Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Charybdotoxin”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 37 records · Page 2Linked to original sources
PubMed1996

Synthesis and structural characterisation of analogues of the potassium channel blocker charybdotoxin.

Charybdotoxin is a 37-residue polypeptide toxin from scorpion venom, which acts by blocking voltage-gated and Ca(2+)-activated K+ channels. We have synthesized charybdotoxin and three mono-substituted analogues using an Fmoc-tBu protocol. The Phe-2 --> Tyr analogues was chosen to introduce a site for Tyr iodination which was distinct from the K+ channel binding surface, while the Glu-12 --> Gln and Arg-19 --> His analogues were studied to probe the roles of charged residues at these positions in the structure and activity of the toxin. The synthetic native molecule was equipped with natural toxin in inhibiting the human erythrocyte Ca(2+)-dependent K+ channel. The affinities of all three analogues for the erythrocyte K+ channel were slightly reduced, with the Arg-19 --> His analogue showing the greatest increase in IC50 (2.30-fold). Two-dimensional 1H-NMR studies of these analogues showed that the Glu-12 to Gln substitution, which appeared to destabilise the N-terminal half of the alpha-helix, possibly due to the weakening of an N-terminal helix capping interaction which is apparent from our NMR data. His-21 has a pKa more than one unit below the value for a non-interacting histidine. Possible reasons for this are that the imidazolium side chain is partly buried and is located near positively charged moieties. Thus, His-21 would be neutral at physiological pH, where charybdotoxin binds to the potassium channel.

Amino Acid Sequence↗
PubMed1992

Charybdotoxin, dendrotoxin and mast cell degranulating peptide block the voltage-activated K+ current of fibroblast cells stably transfected with NGK1 (Kv1.2) K+ channel complementary DNA.

The blocking actions of the K+ channel toxins charybdotoxin, dendrotoxin and mast cell degranulating peptide were studied in B82 mouse fibroblast cells transformed to express NGK1 (Kv1.2) K+ channels. All three toxins were potent blockers of the K+ current in these cells, with KD values of 1.7, 2.8 and 185 nM, respectively. The toxin block exhibited a weak voltage-dependence with the degree of inhibition decreasing at positive membrane potentials. For charybdotoxin and dendrotoxin, reducing [K+]i did not increase the fractional block, demonstrating that the relief of block at positive membrane potentials is not due to displacement of the toxin molecules by outward flow of K+ ions. A voltage-jump protocol was used to determine the rates of binding and unbinding of dendrotoxin and mast cell degranulating peptide; binding of charybdotoxin was too rapid to be quantitatively evaluated in this manner. The binding rates (dendrotoxin, approximately 5 x 10(7)/M per s; mast cell degranulating peptide, approximately 0.8 x 10(7)/M per s) were largely voltage-independent, suggesting that association of the toxin molecules with the channel is diffusion limited. The rates of unbinding (dendrotoxin, approximately 0.3/s; mast cell degranulating peptide, approximately 3/s at +60 mV) of both toxins increased e-fold per approximately 40 mV change in membrane potential, thus accounting for the voltage-dependence of the equilibrium block. Internal perfusion with the three toxins failed to affect the K+ current (in contrast to internal tetraethylammonium which strongly blocked the current), indicating that the toxins exert their blocking action by binding to extracellular sites.

Ampicillin Resistance↗
PubMed1993

Comparison of the distribution of binding sites for the potassium channel ligands [125I]apamin, [125I]charybdotoxin and [125I]iodoglyburide in the rat brain.

