Developmental profile of sinalbin (p-hydroxybenzyl glucosinolate) in mustard seedlings,Sinapis alba L., and its relationship to insect resistance.

Sinalbin was identified as a chemical component of insect anti-xenosis and antibiosis resistance mechanisms in seedlings ofSinapis alba by DEAE-Sephadex chromatography, HPLC, treatment with sulfatase and myrosinase, various feeding tests using artificial and natural substrates, and by measuring sinalbin concentrations in cotyledons and leaves during seedling development. The effects of sinaibin on feeding were dependent upon the insect species and upon the rapidly changing profile of sinaibin concentrations in the developing seedling. The high concentrations of sinalbin found in young cotyledons (up to 20 mM) and leaves (up to 10 mM) deterred the feeding of the flea beetle,Phyllotreta cruciferae Goeze and larvae of the bertha armyworm,Mamestra configurata Walker. The protection that sinalbin confers upon the vulnerable, newly emerged seedling (and upon tiny, young leaves) appears critical for the first few days of survival ofS. alba under feeding pressure from flea beetles in the field. The lower concentrations of sinaibin found in older cotyledons and leaves (2-3 mM) offer little or no protection againstP. cruciferae and may actually stimulate the feeding of this crucifer specialist. These concentrations of sinaibin, however, are still effective in reducing the level of feeding by larvae of the more generalist feederM. configurata.

Glucosinolate levels in cotyledons of mustard,Brassica juncea L. and rape,B. napus L. do not determine feeding rates of flea beetle,Phyllotreta cruciferae (Goeze).

Sinigrin (allyl glucosinolate), the major glucosinolate in the cotyledons ofBrassica juncea cv. Cutlass, occurred in the highest concentration and amount at seedling emergence and declined during growth. Glucobrassicin (3-indolylmethyl glucosinolate), the major glucosinolate in the cotyledons ofB. napus cv. Westar, occurred in the lowest concentration and amount at seedling emergence. The amount of glucobrassicin per cotyledon pair increased about fourfold during 14 days of growth, but its concentration remained relatively unchanged because of "dilution" by increasing cotyledon biomass. These different glucosinolate profiles indicate a different metabolic control and different biological function for sinigrin and glucobrassicin. The flea beetle,Phyllotreta cruciferae Goeze, does not discriminate between cotyledons having sinigrin or glucobrassicin since the two crucifers were fed upon equally in choice tests. Restricting the concentration of sulfur in the nutrient medium accelerated the decline of sinigrin inB. juncea cv. Cutlass but did not alter the feeding rate ofP. cruciferae compared to controls. Sulfur restriction reduced glucobrassicin inB. napus cv. Westar to undetectable levels and somewhat reduced the feeding rate of P.Cruciferae. Nevertheless,P. cruciferae still fed actively on cotyledons ofB. napus cv. Westar depleted of glucosinolates and severely damaged many of them. Since glucosinolate type and concentration had little effect on feeding response, reduction or elimination of foliar glucosinolates alone would not seem a useful strategy for protecting seedlings of these two crucifers from flea beetle damage.

Taurine inhibits octopamine-stimulated cAMP production in cockroach haemocytes.

Taurine and the taurine analogue guanidinoethanesulfonic acid interact with octopamine receptors of cockroach hemocytes to decrease octopamine-stimulated cAMP production. Dopamine-, synephrine- and tyramine-stimulated cAMP production in the haemocytes are also inhibited by taurine.

Effect of temperature on the rate of pupal-adult development of the noctuid moth,Mamestra configurata Wlk.: evidence for differential effects on the initiation of development and subsequent metamorphic development.

Moths eclosed earlier from pupae of the bertha armyworm,Mamestra configurata, that were exposed briefly (1 to 5 days) to a warm temperature (15 or 20°C) at the beginning of postdiapause pupal-adult metamorphosis and then incubated at 10 or 12.5°C than from pupae incubated at 10 or 12.5°C throughout metamorphosis. The differences were greater than could be explained by the additional thermal units received at the higher temperature. Analyses of the times of peak concentrations of ecdysteroids (insect growth and development hormones) in metamorphosing pupae and of moth eclosion after exposure to various combinations of temperatures indicated that the 'warm termperature effect' was not on the rate metamorphic development but on an earlier neuroendocrine process concerned with the initiation of development.The magnitude of the difference in eclosion time between pupac receiving a brief warm temperature "trigger" and the control suggests that the differential effect of temperature on the initiation of development and subsequent metamorphic development is of biological significance and should be considered in the construction of models of insect development under natural conditions.

