Biomedical subjects

D M Soderlund

Publications and source records attributed to D M Soderlund.

At least 19 recordsLinked to original sources

PubMed2003

The molecular biology of knockdown resistance to pyrethroid insecticides.

The term "knockdown resistance" is used to describe cases of resistance to diphenylethane (e.g. DDT) and pyrethroid insecticides in insects and other arthropods that result from reduced sensitivity of the nervous system. Knockdown resistance, first identified and characterized in the house fly (Musca domestica) in the 1950's, remains a threat to the continued usefulness of pyrethroids in the control of many pest species. Research since 1990 has provided a wealth of new information on the molecular basis of knockdown resistance. This paper reviews these recent developments with emphasis on the results of genetic linkage analyses, the identification of gene mutations associated with knockdown resistance, and the functional characterization of resistance-associated mutations. Results of these studies identify voltage-sensitive sodium channel genes orthologous to the para gene of Drosophila melanogaster as the site of multiple knockdown resistance mutations and define the molecular mechanisms by which these mutations cause pyrethroid resistance. These results also provide new insight into the mechanisms by which pyrethroids modify the function of voltage-sensitive sodium channels.

Amino Acid Substitution↗

PubMed2001

The V410M mutation associated with pyrethroid resistance in Heliothis virescens reduces the pyrethroid sensitivity of house fly sodium channels expressed in Xenopus oocytes.

Some strains of Heliothis virescens carry a novel sodium channel mutation, corresponding to the replacement of Val410 by Met (designated V410M) in the house fly Vssc1 sodium channel, that is genetically and physiologically associated with pyrethroid resistance. To test the functional significance of this mutation, we created a house fly Vssc1 sodium channel containing the V410M mutation by site-directed mutagenesis, expressed wildtype and specifically mutated sodium channels in Xenopus laevis oocytes, and evaluated the effects of the V410M mutation on the functional and pharmacological properties of the expressed channels by two-electrode voltage clamp. The V410M mutation caused depolarizing shifts of approximately 9mV and approximately 5mV in the voltage dependence of activation and steady-state inactivation, respectively, of Vssc1 sodium channels. The V410M mutation also reduced the sensitivity of Vssc1 sodium channels to the pyrethroid cismethrin at least 10-fold and accelerated the decay of cismethrin-induced sodium tail currents. The degree of resistance conferred by the V410M mutation in the present study is sufficient to account for the degree of pyrethroid resistance in H. virescens that is associated with this mutation. Although Val410 is located in a sodium channel segment identified as part of the binding site for batrachotoxin, the V410M mutation did not alter the sensitivity of house fly sodium channels to batrachotoxin. The effects of the V410M mutation on the voltage dependence and cismethrin sensitivity of Vssc1 sodium channels were indistinguishable from those caused by another sodium channel point mutation, replacement of Leu1014 by Phe (L1014F), that is the cause of knockdown resistance to pyrethroids in the house fly. The positions of the V410M and L1014F mutations in models of the tertiary structure of sodium channels suggest that the pyrethroid binding site on the sodium channel alpha subunit is located at the interface between sodium channel domains I and II.

Animals↗

PubMed2000

Cloning and functional characterization of a putative sodium channel auxiliary subunit gene from the house fly (Musca domestica).

The functional expression of cloned Drosophila melanogaster and house fly (Musca domestica) voltage-sensitive sodium channels in Xenopus oocytes is enhanced, and the inactivation kinetics of the expressed channels are accelerated, by coexpression with the tipE protein, a putative sodium channel auxiliary subunit encoded by the tipE gene of D. melanogaster. These results predict the existence of a tipE ortholog in the house fly. Using a PCR-based homology probing approach, we isolated cDNA clones encoding an ortholog of tipE (designated Vssc beta) from adult house fly heads. Clones comprising 3444 bp of cDNA sequence contained a 1317 bp open-reading frame encoding a 438 amino acid protein. The predicted Vssc beta protein exhibited 72% amino acid sequence identity to the entire D. melanogaster tipE protein sequence and 97% identity within the two hydrophobic segments identified as probable transmembrane domains. Coexpression of Vssc beta with the house fly sodium channel alpha subunit (Vssc1) in oocytes enhanced the level of sodium current expression five-fold and accelerated the rate of sodium current inactivation 2.2-fold. Both of these effects were significantly larger in magnitude than the corresponding effects of the D. melanogaster tipE protein on the expression and kinetics of Vssc1 sodium channels. These results identify a second example of a putative sodium channel auxiliary subunit from an insect having functional but not structural homology to vertebrate sodium channel beta subunits.