Potassium channels represent a diverse and promising target for drug development. Pharmacological subtypes of K channels have begun to emerge based on the development of both organic molecules and peptide toxins which possess subtype selectivity. In order to evaluate the neuroanatomical distribution of these subtypes we have utilized the ligands [125I]apamin, [125I]charybdotoxin and [125I]iodoglyburide in an autoradiographic study of rat brain. In the rat brain, these ligands have selectivity for the low conductance Ca(2+)-activated, voltage-gated K channels and ATP-sensitive K channels respectively. The distribution of binding sites for these three ligands were distinctly different. [125I]Apamin binding was highest in various thalamic and hippocampal structures, while only low to moderate levels of [125I]charybdotoxin binding were seen in these regions. In contrast, very high levels of [125I]charbydotoxin were seen in white matter regions such as the lateral olfactory tract and fasciculus retroflexus. High levels of [125I]charybdotoxin binding were also seen in gray matter-containing regions such as the zona incerta, medial geniculate and superior colliculus, where low to moderate [125I]apamin binding was found. [125I]Iodoglyburide presented a more uniform binding with the highest levels in the globus pallidus, islands of Calleja, anteroventral nucleus of the thalamus and zonas reticulata of the substantia nigra. These results indicate that subtypes of K channels have very different distributions in the brain. As such, the results imply differing CNS actions for potential modulators of K channel subtypes.

Animals↗
PubMed1997

Cat carotid body chemosensory discharge (in vitro) is insensitive to charybdotoxin.

Charybdotoxin (ChTX), a venom protein, suppresses Ca2+-activated K+ (K+(Ca)) currents in the glomus cell of neonatal rat carotid body. If it works similarly for cat carotid body chemoreceptors, charybdotoxin is expected to stimulate the chemosensory discharge during normoxia, and particularly hypoxia and hypercapnia. We studied the effects of charybdotoxin (20-40 nM) in vitro (perfused/superfused) on the cat carotid chemosensory discharge, and simultaneously tissue PO2 (PtiO2), as a measure of positive control. ChTX (20 nM) only increased PtiO2 and decreased carotid chemosensory discharge during hypoxia, indicating vasodilation. We conclude that K+(Ca) channels do not appear to play a significant role in chemotransduction in the cat carotid body.

Animals↗
PubMed1996

Effects of charybdotoxin on K+ channel (KV1.2) deactivation and inactivation kinetics.

Of particular interest for voltage-gated K+ channels are the effects of membrane voltage and pharmacologic agents on channel kinetics. We have characterized in detail properties of Kv1.2 channel expressed in oocytes as the basis for investigation of its structure-function relationships. This channel exhibited a voltage-dependent rate of activation with a V1/2 of -21 mV. Voltage-dependent steady-state inactivation overlapped the activation curve with half-maximal inactivation occurring at -22 mV. Dendrotoxin inhibited channel activation with an IC50 of 8.6 nM at + 35 mV. Charybdotoxin also blocked this K+ channel (IC50 = 5.6 nM). While dendrotoxin block was not affected by channel activation, charybdotoxin exhibited additional accumulation of block following activation, which was relieved with a time constant of 0.5 s upon repolarization of the membrane. The deactivation of this channel was accelerated in the presence of charybdotoxin while not significantly affected by dendrotoxin.

Animals↗
PubMed1993

Evaluation of the relaxant effects of SCA40, a novel charybdotoxin-sensitive potassium channel opener, in guinea-pig isolated trachealis.