Effects of cold stress during pupal diapause on the survival and development of Mamestra configurata (Lepidoptera: Noctuidae).

The effects of exposing diapausing pupae of Mamestra configurata Wlk. to temperatures from -5° C to-20° C during a 140-day experimental period were reduced survival to post-diapause stages, reduced survival to emergence as malformed or normal adults, and a reduced rate of post-diapause development. Pupae given a continuous exposure of variable length to -10° C and pupae given the same exposure to -10° C, interrupted by 28 days at 0° or -5° C, had the same survival, indicating that repair of cold injury does not occur at 0° or-5° C. Assuming that the effects of low temperatures were additive, not only for exposures to one temperature but also for exposures to different combinations of temperatures, survival was decribed by a polynomial: [Formula: see text] where p=the proportion in post-diapause stages and t=the number of days of exposure to the temperature given by the subscript. The values of the coefficients (C) were inversely related to the temperature of exposure (T): log10 C=-2.87-0.159 T. In validation experiments, the model accurately predicted survival of pupae in the laboratory and in soil under natural snow-cover or in soil where snow cover was maintained at <2 cm through a complete winter. Our results suggest that: (1) symptoms of cold injury, from death in the stage exposed through death, malformation or retarded development in post-exposure stages represent a continuum that can be produced by varying the exposure time at temperatures that cause cold injury; (2) repair does not occur at these temperatures nor at slightly warmer temperatures (e.g. at 0° C for M. configurata pupae); (3) the total amount of injury is not affected by the sequence of temperatures to which the insect is exposed; (4) freezing is not responsible for the observed cold injury.

Characterization of an octopamine-sensitive adenylate cyclase from insect brain (Mamestra configurata Wlk.).

An adenylate cyclase present in the brain of the moth Mamestra configurata Wlk. that is stimulated selectively by low (micromolar) concentrations of octopamine has been characterized with respect to several properties. The optimum pH, optimum ATP:Mg2+ ratio, the concentration of ATP required for half-maximal and maximal reaction velocity, metal ion specificity, effect of NaF, and effects of GTP and 5'-guanylylimidodiphosphate were in general similar to those of catecholamine-sensitive adenylate cyclases from various regions of mammalian brain. However, ethylene glycol bis-(beta-aminoethyl ether)-N,N-tetraacetic acid (EGTA), a calcium chelator, stimulated both basal and octopamine-sensitive enzyme activity in the insect brain, whereas in mammalian brain EGTA is usually observed to inhibit basal activity but not catecholamine-stimulated activity. Adenylate cyclase activity of the 47,000 g particulate fraction of the insect brain was almost undetectable in the absence of added GTP. Addition of saturating concentrations (100 micrometer) of GTP to the particles restored about 30% of the basal and octopamine-sensitive enzyme activity present in the homogenate. Addition of 100,000 g supernatant to the particles doubled both basal and octopamine-sensitive enzyme activity in the presence of saturating concentrations of GTP, indicating that in addition to GTP, a cytosolic factor(s) is necessary for enhanced adenylate cyclase activity.

Correlation between organophosphate poisoning, acetylcholinesterase inhibition, and increased cyclic GMP levels in malathion-treated insects.

Organophosphate poisoning with malathion caused large increases (up to 125 and 440%, respectively) in the level of cyclic GMP in larvae of Mamestra configurata Wlk. and in the fly Sarcophaga bullata Parker. Cyclic AMP was little affected. The malathion-induced increase in cyclic GMP was time and dose dependent. Time-course studies with the head and thorax of S. bullata demonstrated that the increase in cyclic GMP level occurred precipitously after a lag period of about 1 h, during which time the activity of acetylcholinesterase (EC 3.1.1.7) was progressively inhibited. The abrupt increase in cyclic GMP began when acetylcholinesterase activity had been inhibited to a sufficient extent to permit accumulation of acetylcholine. It is suggested that the accumulation of acetylcholine in the malathion-poisoned insects caused cyclic GMP levels to rise. Cyclic GMP may have a role in cholinergic transmission in normally functioning insect neural tissue. Increased levels of cyclic GMP induced by organophosphate and organocholorine (Bodnaryk, R. P. (1976) Can. J. Biochem. 54, 957-962) insecticides appear to be a vital and previously unrecognized biochemical lesion in insects poisoned by these compounds.

