Purification and amino acid sequence of a highly insecticidal toxin from the venom of the Brazilian spider Phoneutria nigriventer which inhibits NMDA-evoked currents in rat hippocampal neurones

Figueiredo, Suely G.; Lima, M.E. de; Cordeiro, Marta N.; Diniz, Carlos R.; Patten, Debra; Halliwell, Robert F.; Gilroy, John; Richardson, Michael

doi:10.1016/s0041-0101(00)00129-x

Cited by 72 publications

(43 citation statements)

References 30 publications

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“…Most of the toxins that have been purified from this venom seem to act on ionic channels, including those for sodium (Araujo et al, 1993;De Lima et al, 2007;Martin-Moutot et al, 2006;Matavel et al, 2002), calcium (Dos Santos et al, 2002;Guatimosim et al, 1997;Leão et al, 2000;Vieira et al, 2005) and potassium (Kushmerick et al, 1999). Some of these toxins have insecticidal activity (for a review see De Lima et al, 2007), interfer with the uptake of L-glutamate on synaptosomes of rat brain (Mafra et al, 1999) and inhibit NMDA-evoked currents in rat hippocampal neurons (Figueiredo et al, 2001). A semi-purified toxic fraction called PhTx2 releases acetylcholine from rat cerebrocortical synaptosomes (Moura et al, 1998).…”

Section: Introductionmentioning

confidence: 99%

Tx2-6 toxin of the Phoneutria nigriventer spider potentiates rat erectile function

Nunes

Costa‐Goncalves

Lanza

et al. 2008

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The venom of the spider Phoneutria nigriventer contains several toxins that have bioactivity in mammals and insects. Accidents involving humans are characterized by various symptoms including penile erection. Here we investigated the action of Tx2-6, a toxin purified from the P. nigriventer spider venom that causes priapism in rats and mice. Erectile function was evaluated through changes in intracavernosal pressure/mean arterial pressure ratio (ICP/MAP) during electrical stimulation of the major pelvic ganglion (MPG) of normotensive and deoxycorticosterone-acetate (DOCA)-salt hypertensive rats. Nitric oxide (NO) release was detected in cavernosum slices with fluorescent dye (DAF-FM) and confocal microscopy. The effect of Tx2-6 was also characterized after intracavernosal injection of a non-selective nitric oxide synthase (NOS) inhibitor, L-NAME. Subcutaneous or intravenous injection of Tx2-6 potentiated the elevation of ICP/MAP induced by ganglionic stimulation. L-NAME inhibited penile erection and treatment with Tx2-6 was unable to reverse this inhibition. Tx2-6 treatment induced a significant increase of NO release in cavernosum tissue. Attenuated erectile function of DOCA-salt hypertensive rats was fully restored after toxin injection. Tx2-6 enhanced erectile function in normotensive and DOCA-salt hypertensive rats, via the NO pathway. Our studies suggest that Tx2-6 could be important for development of new pharmacological agents for treatment of erectile dysfunction.

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Section: Introductionmentioning

confidence: 99%

Tx2-6 toxin of the Phoneutria nigriventer spider potentiates rat erectile function

Nunes

Costa‐Goncalves

Lanza

et al. 2008

Toxicon

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“…Various neurotoxins active on Na + (Cordeiro et al, 1992;Matavel et al, 2002), K + Carneiro et al, 2003) and Ca 2+ (Diniz et al, 1990Santos et al, 2002, Gomez et al, 2002 ion channels, and other highly insecticidal proteins (Figueiredo et al, 1995(Figueiredo et al, , 2001Mafra et al, 1999;De Lima et al, 2002;Oliveira et al, 2003) have been described from the venom of this spider, which is commonly found in Central and South-East Brazil, but very little is known about the venoms of the other species of this genus. Recently, our group has published the comparison of the partial proteomes of venoms from different species of the genus Phoneutria (Richardson et al, 2006).…”

Section: Introductionmentioning

confidence: 99%

Leftward Shift in the Voltage-Dependence for Ca2+ Currents Activation Induced by a New Toxin from Phoneutria reidyi (Aranae, Ctenidae) Venom

et al. 2006

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Various neurotoxins have been described from the venom of the Brazilian spider Phoneutria nigriventer, but little is known about the venoms of the other species of this genus. In the present work, we describe the purification and some structural and pharmacological features of a new toxin (PRTx3-7) from Phoneutria reidyi that causes flaccid paralysis in mice. The observed molecular mass (4627.26 Da) was in accordance with the calculated mass for the amidated form of the amino acid sequence (4627.08 Da). The presence of an alpha-amidated C-terminus was confirmed by MS/MS analysis of the C-terminal peptide, isolated after enzymatic digestion of the native protein with Glu-C endoproteinase. The purified protein was injected (intracerebro-ventricular) into mice at dose levels of 5 microg/mouse causing immediate agitation and clockwise gyration, followed by the gradual development of general flaccid paralysis. PRTx3-7 at 1 microM inhibited by 20% the KCl-induced increase on [Ca2+]i in rat brain synaptosomes. The HEK cells permanently expressing L, N, P/Q and R HVA Ca2+ channels were also used to better characterize the pharmacological features of PRTx3-7. To our surprise, PRTx3-7 shifted the voltage-dependence for activation towards hyperpolarized membrane potentials for L (-4 mV), P/Q (-8 mV) and R (-5 mV) type Ca2+ currents. In addition, the new toxin also affected the steady state of inactivation of L-, N- and P/Q-type Ca2+ currents.

