František Sehnal

Insect silk glands: their types, development and function, and effects of environmental factors and morphogenetic hormones on them

Abstract Insect silk glands are ectodermal organs that evolved several times in the phylogeny from one of the following cell types: epidermal cells associated with bristle formation; sex accessory glands; larval Malpighian tubules; and larval labial salivary glands. As far as is known, the ultrastructure of the silk-producing cells is similar in all gland types, except for the labial glands of lower Hymenoptera. In most Lepidoptera and Trichoptera, the posterior gland region secretes fibroin and one to several small proteins; this core of silk is enveloped in the middle region of the gland by a mixture of sericins. The cuticle-lined anterior region provides the gland outlet. Lepidopteran silk glands become secretory prior to hatching. During larval life they increase their secretory potential by growing in size and ploidy. Their function fluctuates in each instar according to a “larval” pattern, which is characterized by the initiation of intensive RNA transcription with the resumption of feeding, by a high rate of proteosynthesis during feeding, and by cessation of these activities with the advancement of the next molt. In the last-larval instar, the silk glands develop according to a metamorphic pattern that differs from the larval one by enhanced function and, especially, by the programming of silk glands for histolysis. Starvation of larvae causes silk gland regression, cold shock prevents spinning, and exposure of larvae to alcohol vapours kills certain silk gland cells. Development of silk glands is altered with hormonal treatments of larvae. Juvenile hormone (JH) curbs silk gland function, prevents their degeneration, and, indirectly, it can cause an increase in silk production. Low doses of ecdysteroids accelerate silk gland development to increased function, while high doses cause regression and degeneration. Juvenile hormone anologues and anti-JH compounds are used in sericulture to control the yield and quality of silk. Available data are consistent with the following scheme of the hormonal control of silk glands. The change from the larval to the metamorphic developmental pattern is caused by a drop in JH titre. Realization of the pattern depends on consecutive action of several factors: trace amounts of JH in the last-larval instar affect silk glands via regulation of feeding and molt timing; function of silk glands depends on nutrient supply and it is stimulated by a brain neurohormone; slight elevation of ecdysteroid titre, which is associated with the termination of feeding and initiation of cocoon spinning, may be implicated in the culmination of proteosynthesis, and in the initiation of functional regression in the silk glands; the molt-inducing surge of ecdysteroids induces regression in the silk glands developing according to the larval pattern, but degeneration in those completing the metamorphic developmental pattern.

CONTROL OF CORPORA ALLATA FUNCTION IN LARVAE OF GALLERIA MELLONELLA

1. The implantation of three brains into freshly ecdysed last instar larvae which possess at least one of the two corpora allata induces extra larval development. Implanted brains appear to produce a neurohumoral allatotropic factor.2. Corpora allata seem to be inhibited via their nervous connections 48-60 hours after the last larval-larval ecdysis and become insensitive to the allatotropic factor. Severance of the nerves innervating the corpora allata and adjacent corpora cardiaca induces extra larval development in 7% of the experimental insects and restores sensitivity to the implanted brains in as many as 20%.3. Severance of the nerves to the corpora cardiaca—corpora allata complexes or implantation of corpora cardiaca—corpora allata complexes into allatectomized larvae in some instances causes a considerable prolongation of the last larval instar. It is suggested that disconnection of the corpora allata from the brain partly removes their inhibition and induces secretion of a low titer of juvenile ho...

Influence of the corpus allatum on the development of internal organs in Galleria mellonella L.

Effects of corpus allatum hormone (JH) on the growth and differentiation of the internal organs of Galleria larvae were investigated. Some of the internal organs develop continuously, others only for a period in each larval instar. They maintain larval form until the second third of the last larval instar or later and then differentiate towards the imaginal form. Differentiation appears as a continuous process of changes based mostly on degeneration of some parts and more profound growth of the others, which may be interrupted at most points by JH. In this way insects with internal organs intermediate between larval and imaginal structure result. The overall growth of the organs continues after hormone application but at a different rate. Slight morphological change in wing disks which appears in the penultimate larval instar may be also prevented by JH.