The effect of p,p'-dichlorodiphenyltrichloroethane on levels of guanosine 3',5'-cyclic monophosphate and adenosine 3',5'-cyclic monophosphate in two species of insects.

Within 1 h after topical application of a convulsive dose (4 mug per fly, 47 mg/kg) of p,p'-dichlorodiphenyltrichloroethane (DDT) to the adult male of Sarcophaga bullata Parker, guanosine 3',5'-cyclic monophosphate (cyclic GMP) levels rose by 71.5% (P less than 0.05) in the head, 159.5% (P less than 0.01) in the thorax, and 23.4% (P greater than 0.05) in the abdomen compared to controls. Adenosine 3',5'-cyclic monophosphate (cyclic AMP) levels were not significantly affected by the DDT treatment. A convulsive dose (100 mug per larva, 250 mg/kg) of DDT applied to larvae of Mamestra configurata Wlk. caused the whole body level of cyclic GMP to rise by 81.6% (P less than 0.01) after 1 h, and by 95.9% (P less than 0.01) after 3 h. Levels of cyclic AMP were not affected. A hypothesis is advanced suggesting that an abnormally high rate of discharge of acetylcholine (and in the later stages of poisoning, its actual accumulation) at central cholinergic synapses causes cyclic GMP levels to rise, perhaps in post-synaptic cells. The elevated cyclic GMP-cyclic AMP ratio found in DDT-poisoned insects may be of fundamental importance in the complex sequence of events leading to tremor, hyperexcitability, paralysis, and death.

3-O-hydrosulphato-4-hydroxyphenethylamine (dopamine 3-O-sulphate), a metabolite involved in the sclerotization of insect cuticle.

Dopamine 3-O-sulphate (3-O-hydrosulphato-4-hydroxyphenethylamine) was isolated from newly ecdysed cockroaches, Periplaneta americana (L.), and its structure established by chemical and physical techniques and by synthesis. Relatively high concentrations (about 1mumol/g wet. wt.) of dopamine 3-O-sulphate exist in the newly ecdysed insect, and these concentrations decrease sharply as sclerotization of the cuticle proceeds. At least 40% of the radioactivity of (14)C-labelled dopamine 3-O-sulphate injected into newly ecdysed nymphs was recovered in the sclerotized cuticle 7-12 days after the injection. However, less than 1% of the radioactivity of injected dopamine 3-O-[(35)S]sulphate was recovered, and this value was not appreciably different from that for the incorporation of Na(2) (35)SO(4). Apparently, little or none of the sulphate moiety of dopamine 3-O-sulphate is incorporated directly into the cuticle as the intact sulphate ester. These observations are discussed in relation to current concepts of cuticular sclerotization in insects.

Beta-alanyl-L-tyrosine. Chemical synthesis, properties and occurrence in larvae of the fleshfly Sarcophaga bullata Parker.

1. A naturally occurring peptide, termed sarcophagine, has been isolated from larvae of the fleshfly Sarcophaga bullata. 2. The physicochemical properties of the natural peptide are identical with those of synthetic beta-alanyl-l-tyrosine. 3. During larval growth the concentration of the dipeptide increases continuously; in the fully grown larva it is the predominant non-protein ninhydrin-positive material. 4. Sarcophagine is rapidly and extensively utilized at the time of hardening and darkening of the insect puparium. 5. By virtue of its much higher solubility than that of free tyrosine, the peptide may serve as a reservoir to provide large amounts of readily available tyrosine for subsequent puparium formation.

Naturally occurring low-molecular-weight peptides from the blowfly Phormia regina.

Freely occurring alpha-l-aspartyl-l-histidine, beta-alanyl-l-histidine (carnosine), alpha-l-aspartyl-alpha-l-lysine, l-asparaginyl-alpha-l-lysine, alpha-l-glutamyl-alpha-l-lysine, l-isoleucyl-alpha-l-lysine, l-seryl-alpha-l-lysine, l-valyl-alpha-l-lysine and glycyl-alpha-l-lysyl-alpha-l-lysine were identified in young larvae (second and third instars) of the blowfly (Phormia regina).