1. Experiments have been performed in order to analyse the mechanism whereby SCA40, a new imidazo[1,2-a]pyrazine derivative relaxes airway smooth muscle. 2. SCA40 (0.01-10 microM) caused a complete and concentration-dependent relaxation of guinea-pig isolated trachea contracted with 20 mM KCl but failed to inhibit completely the spasmogenic effects of 80 mM KCl. 3. Quinine (30 microM) antagonized the relaxant activity of SCA40 in 20 mM KCl-contracted guinea-pig isolated trachea. The ATP-sensitive K(+)-channel blocker, glibenclamide (3 microM), did not antagonize the relaxant activity of SCA40 in either 20 mM KCl or 1 microM carbachol-contracted isolated trachea. 4. SCA40 (0.01-10 microM) and isoprenaline (0.1 nM-10 microM) caused a complete and concentration-dependent relaxation of guinea-pig isolated trachea contracted with carbachol 1 microM. 5. The large-conductance Ca(2+)-activated K(+)-channel blocker, charybdotoxin (60-180 nM), non-competitively antagonized the relaxant activity of isoprenaline on 1 microM carbachol-contracted trachea. The inhibition was characterized by rightward shifts of the isoprenaline concentration-relaxation curves with depression of their maxima. 6. The relaxant activity of SCA40 in 1 microM carbachol-contracted trachea was antagonized by charybdotoxin (60-600 nM) in an apparently competitive manner. The concentration-relaxation curves to SCA40 were shifted to the right with no significant alteration in the maximum response. 7. It is concluded that SCA40 is a novel potassium channel opener which is a potent relaxant of guinea-pig airway smooth muscle in vitro. The relaxant activity of SCA40 does not involve ATP-sensitive K+-channels but rather large-conductance Ca2'-activated K+-channels or other charybdotoxin sensitive K+-channels.

Animals↗
PubMed1990

Basolateral K+ channels in airway epithelia. I. Regulation by Ca2+ and block by charybdotoxin.

In airway epithelia, adenosine 3',5'-cyclic monophosphate (cAMP) stimulates Cl- secretion by activating apical membrane Cl- channels and basolateral membrane K+ channels. Cl- channels are regulated by cAMP-dependent phosphorylation, whereas K+ channels are regulated by the cytosolic Ca2+ concentration, [Ca2+]c. Our recent observation that cAMP increases [Ca2+]c suggested that cAMP might indirectly regulate K+ channels by increasing [Ca2+]c. To study regulation of K+ channels we measured 86Rb efflux, single K+ channels in membrane patches, and [Ca2+]c with the fluorescent indicator fura-2. Isoproterenol and Ca2+ ionophore, A23187, transiently increased [Ca2+]c and transiently stimulated 86Rb efflux. Stimulation of 86Rb efflux resulted from release of intracellular Ca2+ stores. 86Rb efflux was blocked by Ba2+ or charybdotoxin, but not by tetraethylammonium. Charybdotoxin prevented all of the 86Rb efflux that was stimulated by A23187 or by forskolin. Charybdotoxin also blocked the low-conductance inwardly rectifying K+ channel (KCLIC) in membrane patches. These results indicate that the KCLIC channel is responsible for the Ca2(+)-dependent increase in K+ permeability in airway epithelial cells. They also indicate that cAMP-induced release of intracellular Ca2+ is sufficient to activate K+ channels.

Animals↗
PubMed1993

[Effects of toxins, apamine, charybdotoxin and iberiotoxin on the relaxation of the smooth muscular fiber induced by imidazo(1,2-a)pyrazine derivative].

Experiments have been performed in order to analyse the mechanism whereby SCA40, a new imidazo[1,2-a]pyrazine derivative relaxes airway smooth muscle. We investigated the effect of different toxins, known to be K(+)-channel blockers on guinea-pig smooth muscle relaxant activity of SCA40. The small conductance Ca(2+)-activated K(+)-channel blocker apamin (100 nM) did not antagonize the relaxant activity of SCA40 in 1 microM carbachol-contracted isolated guinea pig trachea. The large conductance Ca(2+)-activated K(+)-channel blocker, iberiotoxin (30, 60 and 180 nM) antagonized the relaxant activity of SCA40 in an apparently competitive manner. The concentration-relaxation curves to SCA40 were shifted to the right with no significant alteration in the maximum response. The relaxant activity of SCA40 in 1 microM carbachol-contracted isolated trachea was antagonized by both charybdotoxin (60 nM) and iberiotoxin (60 nM), but the antagonism induced by iberiotoxin appears to be more potent than that induced by charybdotoxin. It is concluded that the potent relaxant activity of SCA40 on guinea-pig airway smooth muscle in vitro involves a charybdotoxin and iberiotoxin sensitive K(+)-channel.