Amino Acid Sequence↗

PubMed1999

Mutations in the house fly Vssc1 sodium channel gene associated with super-kdr resistance abolish the pyrethroid sensitivity of Vssc1/tipE sodium channels expressed in Xenopus oocytes.

The super-kdr insecticide resistance trait of the house fly confers resistance to pyrethroids and DDT by reducing the sensitivity of the fly nervous system. The super-kdr genetic locus is tightly linked to the Vssc1 gene, which encodes a voltage-sensitive sodium channel alpha subunit that is the principal site of pyrethroid action. DNA sequence analysis of Vssc1 alleles from several independent super-kdr fly strains identified two amino acid substitutions associated with the super-kdr trait: replacement of leucine at position 1014 with phenylalanine (L1014F), which has been shown to cause the kdr resistance trait in this species, and replacement of methionine at position 918 with threonine (M918T). We examined the functional significance of these mutations by expressing house fly sodium channels containing them in Xenopus laevis oocytes and by characterizing the biophysical properties and pyrethroid sensitivities of the expressed channels using two-electrode voltage clamp. House fly sodium channels that were specifically modified by site-directed mutagenesis to contain the M918T/L1014F double mutation gave reduced levels of sodium current expression in oocytes but otherwise exhibited functional properties similar to those of wildtype channels and channels containing the L1014F substitution. However, M918T/L1014F channels were completely insensitive to high concentrations of the pyrethroids cismethrin and cypermethrin. House fly sodium channels specifically modified to contain the M918T single mutation, which is not known to exist in nature except in association with the L1014F mutation, gave very small sodium currents in oocytes. Assays of these currents in the presence of high concentrations of cismethrin suggest that this mutation alone is sufficient to abolish the pyrethroid sensitivity of house fly sodium channels. These results define the functional significance of the Vssc1 mutations associated with the super-kdr trait of the house fly and are consistent with the hypothesis that the super-kdr trait arose by selection of a second-site mutation (M918T) that confers to flies possessing it even greater resistance than the kdr allele containing the L1014F mutation.

Animals↗

PubMed1998

Actions of the pyrethroid insecticides cismethrin and cypermethrin on house fly Vssc1 sodium channels expressed in Xenopus oocytes.

Voltage-sensitive sodium channels encoded by the Vssc1 gene of the house fly (Musca domestica) were expressed in Xenopus laevis oocytes in combination with the tipE gene product of Drosophila melanogaster and were characterized by two-electrode voltage clamp. Vssc1/tipE sodium channels expressed in oocytes were highly sensitive to tetrodotoxin; half-maximal inhibition of sodium currents by tetrodotoxin was obtained at a concentration of 2.4 nM. Cismethrin, a pyrethroid that produces Type I effects on intact nerve, slowed the inactivation of sodium currents carried by Vssc1/tipE channels during a depolarizing pulse and induced a tail current after repolarization that decayed with a first-order time constant of approximately 650 ms. The voltage dependence of activation and steady-state inactivation of cismethrin-modified channels were shifted to more negative potentials. Cypermethrin, a pyrethroid with Type II effects on intact nerve, also prolonged the inactivation of Vssc1/tipE sodium channels and induced a tail current. However, the cypermethrin-induced tail current was extremely persistent, decaying with a first-order time constant of approximately 42 s. Unlike cismethrin, the effect of cypermethrin was use dependent, requiring repeated depolarizing pulses for the full development of modified sodium currents. The divergent effects of cismethrin and cypermethrin on Vssc1/tipE sodium channels expressed in oocytes are consistent with the actions of these and related compounds on sodium channels in invertebrate and vertebrate nerve preparations and provide insight into the mechanisms underlying the production of Type I and II effects on neuronal excitability.