Control of prothoracic gland activity in larvae of Galleria mellonella

Abstract Prothoracic glands of last instar wax moth larvae maintain spontaneous secretory activity both in decapitated larvae and in isolated abdomens into which they have been transplanted, as judged by their ability to induce secretion of a new cuticle. Their activity is hormonally stimulated by the brain and inhibited by the prothoracic and mesothoracic ganglia. The subesophageal ganglion seems to suppress the inhibitory influence of the thoracic ganglia. The prothoracic glands of larvae decapitated at different times during the last instar all respond to brain implantation, and this response does not change when brains are implanted at increasing intervals after decapitation. The prothoracotropic activity of the isolated brain is highest in brains of pupae and adults but is relatively and consistently low in brains of last instar larvae. The results demonstrate that the control of prothoracic glands is a complex process governed by the nervous integration of various stimuli.

Inhibition of adult development in Tenebrio molitor by insect hormones and antibiotics

Abstract Juvenile hormone and its analogues, ecdysterone, actinomycin D, and mitomycin C prevent imaginal differentiation in Tenebrio pupae. All agents produce maximal effects when administered just after the larval-pupal ecdysis (with mitomycin 24 hr after ecdysis); applications at 80 hr after ecdysis or later are ineffective. The juvenile hormone is active in doses as low as 0·01 ng but perfect second pupae are produced with a hundred million times higher amounts. The moderate doses induce development of pupal-adult intermediates. Ecdysterone in amounts exceeding 0·02 μg causes formation of intermediates by accelerating the apolysis process. Administrations of high amounts of actinomycin D or mitomycin C (0·5 and 5 μg, respectively) to freshly ecdysed pupae block apolysis. The lower doses do not hinder the moulding process but bring about retention of some pupal features in emerging adults. These results reveal the significance of the DNA and RNA syntheses for apolysis and adult differentiation. The mode of action of insect hormones is also discussed.

Action of juvenoids on the metamorphosis of cyclorrhaphous Diptera

Abstract Administrations of high doses of juvenoids to the last instar larvae of cyclorrhaphous flies cause occasionally lethal defects in puparium formation but mostly affect only the pupal-adult transformation. In pupae, juvenoids impede proliferation and differentiation of the imaginal disks and of abdominal histoblasts: at low doses they cause incomplete rotation of male genitalia and deformations of the ovipositor, at higher doses their effects gradually spread from the tip of abdomen towards the middle of the body. The highest amounts influence the entire abdomen, size and pigmentation of the eyes, and development of hairs and sclerotization of the integument on the head and thorax. Various species slightly differ in the pattern of morphological effects produced, in the ability of affected insects to leave the puparium, and in the sensitivity to juvenoids of different types. A uniform scale for classification of morphological effects in the species examined is described in this paper. The most potent juvenoids are effective at doses around one nanogramme per specimen. Out of 29 selected compounds tested, isopropyl 11-methoxy-3,7,11-trimethyl-2,4-dodecadienoate is the most active juvenoid for Drosophila, Musca , and Sarcophaga ; methyl 10,11-epoxy-3,7,11-trimethyl-2,6-tridecadienoate is the most active juvenoid for Ceratilis ; and isopropyl 11-chloro-3,7,11-trimethyl-2-dodecenoate possesses the highest activity for Calliphora .

Natural and synthetic materials with insect hormones activity 8. Juvenile activity of the farnesane-type compounds on Galleria Mellonella

Abstract Farnesol, farnesyl ethers, farnesoate esters, and similar compounds act as potent analogues of juvenile hormone on several insect species (1,2,3). In tests on the larvae of Galleria mellonella , however, these chemicals displayed only very slight activities (unpublished observation). Their failure to induce a juvenilizing effect was puzzling since the larvae readily react to tiny amounts of Cecropia juvenile hormone (4,5) which is structurally related to farnesane-type sesquiterpenoids. Being stimulated by the contrast between the high activity of natural juvenile hormone (JH) and the negligible activity of farnesyl ethers and alkyl farnesoates, we started a detail investigation of the relation between the chemical structure and the juvenile hormone activity. This report describes the activity of compounds with the farnesane carbon skeleton but differing in the number of double bonds and the presence of halogen atom(s) or oxirane ring(s) in the molecule. It has been well established that there are great species specific differences in the activity of JH-analogues; e.g. the juvabione and related compounds act on Pyrrchocorids but not on other bugs (6). No attempts have been made, however, to examine whether similar differences do not exist also between successive developmental stages of a single species. Aiming to this goal, we employed in our tests both larvae and pupae of Galleria .