Animals↗
PubMed1997

Extracellular ATP and UTP control the generation of reactive oxygen intermediates in human macrophages through the opening of a charybdotoxin-sensitive Ca2+-dependent K+ channel.

Human monocyte-derived macrophages possess a NADPH oxidase that catalyzes superoxide formation upon phagocytosis. Extracellular ATP per se does not activate NADPH oxidase but potentiates superoxide generation triggered by opsonized zymosan. UTP can substitute for ATP with the same efficiency, suggesting that ATP mediates its effects specifically through P2U receptors. Extracellular UTP stimulates a rapid increase in cytoplasmic Ca2+ concentration in monocytic cells, which results from a release of intracellular Ca2+ stores. Moreover, UTP-induced calcium increase is sufficient to activate a charybdotoxin-sensitive Ca2+-dependent outward K+ channel (K(Ca)). The activity of this channel develops between 0.1 and 1.0 microM free cytoplasmic Ca2+ concentration; it is half-blocked by 10 nM charybdotoxin but insensitive to iberiotoxin. Under asymmetrical K+ conditions, this K(Ca) channel does not depend on membrane potential and is characterized by a linear single-current voltage relationship in the voltage range of -100 to +50 mV, giving a unitary conductance of 10 pico-Siemens. Interestingly, ATP/UTP-induced oxygen radicals release was inhibited by charybdotoxin in the same range of concentration as the UTP-induced K(Ca) channel. Furthermore, we show that ATP or UTP fail to enhance oxygen radicals production before K(Ca) channel is expressed (3 days). The electrogenic nature of the NADPH oxidase, i.e., its level of activation, being dependent on the plasmic membrane potential, might provide the causal link between the reactive oxygen intermediates generation and the opening of the K(Ca) channel.

Adenosine Triphosphate↗
PubMed1992

A small-conductance charybdotoxin-sensitive, apamin-resistant Ca(2+)-activated K+ channel in aortic smooth muscle cells (A7r5 line and primary culture).

A small conductance K+ channel was identified in smooth muscle cells of the rat aortic cell line A7r5 and also in rat aortic smooth muscle cells in primary culture, using conventional single-channel recording techniques. The single-channel conductance shows no rectification, either in the range -70 to +40 mV under asymmetrical conditions (9.1 pS), or in the range -100 to +50 mV in symmetrical 150 mM K+ (37 pS). Channel activity is reversibly inhibited by extracellular application of charybdotoxin, with a concentration of 8 nM producing half-maximal inhibition. It is unaffected by apamin or scyllatoxin. Channel activity depends on the presence of free Ca2+ on the cytosolic face of the membrane, with an activation zone between 0.1 and 1 microM. This small-conductance, charybdotoxin-sensitive, Ca(2+)-regulated K+ channel is activated by vasoconstrictors such as vasopressin and endothelin.

Animals↗
PubMed1994

Charybdotoxin and iberiotoxin but not apamin abolish the slow after-hyperpolarization in myenteric plexus neurons.

Myenteric neurons of guinea-pig ileum were studied with intracellular microelectrodes. The specific toxins charybdotoxin, iberiotoxin and apamin were used to characterize the prolonged after-hyperpolarizations of AH neurons in this preparation. Charybdotoxin and iberiotoxin blocked prolonged after-hyperpolarizations in 23 of 24 AH neurons, but apamin had no effect on 5 of 5 AH neurons. Abolition of the after-hyperpolarizations was accompanied by depolarization and increases in input resistances of those AH neurons affected, but the shapes of action potentials were unchanged. The excitability of the AH neurons was enhanced as shown by an increase in the number of action potentials evoked by a 500-ms depolarizing current pulse or by a train of 15-ms depolarizing current pulses (10Hz). The other class of myenteric neurons, S neurons, was also investigated. The 19 S neurons studied fired action potentials only at the start of a 500 ms depolarization, but the toxins had no effect on this behaviour or on their other properties. Intracellular injection of Neurobiotin into the neurons studied and subsequent immunohistochemical staining to localise the calcium-binding protein, calretinin, indicated that all major classes of S neurons were included in the sample. Thus, the prolonged after-hyperpolarizations in AH neurons may be due to opening of a large-conductance (BK) calcium-dependent potassium channel, but similar channels play little or no role in regulation of the excitability of S neurons.

