were added to the p-amino termini of tetracaine and procaine, thereby interposing a pharmacologically inert "spacer" molecule between the local anesthetic and the biotin moiety. These biotinyl-local anesthetic derivatives produced "tonic" inhibition of the compound action potential of split, desheathed frog sciatic nerves in a concentration-dependent, reversible manner. However, no inhibition of the action potential occurred when sufficient avidin, a 66,000-MW protein that binds four biotins, was present to bind and anchor the biotin-containing end of each derivative outside the plasma membrane. Increasing the "leashed" anesthetic derivative's concentration to 4 times that which reduced impulse height by 50% in the absence of avidin still produced no detectable block when equimolar avidin was present. Apparently, the "spacer" in the derivative compound was too short to permit the avidin-complexed anesthetic to reach its site of action on the sodium channel. In a similar fashion, the local anesthetic derivatives produced "use-dependent" block when drug-treated nerves were stimulated at 40 Hz in the absence of equimolar avidin, but failed to produce "use-dependent" block when equimolar avidin was present. In common with others, we assume that tertiary amine local anesthetics may reach their binding site via hydrophobic (transmembrane) pathways without necessarily entering the cytoplasm. Thus, since our longest local anesthetic derivative, that containing six ethylene glycol subunits, placed the local anesthetic group a maximum of 15-18 A from the surface of the avidin moiety, we conclude that the local anesthetic binding site for block of sodium channels of amphibian nerve must be greater than or equal to 15 A from the outer surface of the plasma membrane.

Action Potentials↗
PubMed1987

Inhibition of sodium currents by local anesthetics in chloramine-T-treated squid axons. The role of channel activation.

In order to test the requirement of Na channel inactivation for the action of local anesthetics, we investigated the inhibitory effects of quaternary and tertiary amine anesthetics on normally inactivating and noninactivating Na currents in squid axons under voltage clamp. Either the enzymatic mixture pronase, or chloramine-T (CT), a noncleaving, oxidizing reagent, was used to abolish Na channel inactivation. We found that both the local anesthetics QX-314 and etidocaine, when perfused internally at 1 mM, elicited a "tonic" (resting) block of Na currents, a "time-dependent" block that increased during single depolarizations, and a "use-dependent" (phasic) block that accumulated as a result of repetitive depolarizations. All three effects occurred in both control and CT-treated axons. As in previous reports, little time-dependent or phasic block by QX-314 appeared in pronase-treated axons, although tonic block remained. Time-dependent block was greatest and fastest at large depolarizations (Em greater than +60 mV) for both the control and CT-treated axons. The recovery kinetics from phasic block were the same in control and CT-modified axons. The voltage dependence of the steady state phasic block in CT-treated axons differed from that in the controls; an 8-10% reduction of the maximum phasic block and a steepening and shift of the voltage dependence in the hyperpolarizing direction resulted from CT treatment. The results show that these anesthetics can bind rapidly to open Na channels in a voltage-dependent manner, with no requirement for fast inactivation. We propose that the rapid phasic blocking reactions in nerve are consequences primarily of channel activation, mediated by binding of anesthetics to open channels, and that the voltage dependence of phasic block arises directly from that of channel activation.

Anesthetics, Local↗
PubMed1987

Dependence of lidocaine potency on pH and PCO2.

Lidocaine solutions with different concentrations of CO2, NaOH, and HCl in two buffering systems were applied to frog sciatic nerves. The peak of the compound action potential (APc) and the firing threshold for single axons were measured. The amount of lidocaine required at steady state to double the firing threshold of single fibers or to reduce the peak of the APc by 40% was used as the index of potency. Acidification with CO2 increased potency (less lidocaine was needed to achieve either criterion), whereas acidification with HCl diminished potency, as compared with alkaline conditions. These results were true whether or not the perineurium was present. Frequency-dependent block (Bf) increased in acid conditions produced by CO2, whereas Bf was less under acid conditions produced with HCl (P less than 0.02). The experiments indicate that CO2 potentiates conduction block with lidocaine either by a direct effect on the membrane or by its indirect action on intracellular pH, but not from effects on the extracellular pH.

Action Potentials↗
PubMed1987

The effect of polyethylene glycol on mammalian nerve impulses.