Animals↗

PubMed1998

Action of the pyrethroid insecticide cypermethrin on rat brain IIa sodium channels expressed in xenopus oocytes.

Pyrethroid insecticides bind to a unique site on voltage-dependent sodium channels and prolong sodium currents, leading to repetitive bursts of action potentials or use-dependent nerve block. To further characterize the site and mode of action of pyrethroids on sodium channels, we injected synthetic mRNA encoding the rat brain IIa sodium channel alpha subunit, either alone or in combination with synthetic mRNA encoding the rat sodium channel beta1 subunit, into oocytes of the frog Xenopus laevis and assessed the actions of the pyrethroid insecticide [1R,cis,alphaS]-cypermethrin on expressed sodium currents by two-electrode voltage clamp. In oocytes expressing only the rat brain IIa alpha subunit, cypermethrin produced a slowly-decaying sodium tail current following a depolarizing pulse. In parallel experiments using oocytes expressing the rat brain IIa alpha subunit in combination with the rat beta1 subunit, cypermethrin produced qualitatively similar tail currents following a depolarizing pulse and also induced a sustained component of the sodium current measured during a step depolarization of the oocyte membrane. The voltage dependence of activation and steady-state inactivation of the cypermethrin-dependent sustained current were identical to those of the peak transient sodium current measured in the absence of cypermethrin. Concentration-response curves obtained using normalized tail current amplitude as an index of the extent of sodium channel modification by cypermethrin revealed that coexpression of the rat brain IIa alpha subunit with the rat beta1 subunit increased the apparent affinity of the sodium channel binding site for cypermethrin by more than 20-fold. These results confirm that the pyrethroid binding site is intrinsic to the sodium channel alpha subunit and demonstrate that coexpression of the rat brain IIa alpha subunit with the rat beta1 subunit alters the apparent affinity of this site for pyrethroids.

Action Potentials↗

PubMed1997

The L1014F point mutation in the house fly Vssc1 sodium channel confers knockdown resistance to pyrethroids.

Voltage-sensitive sodium channels encoded by a full-length cDNA corresponding to the Vssc1 gene of the house fly (Musca domestica) were expressed in Xenopus laevis oocytes either alone or in combination with the tipE gene product of Drosophila melanogaster and were characterized by two-electrode voltage clamp. Vssc1 cRNA alone produced very small (50-150 nA) sodium currents, whereas the combination of Vssc1 and tipE cRNAs produced robust (0.5-3 microA), rapidly inactivating sodium currents. The pyrethroid insecticide cismethrin prolonged the sodium current carried by Vssc1/tipE sodium channels during a depolarizing pulse and induced a tail current after repolarization. The Vssc1 cDNA was specifically mutated to substitute phenylalanine for leucine at position 1014 of the inferred amino acid sequence (L1014F), a polymorphism shown previously to be associated with the kdr (knockdown resistance) trait of the house fly. The L1014F substitution reduced the sensitivity of expressed house fly sodium channels to cismethrin at least 10-fold and increased the rate of decay of pyrethroid-induced sodium tail currents. These results demonstrate that the resistance-associated L1014F mutation confers a reduction in the sensitivity of house fly sodium channels to pyrethroids that is sufficient to account for the kdr resistance trait.

Animals↗

PubMed1996

Characterization of voltage-sensitive sodium channel gene coding sequences from insecticide-susceptible and knockdown-resistant house fly strains.