Animals↗
PubMed1989

Interactions between dendrotoxin, a blocker of voltage-dependent potassium channels, and charybdotoxin, a blocker of calcium-activated potassium channels, at binding sites on neuronal membranes.

Dendrotoxin I (DpI) from black mamba venom (Dendroaspis polylepis) has high affinity binding sites on rat brain synaptic membranes. Native DpI displaced [125I]-DpI binding with a Ki of 1 x 10(-10) M, and over 90% of specific binding was displaceable. Charybdotoxin isolated from the Israeli scorpion venom (Leiurus quinquestriatus hebraeus), also displaced [125I]-DpI binding, with a Ki of approximately 3 x 10(-9) M, although the displacement curve was shallower than with native DpI. Both toxins are thought to be high affinity blockers of specific K+ currents. Charybdotoxin selectively blocks some types of Ca2+-activated K+ channels, whereas dendrotoxins only block certain voltage-dependent K+ channels. The interaction between the two types of toxin at the DpI binding site is unexpected and may suggest the presence of related binding sites on different K+ channel proteins.

Animals↗
PubMed1992

Effect of charybdotoxin and leiurotoxin I on potassium currents in bullfrog sympathetic ganglion and hippocampal neurons.

The effects of charybdotoxin and leiurotoxin I were examined on several classes of K+ currents in bullfrog sympathetic ganglion and hippocampal CA1 pyramidal neurons. Highly purified preparations of charybdotoxin selectively blocked a large voltage- and Ca(2+)-dependent K+ current (IC) responsible for action potential repolarization (IC50 = 6 nM) while leiurotoxin I selectively blocked a small Ca(2+)-dependent K+ conductance (IAHP) responsible for the slow afterhyperpolarization following an action potential (IC50 = 7.5 nM) in bullfrog sympathetic ganglion neurons. Neither of the toxins had significant effects on other K+ currents (M-current [IM], A-current [IA] and the delayed rectifier [IK]) present in these cells. Leiurotoxin I at a concentration of 20 nM had no detectable effect on currents in hippocampal CA1 pyramidal neurons. This lack of effect on IAHP in central neurons suggests that the channels underlying slow AHPs in those neurons are pharmacologically distinct from analogous channels in peripheral neurons.

Animals↗
PubMed1989

Charybdotoxin blocks dendrotoxin-sensitive voltage-activated K+ channels.

Charybdotoxin, a short scorpion venom neurotoxin, which was thought to be specific for the blockade of Ca2+-activated K+ channels also blocks a class of voltage-sensitive K+ channels that are known to be the target of other peptide neurotoxins from snake and bee venoms such as dendrotoxin and MCD peptide. Charybdotoxin also inhibits 125I-dendrotoxin and 125I-MCD peptide binding to their receptors. All these effects are observed with an IC50 of about 30 nM.

Animals↗
PubMed1989

Dendrotoxin and charybdotoxin increase the cytosolic concentration of free Ca2+ in cerebrocortical synaptosomes: an effect not shared by apamin.

Nanomolar concentrations of charybdotoxin or dendrotoxin increase the cytoplasmic free Ca2+ concentration in isolated central nerve terminals. The effects of the two toxins, normally considered to be blockers of K+ channels controlled by voltage in a Ca2+-sensitive or -insensitive manner, respectively, show only marginal additivity. Apamin, and inhibitor of low conductance Ca2+-activated K+ channels, was without effect in either the absence or presence of dendrotoxin. The effect of charybdotoxin on polarized, isolated central nerve terminals seems to be mediated largely by a block of K+ channels sensitive to dendrotoxin. Apparently, these voltage-operated K+ channels make a more significant contribution to maintaining the polarized potential of synaptosomes than do those activated by Ca2+.