Inhibition of reproduction and embryogenesis in the firebrat, Thermobia domestica, by juvenile hormone analogues

Abstract Application of compounds with juvenile hormone (JH) activity to reproducing females may result in shortening of the ovipositor and occasionally in some other changes in appearance. Severe defects occur in the ovaries: the differentiation of oogonia and prefollicular cells seems to be hindered and the mature oocytes are resorbed. The affected ovarioles diminish and in extreme cases tear into the germarium, which remains attached to the terminal filament, and the rest of the ovariole, which shrinks into a small rudiment adjacent to the oviduct. The development of eggs deposited by treated females is often lethally affected. Disorders in embryogenesis also occur in the eggs contaminated with active substances at any time between deposition and 2 days before hatching. The substances interrupt the normal course of embryogenesis but the embryos survive for some time and may develop into strange creatures; in one case, two embryos were found within a single egg shell. One out of 27 tested substances was active in amounts lower than 1 ng/egg.

Hydrolysis and binding of the juvenile hormone in the haemolymph of Galleria mellonella

Abstract Haemolymph of fully grown larvae of Galleria mellonella was chromatographed on Sephadex G-200 and the column eluate assayed for hydrolytic activity and juvenile hormone (JH) binding. Hydrolysis of JH-I and 1-naphthyl acetate (1-NA) by high molecular fractions ( 2 × 10 5 ) was sensitive to 2 × 10 −4 M diisopropylphosphofluoridate (DFP). JH-Hydrolysis by proteins ranging in molecular weight from 1.4 × 10 5 to 3.7 × 10 4 was unaffected by DFP and 2.1 × 10 −4 M p- chloromercuribenzoate (PCMB), whereas the overlapping 1-NA hydrolysis was inhibited by these reagents. Several of the JH-esterase peaks coincided with those which hydrolyzed 4-methylumbelliferyl esters of JH-II acid, JH-III acid and oleic acid. The enzymes hydrolyzing these substrates were also unaffected by DFP and PCMB. Highest non-enzymatic binding of JH occurred in the protein fraction with mol. wt of 2.5 × 10 4 . The binding protected JH against most of the blood esterases.

Age dependent changes in the binding and hydrolysis of juvenile hormone in the haemolymph of last instar larvae of Galleria mellonella

Abstract Binding and hydrolysis of juvenile hormone I (JH) and cleavage of four 1-naphthyl esters were determined in the haemolymph of Galleria mellonella throughout the last larval instar. JH binding per ml haemolymph parallels changes in the protein content with a maximum at the onset of spinning at 120 hr (152 mg protein/ml; binding of 20 n-mole JH/ml) and a drop at the pupal ecdysis. Activity of JH esterase(s) exhibits two maxima: first between cessation of feeding and the start of spinning at 96–120 hr (hydrolysis of 65 n-mole JH/ml/min) and second just before pupal ecdysis at 168 hr (52 n-mole JH/ml/min). Correlations between these changes and available data on the JH content in caterpillars indicate that JH binding protein and JH esterase(s) may play a role in clearing JH from its target tissues rather than in regulating its amounts. Fluctuations in the hydrolysis of 1-naphthyl esters of acetate (max. 4.5 μ-mole/ml/min), butyrate (max. 21 μ-mole/ml/min), palmitate (max. 0.5 μ-mole/ml/min), and laurate (max. 0.4 μ-mole/ml/min) reflect primarily the feeding activity of caterpillars and follow a very different course than changes in the potential to hydrolyze JH.