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“…Despite these significant but modest actions on I Glu-Cl , it is unlikely that an enhancement of an inhibitory response (Cleland, 1996;de Figueiredo et al, 2001) is responsible for the potent excitatory neurotoxicity caused by κ-HXTX-Hv1c. Furthermore, a significant enhancement of I Glu-Cl amplitude (p < 0.05) was only evident at a concentration of 1 µM, which seems unlikely to be effective enough to correlate with the potent toxicity seen in insect bioassays (Gunning et al, 2008;Maggio and King, 2002b;Tedford et al, 2007;Wang et al, 2000).…”

Section: Gaba and Glutamate Gated Chloride Channels Are Not Targeted mentioning

confidence: 99%

Lethal effects of an insecticidal spider venom peptide involve positive allosteric modulation of insect nicotinic acetylcholine receptors

et al. 2017

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Please cite this article as: Windley, M.J., Vetter, I., Lewis, R.J., Nicholson, G.M., Lethal effects of an insecticidal spider venom peptide involve positive allosteric modulation of insect nicotinic acetylcholine receptors, Neuropharmacology (2017), doi: 10.1016/j.neuropharm.2017.04.008. This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. Highlights• The previously characterized BK Ca channel block is not responsible for the lethal excitatory toxicity of κ-HXTX-Hv1c • κ-HXTX-Hv1c is a positive allosteric modulator of the insect nicotinic acetylcholine receptor which acts by prolonging current decay and reversing receptor desensitization • The likely lethal target of κ-HXTX-Hv1c is the nicotinic acetylcholine receptor, acting by a mechanism similar to that of the insecticide spinosyn A. A B S T R A C Tκ-Hexatoxins (κ-HXTXs) are a family of excitotoxic insect-selective neurotoxins from Australian funnel-web spiders that are lethal to a wide range of insects, but display no toxicity towards vertebrates. The prototypic κ-HXTX-Hv1c selectively blocks native and expressed cockroach largeconductance calcium-activated potassium (BK Ca or K Ca 1.1) channels, but not their mammalian orthologs. Despite this potent and selective action on insect K Ca 1.1 channels, we found that the classical K Ca 1.1 blockers paxilline, charybdotoxin and iberiotoxin, which all block insect K Ca 1.1 channels, are not lethal in crickets. We therefore used whole-cell patch-clamp analysis of cockroach dorsal unpaired median (DUM) neurons to study the effects of κ-HXTX-Hv1c on sodium-activated (K Na ), delayed-rectifier (K DR ) and 'A-type' transient (K A ) K + channels. 1 µM κ-HXTX-Hv1c failed to significantly inhibit cockroach K Na and K DR channels, but did cause a 30 ± 7% saturating inhibition of K A channel currents, possibly via a Kv4 (Shal-like) action. However, this modest action at such a high concentration of κ-HXTX-Hv1c would indicate a different lethal target. Accordingly, we assessed the actions of κ-HXTX-Hv1c on neurotransmitter-gated ion channels in cockroach DUM neurons. We found that κ-HXTX-Hv1c failed to produce any major effects on GABA A or glutamate-Cl receptors but dramatically slowed nicotine-evoked ACh receptor (nAChR) current decay and reversed nAChR desensitization. These actions occurred without any alterations to nAChR current amplitude or the nicotine concentration-response curve, and are consistent with a positive allosteric modulation of nAChRs. κ-HXTX-Hv1c therefore represents the first venom peptide that selectively modulates insect nAChRs with a mode of action similar to the excitotoxic insecticide spino...

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Purification and amino acid sequence of a highly insecticidal toxin from the venom of the Brazilian spider Phoneutria nigriventer which inhibits NMDA-evoked currents in rat hippocampal neurones

Cited by 72 publications

References 30 publications

Tx2-6 toxin of the Phoneutria nigriventer spider potentiates rat erectile function

Tx2-6 toxin of the Phoneutria nigriventer spider potentiates rat erectile function

Leftward Shift in the Voltage-Dependence for Ca2+ Currents Activation Induced by a New Toxin from Phoneutria reidyi (Aranae, Ctenidae) Venom

Lethal effects of an insecticidal spider venom peptide involve positive allosteric modulation of insect nicotinic acetylcholine receptors

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