Interaction between Manduca sexta allatotropin and manduca sexta allatostatin in the fall armyworm Spodoptera frugiperda.

A peptide that strongly stimulates juvenile hormone (JH) biosynthesis in vitro by the corpora allata (CA) was purified from methanolic brain extracts of adult Spodoptera frugiperda. Using HPLC separation followed by Edman degradation and mass spectrometry, the peptide was identified as Manduca sexta allatotropin (Mas-AT). Treating the CA from adult S. frugiperda with synthetic Mas-AT (at 10(-6) M) caused an up to sevenfold increase in JH biosynthesis. The stimulation of JH synthesis was dose-dependent and reversible. Synthetic M. sexta allatostatin (Mas-AS) (10(-6) M) did not affect the spontaneous rate of JH secretion from CA of adult S. frugiperda, nor did any of the allatostatins of the Phe-Gly-Leu-amide peptide family tested. However, when CA had been activated by Mas-AT (10(-6) M), addition of synthetic Mas-AS (10(-6) M) reduced JH synthesis by about 70%. This allatostatic effect of Mas-AS on allatotropin-activated glands was also reversible. When CA were incubated in the presence of both Mas-AT (10(-6) M) and various concentrations of Mas-AS (from 10(-8) to 10(-5) M), the stimulation of JH-biosynthesis observed was inhibited in a dose-dependent manner. The experiments demonstrate a novel mechanism of allatostatin action. In S. frugiperda JH synthesis was inhibited only in those glands which had previously been activated by an allatotropin.

Syntheses of analogues of the insect neuropeptide proctolin containing an oxazole ring as an amide bond replacement.

[structure: see text]. Three oxazole analogues of the insect neuropeptide proctolin (H-Arg-Tyr-Leu-Pro-Thr-OH) have been prepared, containing a single oxazole between Tyr and Pro and between Pro and Thr, respectively. A compound containing an oxazole moiety between Tyr and Pro and between Pro and Thr has also been prepared. All compounds have been tested for myotropic activity.

PF1022A--a novel anthelmintic cyclooctadepsipeptide. Modification and exchange of the N-methyl leucine residues.

The first structure-activity relationships of the anthelmintic cyclooctadepsipeptide PF1022A have been established via a systematic exchange of the leucine residues by a series of related N-alkylated amino acids. The data presented strongly suggest that (L)-N-methyl-leucine is crucial for high in vivo activity.

Azadepsipeptides: synthesis and evaluation of a novel class of peptidomimetics.

A general route to azadepsipeptides, a new class of pseudopeptides, has been established. The methodology was applied to the synthesis of a bis-aza analogue of the antiparasitic cyclooctadepsipeptide PF1022A. Comparison of the X-ray crystal structures of natural PF1022A (8) and the chimeric aza analogue 9 revealed that the introduction of nitrogen in the backbone of PF1022A results in almost complete conservation of the 3D structure with only minor deviations at the new nitrogen positions.

Pharmacological characterisation of 5-hydroxytryptamine-induced contractile effects in the isolated gut of the lepidopteran caterpillar Spodoptera frugiperda.

