Pest Control with Enhanced Environmental Safety - American

Chapter 16

Arthropod-Derived Neurotoxic Insecticides A Lead in Pesticide Science? Downloaded by NORTH CAROLINA STATE UNIV on October 26, 2012 | http://pubs.acs.org Publication Date: March 12, 1993 | doi: 10.1021/bk-1993-0524.ch016

T. Piek Department of Pharmacology, University of Amsterdam, Meibergdreef 15, 1105 AZ Amsterdam, Netherlands

Many hymenopteran insects incapacitate their prey by means of a neurotoxin. The glutamatergic excitatory neuromuscular transmission and the nicotinic excitatory synaptic transmission of insects is reversibly blocked by philanthotoxins, as well as by structurally related polyamine toxins isolated from spider venoms. These polyamine toxins are cation channel blockers or change at least the glutamatergic channel kinetics. They therefore are not receptor specific and are, for example, also active at the nicotinic receptors in the insect brain, albeit at higher concentrations. Poneratoxin, a 25 amino acid residue polypeptide isolated from an ant venom, is the first described hymenopteran neurotoxin affecting excitability of nerve and muscle fibres by changing the kinetics of the voltage-dependent sodium channel. The nicotinic synaptic transmission in the insect central nervous system (CNS) is presynaptically and irreversibly blocked by kinins, which probably cause transmitter depletion. Besides this delayed effect the carbohydrate-containing vespulakinins also show a direct and reversible inhibition of the synaptic transmission. Criticism on the possibilities to use venom toxins as leads in pesticide science has been, that these compounds have to be injected into the body or the CNS. For primary synthesis products the incorporation of the genetic codes into entomophilic viruses could solve this problem. Several groups of hymenopteran insects, like wasps and ants, sting their prey to a paralysis or only to a behaviourial inactivation, in order to offer the prey to the own offspring as an incapacitated, but living fresh source of food. These insects have developed venoms containing natural insecticides. The ideas of the 1980 s included the possibility of using natural neurotoxins from arthropods as leads to new pesticides (4, 41-43). f

0097-6156/93/0524-0233S06.00/0 © 1993 American Chemical Society In Pest Control with Enhanced Environmental Safety; Duke, S., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1993.

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PEST CONTROL WITH ENHANCED ENVIRONMENTAL SAFETY

The Reversible Non-competitive Block of Synaptic Transmission by Polyamine Toxins

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f

In the 1960 s investigations started in our laboratories on the paralysing venom of the European solitary sphecid wasp Philanthus triangulum. The excitatory glutamatergic neuromuscular transmission of insects is antagonized by the venom and by its most active toxin (philanthotoxin 4.3.3, Figure 1) through two different effects. Firstly by a presynaptic inhibition of the high affinity glutamate uptake (1-4) and secondly by a block of open ion channels of the glutamate receptorionophore complex (4,5). The same toxin was isolated from the venom of an African subspecies of this wasp: P.t. abdelcader (7). Structure-activity relationship studies showed that the potency as a channel blocker of philanthotoxin depends on the presence of an aromatic moiety (8-11). Attempts to change the molecule into a more potent antagonist resulted in an increase in potency with factors varying from 10-100 (8, 10-13, 22). The pre- and postsynaptic inhibiting properties of the natural philanthotoxin were dissociated into two different analogues (14). Comparable polyamine-like structures have been isolated from spider venoms (15-17). These polyamine toxins are also active at the mammalian CNS glutamate synaptic transmission (18-20) and the vertebrate and insect nicotinic transmission (21-23). The most active philanthotoxin analogue in the mammalian CNS is dideaza-philanthotoxin-12 (19, 20). Irreversible Depletion of Cholinergic Nerve Endings by Kinins

In 1954 Schachter and colleagues discovered a pain-producing substance in the venom of the social wasp Paravespula vulgaris (24). Due to the similarity in pharmacological properties of this substance and the bradykinin (Figure 2) they called this substance wasp kinin (25). The first wasp kinin, which was chemically characterized is polisteskinin 3, an octapeptide isolated from a mixture of Polistes species (26). During the following twenty years a number of kinins were isolated from social vespid wasps, like hornet, (Vespa), yellow jackets (Paravespula), and paper wasps (Polistes) (27), from solitary wasps Megascolia, Colpa (28-30) Tiphia and Dasymutilla

(31), and from several ant species (31).

They were pharmacologically characterized (31) and in some cases the amino acid sequence was determined (27, 30, 33). It was not before the discovery of threonine-6-bradykinin (Thr^BK) in the venom of Megascolia flavifrons (28, 29) that kinins were considered to be neurotoxins. Scoliid wasps sting into the ganglia of their prey, resulting in an irreversible paralysis. Therefore, the action of the venom and of T h r B K was studied using the technique of microperfusion of the sixth abdominal ganglion of the cockroach, Periplaneta americana (28). A t concentrations from 10 M to 10~^M Thr^BK causes an irreversible block of synaptic transmission from the cereal nerve XI to a giant interneuron. The velocity of inhibition is use (frequency) dependent like the antagonism of hemicholinium-3 (32). 6

In Pest Control with Enhanced Environmental Safety; Duke, S., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1993.