Polyethylene glycol (PEG) is a polymeric compound used as a vehicle for depot steroid preparations such as methylprednisolone acetate and triamcinolone diacetate injected into the epidural or intrathecal space to relieve low back pain. There have been reports of neurodysfunction associated with these injections, and it has been postulated that the PEG vehicle is the offending agent. Studies supporting such a possibility have, however, relied upon concentrations of PEG higher than those used clinically (3%) or have used PEG in combination with other drugs. Using an in vitro rabbit sheathed-nerve preparation, we investigated the effects of a 1-hr exposure to different concentrations (3-40%) of PEG in Liley solution on the transmission of impulses of the A, B, and C nerve fibers. The 3% and 10% PEG had no effect on mean amplitudes of the compound action potentials (CAPs) nor did they significantly decrease conduction velocity. Twenty percent PEG slightly depressed and 30% markedly decreased CAPs. Both 20% and 30% PEG significantly slowed the conduction velocities of A, B, and C nerve fibers. Forty percent PEG abolished CAPs. With washout CAPs recovered to at least 80% of their baseline levels, and conduction velocities returned toward baseline levels. The pH of the Liley solution decreased with an increasing concentration of PEG, from 7.4 in the control Liley solution to 6.45 in the solution of 40% PEG.(ABSTRACT TRUNCATED AT 250 WORDS)

Action Potentials↗
PubMed1987

Fundamental properties of local anesthetics. I. The dependence of lidocaine's ionization and octanol:buffer partitioning on solvent and temperature.

The protonation equilibrium and hydrophobic character of lidocaine were characterized by its pKa and the octanol:buffer partition coefficients of the charged (P+) and neutral (Po) drug species. These measurements were accomplished by ultraviolet spectrophotometry of pure lidocaine HCl solutions at different temperatures, ionic strengths, and buffer concentrations. Corroboration of the pKa determination by the potentiometric method and of the partition coefficients by gas chromatography validated the general application of the spectrophotometric technique. The pKa increased with decreasing temperature (7.61 +/- 0.06 at 36 degrees C; 7.94 +/- 0.04 at 26 degrees C, in water; mean +/- SD), increasing ionic strength (8.06 +/- 0.02 at 26 degrees C in 0.165 M NaCl) and increasing buffer capacity (8.28 +/- 0.06 at 25 degrees C in 0.15 M NaCl + 20 mM buffer). Octanol:buffer partition coefficients for both the protonated and the neutral species (expressed as mole fractions) increased upon warning: 0.55 +/- 0.04 and 2666 +/- 202, respectively, at 25 degrees C, and 0.75 +/- 0.09 and 3210 +/- 272, respectively, at 36 degrees C. Ionic strength and buffer concentration had no significant effect on either P value. The increase in pKa at lower temperatures coupled with the decreased partition coefficients resulted in a nearly constant concentration of the protonated species in octanol as the system was cooled, whereas the concentration of the neutral species fell by more than 80%. This finding may explain the large increase in the impulse blocking potency of lidocaine observed upon nerve cooling, if the protonated anesthetic species is the more active form of the drug competing with the neutral species for a common binding site.

1-Octanol↗
PubMed1986

Saxitoxin blocks batrachotoxin-modified sodium channels in the node of Ranvier in a voltage-dependent manner.

The inhibition by saxitoxin (STX) of single Na channels incorporated into planar lipid bilayers and modified by batrachotoxin (BTX) previously has been shown to be voltage dependent (Krueger, B.K.,J.F. Worley, and R. J. French, 1983, Nature [Lond.], 303:172-175; Moczydlowski, E., S. Hall, S. S. Garber, G. S. Strichartz, and C. Miller, 1984, J. Gen. Physiol., 84:687-704). We tested for such a voltage dependence of STX block of the Na current in voltage-clamped frog nodes of Ranvier. The block by STX of normal Na channels showed no modulation in response to maintained (20 s) changes of the membrane potential or to a train of brief pulses to potentials more positive than the holding potential. However, when the nodal channels were modified by BTX, the train of pulses produced a modulation of the block of the Na current by STX. The modulation of STX block depended on the voltage of the conditioning pulses and this voltage dependence agreed well with that predicted from the single channel studies over the membrane potential range used in those studies. In addition, we found that the voltage dependence of STX block was manifest only at potentials equal to or more positive than required to activate the channels. Most of the apparent differences among data from single channels in bilayers, equilibrium binding studies of STX, and the experiments described here are resolved by the hypotheses that (a) STX binding to open channels is voltage dependent, and (b) the affinities of STX for closed and inactivated channels are independent of voltage, equal, and less than the open channel affinity at potentials less than 0 mV. Whether these hypotheses apply to the STX block of all Na channels or just of BTX-modified channels remains to be determined.

Animals↗
PubMed1986

Discrimination of muscle and neuronal Na-channel subtypes by binding competition between [3H]saxitoxin and mu-conotoxins.

