The hormonal control of salivary gland secretion in Drosophila melanogaster: Studies in vitro

The larval salivary gland of Drosophila melanogaster synthesises a complex secretion, known as ‘glue’. which is secreted at puparium formation and then cements the puparium to its substrate. This secretion is made during the third larval instar and is stored in the gland cells as large granules. A few hours before puparium formation it is secreted into the gland's lumen by exocytosis. This process is induced by ecdysone and can be studied in vitro. Secretion is initiated about 3.5 hr after exposure of glands to ecdysone and is complete by 8 hr. The effects of varying the ecdysone concentration, of inhibitors of RNA or protein synthesis, and of withdrawing the hormone at various times after initial exposure on the process of secretion have been studied. We conclude that some event(s) occurring during the first 3 hr exposure to ecdysone is necessary to initiate secretion of the glue into the gland lumen. The possible relationship between this event(s) and the ecdysone induced changes in gene activity (puffs) which occur in the salivary glands at the same time is discussed.     

Dufour gland secretion of Polistes wasp: Chemical composition and possible involvement in nestmate recognition (Hymenoptera: vespidae)

The Dufour gland secretion and cuticle lipids of foundresses of Polistes dominulus have been chemically analysed. The gland secretion (analysed for the first time in a polistine species) contains a complex mixture of hydrocarbons, which are the same as those found on the cuticle. The cuticular hydrocarbon mixture and gland secretion differ, however, in the proportions of components: in the first, linear hydrocarbons are more abundant, while in the second we found a higher quantity of dimethylalkanes. Multivariate statistical analysis has shown that foundresses belonging to different colonies are distinguished on the basis of both the composition of their Dufour gland secretion and cuticular hydrocarbon mixture. Branched hydrocarbons seem to be particularly important in distinguishing females according to their colony. We suggest that the Dufour gland may contribute substances to form the hydrocarbon layer on the cuticle.     

CHANGES IN CHROMATIN TEMPLATE ACTIVITY AND THEIR RELATIONSHIP TO DNA SYNTHESIS IN MOUSE PAROTID GLANDS STIMULATED BY ISOPROTERENOL

Chromatin template activity of mouse parotid glands increases after a single injection of isoproterenol (IPR), a procedure that causes, after a lag period of 20 hr, a marked stimulation of DNA synthesis and cell division in salivary glands of rodents. The increase in chromatin template activity occurs as early as 1 hr and peaks between 6 and 10 hr after IPR, paralleling previously reported changes in the incorporation of uridine-³H into total cellular RNA of mouse parotids. Template activity was measured in vitro in a system in which parotid gland chromatin was incubated with an exogenous RNA polymerase isolated from Escherichia coli. Similar results were obtained when template activity of parotid gland chromatin was assayed using an homologous RNA polymerase from mouse liver. Chromatin template activity in mouse parotids was also studied after the administration of drugs capable of inducing in salivary glands both DNA synthesis and secretion or secretion alone. The results indicate that the increased chromatin template activity occurring 6 hr after IPR is related to the subsequent onset of DNA synthesis. Furthermore, the increased chromatin template activity caused by IPR is inhibited by the previous administration of puromycin, an inhibitor of IPR-stimulated DNA synthesis.     

The differential regulation of the synthesis of ribosomal RNA, 5 S RNA, and 4 S RNA in the polytenic salivary gland cells of the blowfly, Calliphora erythrocephala

Third-instar larvae of the blowfly Calliphora erythrocephala were injected with [2-³H]adenosine, and its flow into the salivary gland ATP pool and each of several electrophoretically resolved salivary gland RNA species were quantitated. From these data, the individual in vivo rates of synthesis, accumulation, and processing of salivary gland ribosomal RNA (rRNA), 4 S RNA, and 5 S RNA have been measured at several different developmental stages. These results indicate that the synthesis of 5 S RNA and rRNA are coordinate, developmentally regulated, and independent of the synthesis of 4 S RNA. A nonribosomal, heterodisperse RNA component (hdRNA) was also identified. This species contributes to both the rapidly turning over pulse-labeled RNA and the accumulating pulse-labeled RNA populations. Indirect measurements suggest that the developmental pattern of regulation of this RNA species is also independent of 5 S RNA and rRNA synthesis. The rate of synthesis and accumulation of each of these RNA species either remained constant or declined during the first three-fourths of the instar, despite a six- to sevenfold increase in the content of cellular DNA.     