16. PIEK

235

Arthropod-Derived Neurotoxic Insecticides f

During the 1960 s Pisano and colleagues isolated, from the venom of the most offensive wasp in the United States: Paravespula maculifrons a heptadecapeptide called vespulakinin 1. This kinin is highly basic and contains the nonapeptide bradykinin at the carboxyterminus (Figure 2). Most interesting is the presence of carbohydrate moieties connected to threonine residues (33). Synthesis of some mono- and di-glycosylated analogues of the polypeptide called "carbohydrate free" vespulakinin 1 (Figure 3) were described, (34-36). Recently the effects of these synthetic vespulakinin analogues on the synaptic transmission of the insect CNS were studied. Two distinctly different effects were found: a direct reversible block of excitatory nicotinic transmission and a delayed irreversible block of transmission as has been described earlier for threonine-6bradykinin (32).


f

Poneratoxin Induces an Interconversion between Two Gating Modes of Sodium Channels

Evenomation by the ponerine ant Paraponera clavata causes pain and uncontrollable trembling (37). Recently, three neurotoxin principles, which block the synaptic transmission from the cereal excitatory nerve to a giant interneuron in the sixth abdominal ganglion of the cockroach have been separated from this venom (38,39). One of the principles was pharmacologically characterized as a kinin. Another and also the most active neurotoxic fraction was rechromatographed, resulting in the purification of a peptide of 25 amino acid residues, called poneratoxin (PoTX, Figure 4). A t concentrations varying from 10"**M to 10~^M synthetic PoTX is a strong, but very slowly acting agonist of smooth muscles, it also blocks synaptic transmission in the insect CNS in a concentration dependent manner and it depolarizes giant interneurons (40). PoTX induces a concentration dependent (10~^M to 5 x 10"^M) prolongation of action potentials and at saturating concentration, a slow repetitive activity, developing at negative potentials (40). PoTX specifically acts on voltage-dependent sodium-ion channels by decreasing the peak sodium current ( I ) and by simultaneously inducing a slow I which starts to activate at -85 mV and inactivates very slowly. Both the fast and the slow components of I are suppressed by tetrodotoxin and reverse at equal potentials, corresponding to the equilibrium potential for sodium ions. The fast component of I has a voltage-dependence, activation and steady-state inactivation almost similar to those of the control I . The voltage dependence of the slow I is 40 mV more negative than that of the fast one. These results suggest that PoTX affects all the sodium channels and that the fast and slow I -components originate from a possible PoTX-induced interconversion between a fast and a slow operating mode of the sodium channel (40). Na

N a

N a

N a

N a

N a

Na

Discussion

The present paper describes two types of arthropod neurotoxins selected by nature to incapacitate insects and spiders. The first group consists of low molecular weight neurotoxins that contain a polyamine part. These toxins are highly specific for modifying the kinetics of cation channels coupled to excitatory receptors like glutamate or acetylcholine receptors. Their action is In Pest Control with Enhanced Environmental Safety; Duke, S., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1993.

236


c=o I NH

HO—(C))-- CH CHCNH(CH ) NH(CH ) NH(CH ) NH I 2

2

4

2

3

2

3

2


O Figure 1. Structure of 6-philanthotoxin (philanthotoxin-4.3.3) isolated from the wasp Philanthus triangulum.

Carbohydrate

(T)

Carbohydrate @

Ti Thr-Ala-Thr-Thr-Arg-Arg-Arg-Gly-Bradykinin

Carbohydrate 1 : N;Ac.Galactosamine 1-2, Galactose 1 Carbohydrate 2 : N.Ac.Galactosamine 2-3, Galactose 2 Bradykinin i s Arg-Pro-Pro-Gly-Phe-Ser-Pro-Phe-Arg Figure 2 . Structure of vespulakinin 1 (33). A

Carbohydrate

B

(Gala)Thr

C

(Gala)Thr ,

D

3

3

(Galfl)Thr

3

free

peptide

(VSK 1)

- VSK 1 (Gala)Thr

4

- VSK 1

- VSK 1

Figure 3- Synthetic vespulakinin-1-analogues (34, 35).

P i n e — Leu—Pro—Leu-Leu

i Leu— Ser—Gly — L e v a . —He

i Leu-Met-Thr-Pro-Pro

I H e — A X « a — G i n - l i e - V a X

I His-Asp-Ala-Gln-Arg

I Figure 4- Amino acid sequence of poneratoxin (PoTX), a neurotoxin isolated from the ant Paraponera clavata. In Pest Control with Enhanced Environmental Safety; Duke, S., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1993.