The effect of two mu-conotoxin peptides on the specific binding of [3H]saxitoxin was examined in isolated plasma membranes of various excitable tissues. mu-Conotoxins GIIIA and GIIIB inhibit [3H]saxitoxin binding in Electrophorus electric organ membranes with similar KdS of approximately equal to 50 X 10(-9) M in a manner consistent with direct competition for a common binding site. GIIIA and GIIIB similarly compete with the majority (80-95%) of [3H]saxitoxin binding sites in rat skeletal muscle with KdS of approximately 25 and approximately 140 X 10(-9) M, respectively. However, the high-affinity saxitoxin sites in lobster axons, rat brain, and rat heart are virtually insensitive to GIIIA concentrations up to 10 microM. These results and previously published data suggest that three Na-channel subtypes can be distinguished on the basis of toxin pharmacology: Na channels of skeletal muscle and Electrophorus electroplax have high affinity for mu-conotoxins and tetrodotoxin, neuronal Na channels have low affinity for mu-conotoxins and high affinity for tetrodotoxin, while heart Na channels and a similar subtype also found in denervated muscle have low affinity for both mu-conotoxin and tetrodotoxin.

Animals↗
PubMed1986

Rapid voltage-dependent dissociation of scorpion alpha-toxins coupled to Na channel inactivation in amphibian myelinated nerves.

The voltage-dependent action of several scorpion alpha-toxins on Na channels was studied in toad myelinated nerve under voltage clamp. These toxins slow the declining phase of macroscopic Na current, apparently by inhibiting an irreversible channel inactivation step and thus permitting channels to reopen from a closed state in depolarized membranes. In this article, we describe the rapid reversal of alpha-toxin action by membrane depolarizations more positive than +20 mV, an effect not achieved by extensive washing. Depolarizations that were increasingly positive and of longer duration caused the toxin to dissociate faster and more completely, but only up to a limiting extent. Repetitive pulses had a cumulative effect equal to that of a single pulse lasting as long as their combined duration. When the membrane of a nonperfused fiber was repolarized, the effects of the toxin returned completely, but if the fiber was perfused during the conditioning procedure, recovery was incomplete and occurred more slowly, as it did at lower applied toxin concentrations. Other alpha-type toxins, from the scorpion Centruroides sculpturatus (IVa) and the sea anemone Anemonia sulcata (ATXII), exhibited similar voltage-dependent binding, though each had its own voltage range and dissociation rate. We suggest that the dissociation of the toxin molecule from the Na channel is coupled to the inactivation process. An equivalent valence for inactivation gating, of less than 1 e per channel, is calculated from the voltage-dependent change in toxin affinity.

Animals↗
PubMed1986

The interaction between the activator agents batrachotoxin and veratridine and the gating processes of neuronal sodium channels.

The depolarization of frog sciatic nerves by the Na channel-activating toxins, batrachotoxin and veratridine, was studied using the sucrose-gap technique. To study the interaction between the activators and the gating processes of Na channels, we measured the depolarizations of unstimulated nerves, of nerves during repetitive stimulation, and of nerves whose Na channel inactivation process had been pharmacologically modified. Stimulation enhanced the rates of depolarization by the activators but did not effect the steady state depolarization values. Of the three inhibitors of Na channel inactivation that were tested (Leiurus alpha-scorpion toxin, chloramine T, and Ni2+), only Leiurus toxin enhanced the potencies of the activators. Neither chloramine T nor Ni2+ had any effect on the steady state level of depolarization produced by either activator. Both chloramine T and Ni2+, however, enhanced the rate of batrachotoxin action, although neither affected the rate of veratridine action. Leiurus toxin also potentiated the effects of the activators in chloramine T-treated nerves. We tested the interaction between the Na channel activators and a class of agents, local anesthetics, that stabilize a non-conducting state of the Na channel. The presence of lidocaine inhibited the depolarization produced by addition of either activator, although the addition of lidocaine subsequent to the development of batrachotoxin-induced depolarization produced repolarization very weakly and slowly. We also found that the lidocaine homologue, RAC 109I, was about 3 times as potent as its stereoisomer, RAC 109II, in its ability both to reduce the compound action potential amplitude and to inhibit the veratridine-induced depolarization.

Action Potentials↗
PubMed1985

Tachykinin antagonists have potent local anaesthetic actions.