The nasus gland: A new gland in soldiers of Angularitermes (Termitidae,Nasutitermitinae)

Termites have developed many exocrine glands, generally dedicated to defence or communication. Although a few of these glands occur in all termite species, or represent synapomorphies of larger clades, others are morphological innovations of a single species, or a few related species. Here, we describe the nasus gland, a new gland occurring at the base of the nasus of Angularitermes soldiers. The nasus gland is composed of class 1, 2, and 3 secretory cells, a rare combination that is only shared by the sternal and tergal glands of some termites and cockroaches. The ultrastructural observations suggest that the secretion is produced by class 2 and 3 secretory cells, and released mostly by class 3 cells. The base of the nasus has a rough appearance due to numerous pits bearing openings of canals conducting the secretion from class 3 secretory cells to the exterior. We tentatively assign a defensive function to the nasus gland, although further research is needed to confirm this function. Although the gland is described only from species of Angularitermes, other genera of Nasutitermitinae also present a rough nasus base, suggesting the presence of a similar, possibly homologous, gland.     

Protein metabolism of Drosophila melanogaster male accessory glands—III: Stimulation of protein synthesis following copulation

Stimulation of the accessory gland of Drosophila melanogaster males by copulation was studied at the molecular level by in vivo radiolabelling experiments. In addition, experiments were carried out to clarify the molecular basis of regulation of the gland's synthetic activityThe stimulatory effect of copulation on secretion protein synthesis is observed within minutes after copulation is terminated. Repeated copulations elicit a stronger response. The data indicate a coordinate synthesis of the secretory proteins.In a reticulocyte-lysate system with added total RNA extract from male accessory glands, secretion proteins appear as translation products. It is shown that synthesis of secretory protein messages is not enhanced by copulation and that the messages are stable.Synthesis of ribosomal proteins and ribosomal RNA increases upon copulation. The increase is observed in the time period from 1 to 5 hr after copulation. Again, the synthesis of the various ribosomal proteins is well coordinated. There is no elevation in the accumulation of ribosomal protein mRNAs after copulation.     

SOURCES OF LARVAL SALIVARY GLAND SECRETION IN THE DIPTERAN CHIRONOMUS TENTANS

The soluble proteins in the hemolymph, the salivary gland, and the salivary secretion of fourth instar Chironomus tentans were examined by disc electrophoresis in acrylamide gels. Of the 11 protein fractions detected in buffered saline extracts of the gland, 10 are present also in the hemolymph. Amino acid isotope incorporation experiments indicate that the protein fractions shared by the salivary gland and the hemolymph are not synthesized in the gland but are synthesized in other larval tissues. Immunochemical studies show that most of these proteins eventually are secreted from the gland. The salivary gland in vivo and in vitro is active in de novo protein synthesis. The protein synthesized tends to form large molecular weight aggregates. As demonstrated by radioautography, at least 80% of this protein is secreted from the 30 large cells forming most of the gland. The proteins synthesized in the salivary gland cannot be detected in the hemolymph. The results of this investigation are consistent with a mechanism of secretion formation involving both de novo synthesis of some secretion proteins and the selective uptake, transport, and secretion of hemal proteins by the salivary gland.     

The secretion and action of the digestive enzymes of the salivary glands of the blowfly, Calliphora

The salivary glands of adult Calliphora contain enzymes which hydrolyze starch, sucrose and trehalose. Amylase and sucrase are shown to be secretory enzymes, while trehalase remains in the gland. Results of electrophoretic and ultrastructural studies suggest that protein secretion is confined to the abdominal region of the gland. Secretion of both fluid and protein occurs from a single type of cell. While a fly is feeding on solid sugar, amylase and sucrase are lost from the gland and appear in saliva, while the level of trehalase in the gland increases slightly. The mixture of food and saliva passes mainly to the crop where carbohydrate is digested by the salivary enzymes.     