16. PIEK

Arthropod-Derived Neurotoxic Insecticides

237

non-competitive and reversible. Attempts to change the molecules into more potent antagonists resulted in an increase in potency with factors varying from only 10 to 100 (8, 10-12). If no further increase in potency and/or decrease in the reversibility can be obtained, these interesting class of neurotoxins may be only important as tools for the study of insect neurophysiology. The second group of medium-sized low molecular weight arthropod neurotoxins consists of polypeptides. Examples of such neurotoxins are the (wasp) kinins (27-29) and the ant venom.component poneratoxin (38-40). It is conceivable that a large number of potent polypeptide neurotoxins could be isolated from venoms of insectivore ants. Especially in case of irreversible or slowly reversible action of primary synthesis products, as has been demonstrated for kinins, a pesticide action could be obtained by means of genetic manipulation. Moreover, it might be possible to design a mimic pharmacophore (loop) of a peptide (44). Literature Cited

1. 2. 3. 4.

5. 6. 7. 8. 9.

10. 11. 12.

13. 14. 15. 16. 17.

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23. Rozenthal,R.;Scoble,G.T.;Albuquerque,E.X.;Idriss,M.;Sherby,S.;Sattelle,D.B.;

Nakanishi,K.;Konno,K.;Eldefrawi,A.T.;Eldefrawi,M.E.J.Pharm.Exp. Therapeut.1989,249,123-130. 24. Jaques,R.;Schachter,M.Brit.J.Pharmacol.1954,9,53-57. 25. Schachter,M.;Thain,E.M.Brit.J.PAarmacol.1954,9,352-359. 26. Udenfriend,S.;Nakajima,T.;Pisano,J.J.Proc.Int.Congr.Biochem.7th VIII4,1967,501. 27. Nakajima,T.In Venoms of the Hymenoptera,Biochemical,Pharmacological and Behavioural Aspects;Editor T.Piek;Academic Press,London Chapt.6 pp.309-327. 28. Piek,T.;Hue,B.;Mony,L.;Nakajima,T.;Pelhate,M.;Yasuhara,T.Comp.Biochem. Physiol.1987, 87C,287-295. 29. Yasuhara,T.;Mantel,P.;Nakajima,T.;Piek,T.Toxicon 1987,25,527-535. 30. Piek,T.;Hue,B.;Mantel,P.;Nakajima,T.;Pelhate,M.;Yasuhara,T.Comp. Biochem. Physiol.1990,96C,157-162. 31. Piek,T.;Schmidt,J.O.;De Jong,J.M.;Mantel,P.Comp.Biochem.Physiol.1989, 92C,117-124. 32. Hue,B.;Piek,T.Comp.Biochem.Physiol.1989,93C,87-89. 33. Yoshida,H.;Geller,R.G.;Pisano,J.J.Biochemistry 1976,15,61-64. 34. Rocchi,R.;Biondi,L.;Filira,F.;Scolaro,B.Int.J.Pept.Prot.Res.1987,30,240-256. 35. Biondi,L.;Cavaggion,F.;Filira,F.;Rocchi,R.Gazz.Chim.Ital.1988,118,427-433. 36. Gobbo,M.;Biondi,L.;Filira,F.;Scolaro,B.;Rocchi;Piek,T.Int.J.Pept.Prot.Res. 1992,in press. 37. Schmidt,J.O.In Venoms of the Hymenoptera Biochemical,Pharmacological and Behavioural Aspects;Editor T.Piek;Academic Press London,Chapt.6 pp.425-508. 38. Piek,T.;Hue,B.;Mantel,P.;Nakajima,T.;Schmidt,J.O.Comp.Biochem.Physiol. 1991,99C, 481-486. 39. Piek,T.;Duval,A.;Hue,B.;Karst,H.;Lapied,B.;Mantel,P.;Nakajima,T.; Pelhate,M.; Schmidt,J.O.Comp.Biochem.Physiol.1991,99C,487-495. 40. Duval,A.;Malécot,C.O.;Pelhate,M.;Piek,T.Pflügers Arch.1992,in press 41. Piek,T.In Neuropharmacology of Insects; Publisher:Pitman,London,1982.Ciba Found.Symp.88 pp.275-290. 42. Piek,T.Pestic.Sci.1987,19,317-322. 43. Hildebrand,J.G. In Neurotox'88:Molecular Basis Drug & Pesticide Action; Editor G.G.Lunt;Elsevier,Amsterdam Exerpta Medica Int.Congr.Ser 832 pp. 583-588. 44. Milner-White,E.J. TIPS 1989,10,70-74. RECEIVED October 6, 1992 In Pest Control with Enhanced Environmental Safety; Duke, S., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1993.

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