Animals↗
PubMed1994

Functional role of charybdotoxin-sensitive K+ channels in the resting state of dog basilar artery.

To determine the possible role of Ca(2+)-activated K+ (KCa) channels in the regulation of resting tone of cerebral arteries, the effect of agents which interact with these channels on tension and 86Rb efflux was examined in endothelium-denuded strips of dog basilar artery, and data were compared with findings in the mesenteric artery. The basilar artery maintained a myogenic tone; that is, the resting tone decreased when nifedipine was added. The addition of charybdotoxin, a blocker of large conductance KCa channels, caused a concentration-dependent contraction in the basilar artery but not in the mesenteric artery. In strips preloaded with 86Rb, the basal 86Rb efflux rate constant was significantly greater in the basilar artery than in the mesenteric artery. Nifedipine decreased the basal 86Rb efflux rate constant only in the basilar artery. The basal 45Ca influx in the resting state of the basilar artery was significantly increased when compared with the mesenteric artery and this increase in the basilar artery was reduced by nifedipine. The results suggest that the charybdotoxin-sensitive KCa channels regulate the myogenic tone in the resting state of the basilar artery. The activation of KCa channels in the basilar artery appears to be secondary to the increased transmembrane Ca2+ influx probably via the activation of L-type voltage-dependent Ca2+ channels in the resting state of this artery.

Animals↗
PubMed1988

Presence of a charybdotoxin sensitive Ca2+-activated K+ channel in rat glioma C6 cells.

A study was made of the 86Rb+ influx and efflux through Ca2+-activated K+ channels of intact rat glioma C6 cells after addition of a Ca2+ ionophore to the incubation medium. Half-maximal activation of the channels was obtained at a cytoplasmic Ca2+ concentration of approximately 400 nM. The 86Rb+ ion flux through the Ca2+-activated K+ channels was insensitive to apamin, but was inhibited by low concentrations of charybdotoxin (IC50 = 1.6 nM). This is the first evidence for the presence of charybdotoxin-sensitive Ca2+-activated K+ channels in glial cells.

Animals↗
PubMed1996

Presence of the voltage-gated potassium channels sensitive to charybdotoxin in inhibitory presynaptic terminals of cultured rat hippocampal neurons.

To determine whether the charybdotoxin-sensitive subtypes of voltage-gated K+ channels (Kv1.2 and Kv1.3) exist in inhibitory pre-synaptic terminals, effects of K+ channel blockers including TEA, charybdotoxin (ChTX), iberiotoxin (IbTX), kaliotoxin (KTX) and margatoxin (MgTX) on the inhibitory transmission were examined with cultured rat hippocampal neurons. Monosynaptic inhibitory postsynaptic currents (IPSCs) evoked by electrical stimulation of single presynaptic neurons were recorded from the whole-cell clamped postsynaptic neurons. In the presence of TEA, application of ChTX greatly increased the amplitude of IPSCs. A specific maxi-K+ channel blocker IbTX failed to augment IPSCs. KTX and MgTX, both of which block Kv1.3 but not Kv1.2, mimicked the facilitating effect of ChTX. In the absence of TEA, application of ChTX increased the IPSC amplitude significantly, while IbTX was without effect. These results indicate that the ChTX-sensitive subtypes of voltage-gated K+ channels, most likely Kv1.3, contribute to the repolarization of action potentials at presynaptic terminals of hippocampal inhibitory neurons, and that the ChTX-induced facilitation of the transmission can be explained by its effects on the Kv channels rather than maxi-K+ channels.

Animals↗