The kdr insecticide resistance trait of the house fly (Musca domestica .L.), which confers reduced neuronal sensitivity to DDT and pyrethroid insecticides, was previously shown to exhibit tight genetic linkage to restriction fragment length polymorphism markers lying within a voltage-sensitive sodium channel gene that is homologous to the para gene of Drosophila melanogaster. In the present study, the 6315 nucleotide coding sequences of this voltage-sensitive sodium channel gene from insecticide-susceptible (NAIDM strain) and kdr (538ge strain) house flies were determined by automated direct DNA sequencing of PCR fragments obtained by amplification on first strand cDNA from adult heads. The deduced 2105-residue amino acid sequence from each strain exhibited overall structure and organization typical of sodium channel alpha subunit genes and was 90.0% identical to that of the D. melanogaster para gene product. We did not detect any splice variants among voltage-sensitive sodium channel cDNAs obtained from adult house fly head preparations. Comparison of the coding sequence of the voltage-sensitive sodium channel gene of the kdr house fly strain to that of the NAIDM strain revealed 12 amino acid differences in the 538ge strain. The significance of these polymorphisms as candidate resistance-conferring mutations is discussed.

Amino Acid Sequence↗

PubMed1994

Tight genetic linkage between the kdr insecticide resistance trait and a voltage-sensitive sodium channel gene in the house fly.

The kdr insecticide resistance trait in the house fly, Musca domestica, confers resistance to the rapid paralysis (knockdown) and lethal effects of 1,1,1-trichloro-2,2-bis(p-chlorophenyl)ethane (DDT) and pyrethroids. Flies with the kdr trait exhibit reduced neuronal sensitivity to these compounds, which are known to act at voltage-sensitive sodium channels of nerve membranes. To test the hypothesis that a mutation in a voltage-sensitive sodium channel gene confers the kdr phenotype, we have cloned genomic DNA corresponding to a segment of the house fly homologue of the para sodium channel gene of Drosophila melanogaster, identified restriction-site polymorphisms within this segment between the kdr strain 538ge and an inbred insecticide-susceptible lab stain, and developed a sensitive polymerase chain reaction-based diagnostic procedure to determine the sodium channel genotype of individual flies. A genetic linkage analysis performed with these molecular markers shows that the kdr trait is tightly linked (within about 1 map unit) to the voltage-sensitive sodium channel gene segment exhibiting the DNA sequence polymorphism. These findings provide genetic evidence for a mutation at or near a voltage-sensitive sodium channel gene as the basis for kdr resistance.

Amino Acid Sequence↗

PubMed1994

PCR-based homology probing reveals a family of GABA receptor-like genes in Drosophila melanogaster.

A polymerase chain reaction (PCR)-based homology probing strategy was employed to screen Drosophila melanogaster genomic DNA for sequences encoding a conserved amino acid 'signature motif' known to be present in vertebrate GABA receptor and glycine receptor subunit genes. This approach yielded three discrete amplified sequence elements (designated LCCH1, LCCH2, and LCCH3) that contained open reading frames and > 40% amino acid sequence identity to the corresponding regions of vertebrate ligand-gated chloride channel genes. Genomic DNA clones corresponding to each element were isolated and sequenced, and predicted amino acid sequences corresponding to the second (M2) and third (M3) transmembrane domains of vertebrate genes were analyzed for identity or similarity to known sequences. LCCH1 was identical to the Rdl gene, a known GABA receptor subunit gene from D. melanogaster, whereas LCCH2 and LCCH3 were novel D. melanogaster sequences that exhibited structural similarity to other members of the ligand-gated chloride channel gene family. LCCH2 was equally divergent in M2 and M3 (46-49% amino acid identity) from all other known members of this family and may therefore represent a new subunit or receptor class within this family. LCCH2 was localized by in situ hybridization to cytogenetic region 75A on the left arm of chromosome 3. LCCH3 was closely related to mammalian (79% amino acid identity) and snail (96% amino acid identity) GABA receptor beta subunits and may therefore be the homologue in D. melanogaster of this subunit class. LCCH3 was localized by in situ hybridization to cytogenetic region 13F on the X chromosome.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗

PubMed1993

Characterization of a putative gamma-aminobutyric acid (GABA) receptor beta subunit gene from Drosophila melanogaster.