The indolealkylamine 5-hydroxytryptamine (5-HT, 0.1nM-1&mgr;M) caused dose-dependent increases in the number of contractions observed in guts isolated from the caterpillar Spodoptera frugiperda. Of the 5-HT analogues tested for agonist action, 2-methyl-5-HT (0.1-10&mgr;M) was a full agonist with reduced potency while alpha-methyl-5-HT (0.1-100&mgr;M), 5-carboxamidotryptamine (0.1-100&mgr;M), 5-methoxytryptamine (5-MeOT) (10nM-10&mgr;M), and tryptamine (1-100&mgr;M) were partial agonists. Incubation of isolated guts with proven mammalian 5-HT receptor antagonists showed that cyproheptadine (10nM-1&mgr;M), MDL 72222 (1-10&mgr;M), tropisetron (1-10&mgr;M) and 5-benzoyloxygramine (1-10&mgr;M) were potent non-competitive antagonists of 5-HT-induced tissue contraction. In comparison, ketanserin (0.1-1&mgr;M) was a competitive antagonist. The mammalian selective serotonin reuptake inhibitors, clomipramine (10nM-10&mgr;M) and fluoxetine (10nM-10&mgr;M) also caused non-competitive inhibition of 5-HT-induced contraction while fluvoxamine (10nM-10&mgr;M) was a weak competitive antagonist. Low doses of clomipramine (0.1&mgr;M) caused potentiation of 5-HT-induced gut contraction thereby suggesting the presence of 5-HT reuptake systems in this tissue. The contractile effects of 5-HT were inhibited by verapamil, Li(+) and H7 and potentiated by theophylline thereby indicating that L-type Ca(2+) channels, phosphatidylinositol second messengers and cAMP, respectively, are involved in 5-HT-induced tissue contraction. The 5-HT receptors mediating contractility in the gut of S. frugiperda have properties in common with mammalian 5-HT(2) and Drosophila 5-HT(dro2A/2B) receptors. In addition, these data suggest that the tissue also contains receptors that are similar to mammalian 5-ht(6) and 5-HT(7) as well as Drosophila(dro1) receptors. However, the primary amino acid sequence of these lepidopteran 5-HT receptors will have to be elucidated before full comparisons can be made.

Structure-activity relationship of contractile effects induced by helicokinins in the isolated gut of the lepidopteran caterpillar Spodoptera frugiperda.

The diuretic helicokinins YFSPWG-amide (Hez KI), VRFSPWG-amide (Hez KII) and KVKFSAWG-amide (Hez KIII) are potent contractants of the isolated gut of the caterpillar Spodoptera frugiperda at doses ranging from 0.1 to 10nM. In comparison, the pentapeptide FSPWG-amide was a full agonist with greatly reduced potency while SPWG-amide and PWG-amide were weak partial agonists. Substitution of individual amino acids in Hez KI with alanine revealed that replacement of the [phenylalanine(2)] residue caused a large fall in potency while replacement of [tryptophan(5)] residue caused complete loss of myogenic activity. The striking fall in potency of YASPWG-amide and the lack of activity of YFSPAG-amide confirm the requirement for aromatic groups in positions 2 and 3 of the core pentapeptide as well as supporting the ideas that the active core of these peptides adopts a beta-turn when interacting with receptors, bringing together the [Phe] and [Trp] residues that are critical for activity. Neither the pentapeptide proctolin nor the potent mammalian gut contractant Substance P were able to cause contraction when applied to caterpillar gut tissue. Incubation of isolated gut tissue in the phosphodiesterase inhibitor theophylline (10-100&mgr;M) caused significant potentiation of the response to applied Hez KI. Conversely, in the presence of the L-type Ca(2+) channel blocker verapamil (10&mgr;M-1mM) or Co(2+) (1-50mM) the contractile effects of Hez KI were attentuated significantly. These data suggest that the gut of S. frugiperda contains G-protein-linked kinin receptors that utilise cyclic AMP as their second messenger system and cause contraction by promoting the entry of extracellular Ca(2+).

Effect of helicokinins and ACE inhibitors on water balance and development of Heliothis virescens larvae.

The diuretic activity of the helicokinins I (YFSPWG-amide), II (VRFSPWG-amide) and III (KVKFSAWG-amide) was tested on Heliothis virescens larvae. All three kinins increased fluid secretion in isolated Malpighian tubules in a dose-dependent manner. Injections into the haemolymph caused a significant reduction in weight gain after 24 h and, in the case of helicokinin I, led to an increased mortality of 43% within 6 days. When truncated analogues of helicokinin I were tested in vitro, only the pentapeptide (FSPWG-amide) stimulated fluid secretion. Tested in vivo the pentapeptide did not influence normal development of the larvae. An alanine scan of helicokinin I showed that the substitution of phenylalanine, tryptophan and glycine led to a massive decrease or even loss of diuretic activity. The substitution of the other amino acids had no effect in vitro. The ACE inhibitors captopril, enalapril-maleate and lisinopril were tested for their influence on the development of the larvae. In combination with one of the helicokinins the in vivo injection of the ACE inhibitors led to increased rates of mortality and/or reductions in pupal weight.