Electron microscope studies of Fasciola hepatica. X. Egg formation

Evidence obtained would suggest that the processes involved in egg formation in Fasciola hepatica occur in the following sequence. The egg constituents, namely an ovum and some vitelline cells, pass through the proximal ootype and as they do so they are smeared by the secretions of Mehlis' gland which have accumulated there. A temporary interface is set up between the Mehlis' gland secretion and the fluid which surrounds the egg constituents. Shell globules are released by the vitelline cells and coalesce on the inner aspect of the interface. At the same time some of the Mehlis' secretion diffuses across the interface, thereby bringing about the dissociation of the interface. It is suggested that Mehlis' gland secretion brings about the fusion of the shell layer. During the initial stages of the process the developing egg is supported by the cells of the ootype epithelium. Later the egg passes into a wider, more distal part of the ootype where the process of shell deposition is continued. When complete, or almost complete, the egg passes into the uterus where changes indiciating the initial process of tanning are seen to take place. A thin, very uniform layer of Mehlis' gland secretion can still be identified on the surface of mature eggs.     

Phéromones agrégeant les ouvrières de Myrmica rubra

The workers of Myrmica rubra aggregate around a source of one of their secretions, which can be called ‘alarm pheromone’, and also around workers of Lasius flavus. The mechanism of these aggregations differ.Both L. flavus workers and a solution in liquid paraffin of 3-octanol, one of the mandibular gland compounds, act as an arrestant for the workers of M. rubra. Both Dufour's gland secretion and a source of 3-octanone, the major compound of the mandibular gland secretion, are true attractants.The poison gland secretion, a mixture of 3-octanone and 3-octanol in liquid paraffin and a solution in liquid paraffin of 3-nonanone, a minor mandibular gland compound, all induce klinokinesis. The secretion of the mandibular glands and the secretion of the venom apparatus both cause positive klinokinesis and taxis. These locomotory reactions increase the probability that an object, marked by nest mates with these secretions, will be detected by several workers.When presented alone, 3-octanone is the only attractive compound in the mandibular gland secretion. However, a mixture of 3-octanone and 3-octanol (15 per cent of 3-octanol in the vapour phase) is detected more easily by the ants. The diffusion coefficients of the two compounds are different, and a mixture of these substances creates not only a quantitative but also a qualitative odour gradient. This may explain the synergy of the mixture.     

Cutaneous application of an accessory-gland secretion after sperm exchange in a terrestrial slug (Mollusca: Pulmonata)

Competition for fertilisation in hermaphroditic animals seems to have led to many odd behaviours and complex morphologies involved in the transfer of accessory-gland products to the partner. Terrestrial slugs of the genus Deroceras show remarkably elaborate and interspecifically diverse penis morphologies and mating behaviours. Most species have an appending penial gland, which in Deroceras panormitanum consists of a few long fingers that are everted after sperm exchange and laid onto the partner’s back. To investigate whether this gland transfers a secretion onto the partner’s skin, we killed slugs at different mating stages and studied their penial glands and skin histologically. Two types of secretion granules appeared at a very early stage of courtship, and the penial gland was already filled 15 min into the courtship. At copulation, the gland everted this secretion onto the partner’s body, where it remained for at least 50 min. No lysis of skin tissue or other effects on the skin were observed. The slugs tried to lick the received secretion off their own body, and some droplets were observed to be shed with the body mucus. Our results indicate the external application of a glandular substance that could function as either a pheromone or allohormone. The behaviours of the recipients suggest sexual conflict, although mutual interest cannot be ruled out.     