A cDNA encoding a novel member of the ligand-gated chloride channel gene family of insects has been isolated from Drosophila melanogaster and characterized. The 1488 nucleotide open reading frame of this cDNA encodes an amino acid sequence having structural features conserved among ligand-gated chloride channel subunit proteins, including four hydrophobic domains capable of forming transmembrane helices (M1-M4), an octapeptide "signature motif" occurring in M2, a large intracellular domain between M3 and M4, and cysteine residues postulated to form a disulfide-bridged loop structure in the extracellular domain. Among characterized members of this gene family, this gene exhibits the greatest similarity to GABA receptor beta subunit genes. This high level of structural similarity suggests that its encoded protein may be a functional homologue of the beta subunit family in the D. melanogaster nervous system.

Amino Acid Sequence↗

PubMed1993

Binding of [3H]batrachotoxinin A-20-alpha-benzoate and [3H]saxitoxin to receptor sites associated with sodium channels in trout brain synaptoneurosomes.

1. [3H]Batrachotoxinin A-20-alpha-benzoate ([3H]BTX-B) and [3H]saxitoxin ([3H]STX), radioligands that bind to distinct sites on the voltage-sensitive sodium channel, were bound specifically to saturable sites in rainbow trout (Oncorhynchus mykiss) brain synaptoneurosomes. 2. Specific [3H]BTX-B binding was temperature dependent with highest levels of specific [3H]BTX-B binding observed at 7 degrees C. Specific binding was inversely correlated with assay temperature at temperatures above 7 degrees C. 3. Saturating concentrations of scorpion (Leiurus quinquestriatus) venom (ScV) stimulated specific [3H]BTX-B binding at 27 degrees C, but not at 7 degrees C. The dihydropyrazole insecticide RH 3421 inhibited specific [3H]BTX-B binding at 7 degrees C but had no effect on specific binding at 27 degrees C. The sodium channel activators veratridine and aconitine and the local anesthetic dibucaine inhibited specific [3H]BTX-B binding at both 7 degrees C and 27 degrees C. 4. Displacement experiments in the presence of ScV at 27 degrees C gave an equilibrium dissociation constant (KD) for [3H]BTX-B of 710 nM and a maximal binding capacity (Bmax) of 11.3 pmol/mg protein. Kinetic experiments established the rates of association (1.17 x 10(5) min-1 nM-1) and dissociation (0.0514 min-1) of the ligand-receptor complex. 5. The binding of [3H]STX reached apparent saturation at 7.5 nM. Scatchard analysis of the saturation data indicated a KD of 3.8 nM and a Bmax of 1.9 pmol/mg protein. 6. These studies provide evidence for high affinity, saturable binding sites for [3H]BTX-B and [3H]STX in trout brain preparations. Whereas certain neurotoxins modified the specific binding of [3H]BTX-B in trout brain synaptoneurosomes in a predictable fashion, other compounds known to affect specific [3H]BTX-B binding in mammalian brain preparations had no effect on specific [3H]BTX-B binding in the trout.

Animals↗

PubMed1992

Mechanisms of action of ibogaine and harmaline congeners based on radioligand binding studies.

Assays using radioligands were used to assess the actions of ibogaine and harmaline on various receptor types. Ibogaine congeners showed affinity for opiate receptors whereas harmaline and harmine did not. The Ki for coronaridine was 2.0 microM at mu-opiate receptors. The Kis for coronaridine and tabernanthine at the delta-opiate receptors were 8.1 and 3.1 microM, respectively. Ibogaine, ibogamine, coronaridine and tabernanthine had Ki values of 2.08, 2.6, 4.3 and 0.15 microM, respectively, for kappa-opiate receptors. Long-lasting, dose-dependent behavioral effects of ibogaine have been reported. The possibility that these effects were due to irreversible binding properties of ibogaine at kappa-receptors was considered; however, radioligand wash experiments showed a rapid recovery of radioligand binding after one wash. A voltage-dependent sodium channel radioligand demonstrated Ki values in the microM range for all drugs tested. Using radioligand binding assays and/or 36Cl- uptake studies, no interaction of ibogaine or harmaline with the GABA receptor-ionophore was found. The kappa-activity of ibogaine (or an active metabolite) may be responsible for its putative anti-addictive properties whereas the tremorigenic properties of ibogaine and harmaline may be due to their effects on sodium channels.

Animals↗

PubMed1991

PCR-generated conspecific sodium channel gene probe for the house fly.