Contrary to what would have been expected, an antagonist of substance P (SP) [Arg5,D-Trp7,9]SP-(5-11) inhibited the neurogenic contraction of isolated guinea-pig hilus bronchi more readily than a contraction produced by exogenous SP. Furthermore, it has previously been shown that a tachykinin antagonist given intrathecally produced motor blockade as do local anaesthetic drugs. We therefore examined whether tachykinin antagonists had a depressant action on axonal neurotransmission. The compound action potential (APc) of the frog isolated sciatic nerve was suppressed in a concentration-dependent manner by the tachykinin antagonists [D-Pro2,D-Trp7,9]SP and [Arg5,D-Trp7,9]Sp-(5-11), both being about 4 times more potent than lidocaine. SP itself was without effect. Similarly in the rat isolated sciatic nerve [D-Pro2,D-Trp7,9]SP suppressed the APc. It was more potent in the A alpha- than in the C-fibres. SP did not affect conduction in either fibre type. In conscious guinea-pigs [D-Pro2,D-Trp7,9]SP injected adjacent to the sciatic nerve was found to block motor but not sensory functions of the limb. Thus, commonly used tachykinin antagonists, but not SP itself, have potent local anaesthetic properties. This should be considered when these agents are employed as pharmacological tools.

Action Potentials↗
PubMed1985

Genetic modifications of voltage-sensitive sodium channels in Drosophila: gene dosage studies of the seizure locus.

We have identified a genetic locus in Drosophila melanogaster whose product appears to have a structural role in the formation of functional voltage-sensitive sodium channels. This locus, designated seizure, is defined by two temperature-sensitive alleles (seits-1 and seits-2), each of which causes convulsive seizures followed by a rapid but reversible paralysis of adults at restrictive temperatures above 38 degrees C. Previous work had shown that seits-2 extracts display an altered pH dependence and an abnormally high Kd for [3H]-saxitoxin binding at high temperatures, suggesting that sodium channels in seits-2 mutants have an altered structure (Jackson F. R., S. D. Wilson, G. R. Strichartz, and L. M. Hall (1984) Nature 308: 189-191). These binding studies have now been extended to extracts of seits-1 which have a Kd not significantly different from wild-type at all assay temperatures. However, seits-1 extracts show a reduced number of saxitoxin binding sites (Bmax) relative to wild-type. This reduction is only 5 to 18% at 0 degree C but is 17 to 37% at 39 degrees C, suggesting that under certain conditions sodium channels in the seits-1 mutant are more labile than those of wild-type. Cytogenetic studies demonstrate that the seizure locus maps within region 60A to 60B8-10 on the second chromosome. Gene dosage analysis of approximately 99.7% of the genome, including this second chromosome region, failed to detect a wild-type locus whose dose affected saxitoxin-binding activity. Nevertheless, the mutant seits-2 allele has codominant and dose-dependent effects on paralytic behavior and saxitoxin-binding activity.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1984

Components of the plasma membrane of growing axons. III. Saxitoxin binding to sodium channels.

The density of sodium channels was measured in growing and mature axons of the olfactory nerve of the bullfrog, using as a probe the drug saxitoxin (STX). The toxin binds to control nerves from adult animals in a saturable manner with a dissociation constant of approximately 23 nM at 4 degrees C and a capacity of 72 fmol/mg wet weight, equivalent to about five sites per square micrometer of axolemma. In growing nerves, obtained from adult frogs 4-5 wk following removal of the original nerve, the STX-binding capacity per wet weight of tissue is markedly reduced, to approximately 25% of control values, and appears to decrease in the proximodistal direction. STX-binding data, expressed as STX/mg wet weight, was converted to STX/micron 2 of axolemma using stereologically derived values of membrane area per milligram wet weight of nerve. The axolemmal content (area/mg wet weight) of all regions of growing nerve is substantially decreased compared to controls, but increases in the proximodistal direction by 60%. These changes in axolemmal area result in calculated STX receptor densities (per unit axolemmal area) which, in distal regions, are approximately at the level of the mature nerve and, in proximal regions, are actually increased above controls by 50 to 70%. Upon comparing the axolemmal density of intramembrane particles, reported in the companion paper, with the calculated density of STX receptors in both mature and growing nerves, we find a correlation between STX receptors and intramembrane particles with diameters of 11.5-14.0 nm. The growing axon's gradient of sodium channels and the shift from this gradient to a uniform distribution in the mature axon suggest (a) that sodium channels are inserted into the perikaryal plasmalemma and diffuse from there into the growing axolemma, and (b) that the axolemma undergoes functional maturation during growth.

Amphibian Proteins↗
PubMed1983

Phenol solutions differentially block conduction in cutaneous nerve fibers of the cat.