Evolutionarily Conserved Repulsive Guidance Role of Slit in the Silkworm Bombyx mori

Axon guidance molecule Slit is critical for the axon repulsion in neural tissues, which is evolutionarily conserved from planarians to humans. However, the function of Slit in the silkworm Bombyx mori was unknown. Here we showed that the structure of Bombyx mori Slit (BmSlit) was different from that in most other species in its C-terminal sequence. BmSlit was localized in the midline glial cell, the neuropil, the tendon cell, the muscle and the silk gland and colocalized with BmRobo1 in the neuropil, the muscle and the silk gland. Knock-down of Bmslit by RNA interference (RNAi) resulted in abnormal development of axons and muscles. Our results suggest that BmSlit has a repulsive role in axon guidance and muscle migration. Moreover, the localization of BmSlit in the silk gland argues for its important function in the development of the silk gland.     

A calcium ionophore-induced secretion in rabbit lacrimal gland in vivo

Local administration of the calcium ionophore, A-23187 increased basal fluid secretion (non-stimulated) from the cannulated main excretory duct of rabbit lacrimal gland in vivo. A-23187 also facilitated fluid secretion induced by submaximal dose of methacholine (0.1 μg/kg, intraarterially). The stimulatory effect of A-23187 was dependent on the extracellular calcium concentration. Lowering the extracellular calcium by addition of EGTA markedly depressed or abolished the responses to the ionophore while increasing the extracellular calcium with CaCl₂ enhanced it. The results suggest that A-23187 causes increase in cell membrane permeability to extracellular calcium and the rise in intracellular calcium activates the secretory process(es) by an unknown mechanism to produce fluid secretion in the rabbit lacrimal gland.     

In-situ Hybridization to Messenger RNA in Heterodera glycines

A method is presented for in-situ hybridization to mRNA in second-stage juveniles (J2) of the soybean cyst nematode Heterodera glycines. The protocol was developed using a digoxigenin-labeled RNA probe transcribed from cDNA of a cellulase gene that was known to be expressed in the subventral esophageal glands of H. glycines. Formaldehyde-fixed J2 were cut into sections with a vibrating razor blade to make the inside of the nematodes accessible for probing. These nematode fragments then were hybridized in suspension with riboprobe, and labeled with an alkaline phosphatase-conjugated antibody to digoxigenin. Staining with nitroblue tetrazolium and bromo-chloro-indolyl phosphate revealed a highly specific hybridization signal to mRNA within the cytoplasm of the subventral gland cells, using this specific antisense probe. This in-situ hybridization protocol will be useful for the characterization and identification of esophageal gland secretion genes in plant-parasitic nematodes, among other applications.     

Histochemistry of the glands associated with the reproductive tract ofLymnaea stagnalis

Summary The albumen gland, the muciparous gland and the oöthecal gland of female genital tract of Lymnaea stagnalis, collected in spring, autumn and winter have been studied.The reactions for polysaccharides, proteins and RNA have been performed in order to characterise the secretion of the glands.The albumen gland secretion consists almost exclusively of slightly acid polysaccharides whose histochemical reactions, according to Lison and Grainger and Shillitoe confirm the presence of galactogen. Proteins are also present in the secretion. The muciparous gland secretion consists of strongly acid mucopolysaccharides (non sulphated) produced by large cells among which small cells containing sulphated mucopolysaccharides are present.In the oöthecal gland two zones are present, one with a single type of cells containing acid mucopolysaccharides, and the other with two different types of cells: the first with mucopolysaccharides and the second with sulphated mucopolysaccharides, proteins and glycogen at the basis of the cell. Sialic acids are not present in the secretion of the glands studied.The polysaccharidic composition of the secretion of the glands is different from gland to gland. The secretion of the glands gradually changes and gets acid according to the composition of the various membranes and envelopes wrapping up the eggs.     