A segment of the house fly (Musca domestica) homologue of the para (paralytic) sodium channel gene of Drosophila melanogaster was isolated by using mixed sequence oligonucleotide primers in the polymerase chain reaction (PCR). The specificity of the procedure was demonstrated by genomic Southern analysis using the housefly PCR amplification product as a probe and by DNA sequence analysis. The latter showed structural homology to the para gene, but not to the corresponding region of DSC1, another D. melanogaster gene with structural similarity to vertebrate sodium channel genes.

Amino Acid Sequence↗

PubMed1991

Activation of gamma-aminobutyric acid insensitive chloride channels in mouse brain synaptic vesicles by avermectin B1a.

The interaction of avermectin B1a (AVMB1a) with mouse brain chloride channels was characterized using a radiochloride efflux assay. The loss of intravesicular chloride from synaptoneurosomes preloaded with 36Cl involved an initial rapid phase followed by a slower phase that approached equilibrium within 10 min. AVMB1a stimulated a 30% loss of intravesicular chloride within the first 2 s of exposure; however, AVMB1a had no effect on the rate of the slower phase of chloride loss. Experiments with lysed synaptoneurosomes showed that both chloride loading and basal and AVMB1a-stimulated chloride release required the presence of intact vesicles. The efflux of 36Cl from mouse brain synaptosomes and the stimulation of efflux by AVMB1a were qualitatively similar to the results obtained with synaptoneurosomes but involved much lower overall levels of chloride loading and release. AVMB1a produced half-maximal stimulation of chloride efflux from synaptoneurosomes at a concentration of 2.1 +/- 0.3 microM and a 35.4 +/- 1.4% maximal loss of intravesicular chloride at saturating concentrations. gamma-Aminobutyric acid (GABA), bicuculline, or the chloride channel blockers picrotoxinin, t-butylbicyclophosphorothionate (TBPS) 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid (DIDS), and anthracene 9-carboxylic acid (9-CA) had little or no effect on the loss of chloride from synaptoneurosomes either in the presence or the absence of AVMB1a. However, the chlorinated cycloalkane insecticides dieldrin and lindane were equally effective as inhibitors of GABA-dependent chloride uptake and AVMB1a-stimulated chloride efflux. These data demonstrate that AVMB1a-stimulated chloride efflux from mouse brain synaptic vesicles results from the activation of GABA-insensitive chloride channels and that this action is distinct from their previously documented effects on GABA-gated chloride channels in mouse brain preparations. Our findings imply that both GABA-gated and GABA-insensitive chloride channels may be toxicologically significant targets for the action of avermectins.

Animals↗

PubMed1991

Prolonged exposure to GABA activates GABA-gated chloride channels in the presence of channel-blocking convulsants.

1. In assays of 36Cl- uptake into mouse brain vesicles, 100 microM GABA markedly increased both the initial rate of 36Cl- uptake and the total amount of chloride taken up over a 120-sec incubation period. Specific GABA-dependent 36Cl- uptake (the difference between total and background uptake) was essentially complete within 15 sec of incubation. 2. Incubation with GABA following preincubation with 10 microM endrin, a polychlorocycloalkane insecticide and established blocker of GABA-gated chloride channels, showed a stimulation of uptake over background levels that was much slower in onset than that observed with GABA alone but nevertheless achieved virtually the same level of stimulation above background levels after 90 sec of incubation with GABA. 3. In electrophysiological assays of GABA receptors expressed in Xenopus oocytes following injection with rat brain mRNA, endrin (20 microM) effectively blocked the transient currents elicited by brief exposure of oocytes to GABA (200 microM). However, prolonged exposure to GABA in the absence of perfusion produced a large, slowly-developing inward current. 4. The actions of several known GABA antagonists were also compared as inhibitors of GABA-dependent 36Cl- uptake into mouse brain vesicles at short (4 sec) and long (120 sec) incubation times using concentrations of inhibitors known to produce approximately 70-90% inhibition of GABA-dependent chloride uptake in 4-sec incubations. Picrotoxinin and TBPS, like endrin, were completely ineffective as inhibitors in 120-sec incubations. In contrast, bicuculline was almost as effective at 120 sec as at 4 sec, and avermectin Bla produced approximately 50% inhibition of the GABA response after 120 sec.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1989

Binding of [3H]batrachotoxinin A-20-alpha-benzoate to a high affinity site associated with house fly head membranes.