The action of phenol in Tyrode solution on nerve conduction has been investigated in vivo in the desheathed sural nerve of cats. A section of 8 mm of the nerve was exposed to phenol solution. At low phenol concentrations (0.05-0.125%) a reversible block of A- and C-fibers occurred. At higher concentrations (0.6-1%) a selective and persistent C-fiber block could be achieved. The size of the residual A-volley was between 50% and 81% when all C-fibers were blocked.

Animals↗
PubMed1983

Purification and physiological characterization of neurotoxins from venoms of the scorpions centruroides sculpturatus and leiurus quinquestriatus.

Several different toxins having specific effects on the kinetics of sodium channels have been isolated from the venoms of two scorpions. A combination of two steps of ion-exchange chromatography has been used to purify these toxins, whose sizes and purities have been assayed by gel filtration, urea/sodium dodecyl sulfate/polyacrylamide gel electrophoresis, and isoelectric focusing. The actions of the toxins and their relative potencies have been determined by studying the modifications they produce in action potential shape, using the sucrose-gap method, and in ionic current kinetics, measured under voltage-clamp, both assays performed on myelinated axons of frogs and toads. The venom of Leiurus quinquestriatus scorpions yielded two active neurotoxins; the major neurotoxin has a mass of approximately 7000 daltons. This major toxin affected the sodium channel inactivation process exclusively, slowing the rates of inactivation as well as preventing complete inactivation from occurring in some of the channels. Such slowed and incomplete sodium inactivation resulted in action potentials that were prolonged, from their usual duration of 5-8 msec to hundreds of milliseconds or even seconds. Five toxins were isolated from the venom of Centruroides sculpturatus Ewing scorpions, all of which also had masses of approximately 7000-7500 daltons. Four of these toxins acted primarily on the activation process of sodium channels, producing a novel increase in sodium permeability upon repolarization of the nerve membrane following a depolarizing pulse, as previously described for the crude venom [Cahalan, M. D. J. Physiol. (Lond.) 244:511-534 (1975)]. These toxins also caused repetitive firing of action potentials in single axons in response to one stimulating pulse, as well as spontaneous impulse firing. A fifth neurotoxin from C. sculpturatus venom had effects similar to those of the L. quinquestriatus toxins, slowing and preventing complete sodium inactivation. The effect of this toxin was slowly removed during external perfusion by Ringer's solution.

Action Potentials↗
PubMed1981

Saxitoxin binding in nerves from walking legs of the lobster Homarus americanus. Two classes of receptors.

The binding of exchange-labeled saxitoxin (STX) to sodium channels has been investigated in the nonmyelinated fibers of the walking leg nerves of the lobster. The properties of the STX binding site differed systematically among the nerves from different walking legs. The equilibrium dissociation constant for STX binding (KSTX) to the front legs is approximately twice that for the binding to the rear legs; the average ratio of KSTX (front): KSTX (rear) from five separate experiments was 1.80 +/- 0.21 (mean +/- SE). The actual KSTX values ranged from 124.0 to 22.7 nM for the front leg nerves and from 8.6 to 12.7 nM for the rear leg nerves. KSTX values for the middle two walking leg nerves fell between those for the front and rear legs. The inhibitory dissociation constant for tetrodotoxin (KTTX), calculated from tetrodotoxin's inhibition of labeled STX binding, was 3.02 +/- 0.27 nM for the front legs and 2.20 +/- 0.33 nM for the rear legs. The ratio KSTX: KTTX was different in the front and rear leg nerves, being 5.5 and 4.2, respectively. The apparent P pKa of the STX receptor also differed between the two legs, being 4.6 +/- 0.3 for the front legs and 5.1 +/- 0.1 for the rear legs. These results demonstrate that one tissue type in one organism can contain different toxin binding sites. The difference in the receptors can be qualitatively accounted for by the location of an additional negative charge near the receptor site of the rear walking leg.

Animals↗
PubMed1981

Current concepts of the mechanism of action of local anesthetics.

Recent experiments are reviewed to present a current view of the mechanisms of conduction block by local anesthetics. Local anesthetics block the sodium channels whose opening causes the rising phase of the action potential. Both charged and neutral forms of local anesthetics are able to block channels. Charged anesthetics interfere with a "gating" mechanism after gaining access to a "receptor" site in the aqueous pore of the channel from the axoplasmic surface of the membrane. This site is only available to charged compounds when the gate of the channel is open. In contrast, uncharged compounds (including the free base form of local anesthetics) appear to reach the site through the membrane's lipid interior, bypassing the channel "gates." Anesthetics blocking the gate of the channel can either enhance or inhibit the normal inactivation mechanism of the sodium channel, depending on the particular anesthetic.

Action Potentials↗