In vitro release of house fly,Musca domestica l. (Diptera : Muscidae), acrosomal material after treatments with secretion of female accessory gland and micropyle cap substance

Mature house fly, Musca domestica L. (Diptera : Muscidae), sperm were treated with female accessory gland secretion and micropyle cap substance to determine their effectiveness in eliciting an acrosome response. Scanning and transmission electron microscopy revealed that release of acrosomal material was achieved by treatment of the sperm with a combination of accessory gland secretion and micropyle cap substance but not by gland secretion alone. The gland secretion was utilized to dissolve the cap substance from mature ovarian eggs, and this combined solution was applied to sperm removed from the spermathecae of mated females. The acrosomes of several of the sperm were completely lacking, while others showed a partial effect of the treatment, i.e. extensive acrosomal membrane fragmentation and vesiculation. Most sperm that reacted to the treatment lacked a plasma membrane around the acrosome, suggesting that this membrane, along with the acrosomal membrane, is involved in formation of the vesicles alongside the acrosomal cavity.     

The clypeal gland: A new exocrine gland in termite imagoes (Isoptera: Serritermitidae,Rhinotermitidae, Termitidae)

Social insects possess a rich set of exocrine organs producing diverse pheromones and defensive compounds. This is especially true for termite imagoes, which are equipped with several glands producing, among others, sex pheromones and defensive compounds protecting imagoes during the dispersal flight and colony foundation. Here, we describe the clypeal gland, a new termite exocrine organ occurring in the labro-clypeal region of imagoes of most Rhinotermitidae, Serritermitidae and Termitidae species. The clypeal gland of Coptotermes testaceus consists of class 1 (modified epidermal cell) and class 3 (bicellular gland unit) secretory cells. Ultrastructural features suggest that the gland secretes volatile compounds and proteins, probably after starting the reproduction. One peculiar feature of the gland is the presence of multiple secretory canals in a single canal cell, a feature never observed before in other insect glands. Although the function of the gland remains unknown, we hypothesize that it could produce secretion signalling the presence of functional reproductives or their need to be fed.     

Dermal gland secretion improves the heat tolerance of the brown dog tick, Rhipicephalus sanguineus, allowing for their prolonged exposure to host body temperature

Dermal glands (sensilla sagittiformia) secrete when brown dog ticks, Rhipicephalus sanguineus, are mechanically disturbed, presumably as a defensive mechanism. Recently, we observed that these glands secrete due to the pressure stimulation of engorgement. In this study, we examine how dermal gland secretion alters the physiology of R. sanguineus, particularly if this secretion is an important mechanism during blood feeding. The ability of ticks to retain water was not modified by dermal gland secretion, but heat tolerance was enhanced. Short-term heat shock was improved slightly (1 h at 50 °C to 1 h at 56 °C) and featured reduced injury responses and greater recovery after heat shock. When exposed to their host body temperature (37 °C) for prolonged periods, individuals that had secreted survived over 1 week longer than individuals that did not secrete. Dorsal application of squalene, the main component of dermal gland secretion, did not increase temperature tolerance, suggesting that the act of secreting rather than the physical properties of the secretion itself is responsible for the increase in heat tolerance. Based on our results, dermal gland secretion may be an essential mechanism in certain tick species (Amblyomma, Dermacentor, Hyalomma, Rhipicephalus, but not Ixodes) for tolerating body temperature and not succumbing to heat stress during the extended time periods of feeding on a mammalian host, serving as a mechanism to prevent heat damage from the host during feeding.     

Male specific natural products in the bug, Leptoglossus phyllopus: Chemistry and possible function

In the Coreoidea and some allied groups, male adults possess an abdominal gland opening through a midventral ostiole in the 7–8th abdominal intersegmental membrane. Using a GC-MS system, the following aromatic volatiles were identified in the abdominal gland secretion from males of the leaf footed bug, Leptoglossus phyllopus: guaiacol, benzyl alcohol, syringaldehyde, methyl p-hydroxybenzoate, acetosyringone, and vanillin. Males from which the glands were removed still mated and were competitive with control males for a limited number of females. The ventral abdominal gland secretion may act as a long-range attractant of females. The possibility that attraction of females by males in Heteroptera is an adaptation facilitating colonization of successional habitats is discussed.