1. [3H]Batrachotoxinin A-20-alpha-benzoate (BTX-B), a radioligand that labels the alkaloid activator recognition site of the voltage-sensitive sodium channel, was bound specifically to high affinity, saturable sites in a subcellular preparation from house fly (Musca domestica L.) heads that was shown previously to contain binding sites for other sodium channel-directed ligands. 2. Specific binding of [3H]BTX-B was observed in the presence of 140 mM sodium or potassium and was inhibited by choline ion. 3. Saturating concentrations of scorpion (Leiurus quinquestriatus) venom stimulated the specific binding of [3H]BTX-B four-fold, increasing the proportion of specific binding of 10 nM [3H]BTX-B from less than 15% to 40%. Equilibrium dissociation studies in the presence of scorpion venom gave an equilibrium dissociation constant (KD) for [3H]BTX-B of 80 nM and a maximal binding capacity (Bmax) of 1.5 pmol/mg protein. 4. Parallel experiments in the absence of venom gave a KD value of 140 nM and a Bmax of 1.3 pmol/mg protein, indicating that scorpion venom stimulated [3H]BTX-B binding by increasing the affinity of this site approximately two-fold. 5. The specific binding of [3H]BTX-B was inhibited by the sodium channel activators aconitine and batrachotoxin and, to a lesser extent, by the anticonvulsant diphenylhydantoin. However, several other sodium channel-directed neurotoxins known to exert allosteric effects on the binding of [3H]BTX-B to mammalian brain preparations did not affect the binding of [3H]BTX-B to house fly head membranes.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1989

Neurotoxic actions of pyrethroid insecticides.

Pyrethroid insecticides interact with a variety of neurochemical processes, but not all of these actions are likely to be involved in the disruption of nerve function. Several lines of evidence suggest that the voltage-sensitive sodium channel is the single principal molecular target site for all pyrethroids and DDT analogs in both insects and mammals. The alterations of sodium channel functions identified in both biophysical and biochemical studies are directly related to the effects of these compounds on intact nerves. The pyrethroid recognition site of the sodium channel exhibits the stringent stereospecificity predicted by in vivo estimates of intrinsic neurotoxicity in both insects and mammals. Type I and Type II compounds produce qualitatively different effects on sodium channel tail currents, divergent actions on intact nerves, and different effects on the excitability of vertebrate skeletal muscle. Moreover, compounds that are defined as intermediate in the Type I/Type II classification scheme are also intermediate in their effects on sodium channel kinetics. The range of different actions on sensory and motor nerve pathways arising from these qualitatively different effects at the level of the sodium channel appear to be sufficient to explain the distinct poisoning syndromes that have been identified in both insects and mammals. Thus, it does not appear necessary to invoke different primary target sites for Type I and Type II compounds to explain their actions in whole animals. Although the voltage-sensitive sodium channel is likely to be the principal site of pyrethroid action, it is probably not the only site involved in intoxication. Insect neurosecretory neurons are sensitive to very low concentrations of pyrethroids, and disruption of the neuroendocrine system has been implicated as a factor contributing to the irreversible effects of pyrethroid intoxication in insects. Since action potentials in these nerves are carried by calcium ions through TTX-insensitive voltage-gated cation channels, these findings provide evidence that pyrethroids can alter neuronal excitability through an action on voltage-sensitive channels other than the sodium channel. Actions on voltage-sensitive calcium channels may also be involved in the effects of pyrethroids on neurotransmitter release in mammals. The proconvulsant actions of pyrethroids mediated through the peripheral-type benzodiazepine receptor may also contribute to pyrethroid intoxication. Both Type I and Type II compounds are potent proconvulsants in vivo at doses well below those required to produce pyrethroid-dependent intoxication.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