Salivary Glands
By: Clarence Collison
Salivary glands are involved in digestion, cleaning, softening foods, metabolism, growth, labor transition, brood pheromone and hormone regulation, saliva synthesis and even silk production.
Figure 1. A longitudinal section through the head of a worker showing the position of the hypopharyngeal glands (hyg) and the post cerebral salivary glands (sh) and thoracic salivary glands (st). Hypopharynx (hy). (Goodman 2003).
Honey bee salivary glands are important exocrine glands (chemical products are secreted to the body exterior). Salivary glands, also known as labial glands, are composed of two parts, the cephalic salivary glands or post-cerebral glands located in the head behind the brain and the thoracic salivary glands in the ventral part of the thorax, all of which connect through a common duct to the mouth (Figure 1). The two pairs of salivary glands open into a narrow sac or salivarium behind the posterior flange of the hypopharynx, by means of the common duct (Goodman 2003) (Figure 2). The adult cephalic and thoracic salivary glands begin to develop during the pupal period and progressively increase in size from the newly emerged workers to foragers, reaching their maximal size when the workers begin to forage (Katzav-Gozansky et al. 2001). Thoracic salivary glands are large in queens and workers and small in drones. Cephalic salivary glands are also large in queens and workers but vestigial in drones. The cephalic glands of queens increase in activity when they start egg laying.
Figure 2. Salivary gland system of adult worker honey bees. Head or post-cerebral salivary glands (HGld), thoracic salivary glands, common salivary duct (slDct), salivarium (Slv), thoracic salivary gland reservoir (Res). D, detail of head gland. E. detail of thoracic gland. (Snodgrass 1956).
In the worker each head gland consists of a loosely arranged mass of small pear-shaped bodies with individual ducts that unite irregularly with each other and eventually come together in a single duct that joins the common median duct from the thoracic glands (Figure 2D). Each thoracic gland consists of a mass of many-branched glandular tubules opening into short collecting ducts that unite in several major ducts which end in a sac-like reservoir at the anterior end of the gland (Figure 2E). The final outlet duct proceeds forward from the reservoir and is joined by the ducts from the glands in the back of the head (Snodgrass 1956).
The thoracic and cephalic salivary glands differ in protein expression, producing different kinds of secretions that have different functions, even though they have a common embryonic origin and excretory duct (Poiani and Cruz-Landim 2010). The secretion of the thoracic salivary gland is an aqueous solution of digestive enzymes that are involved in honey and sugar digestion as well as pollen and wax moistening (Simpson 1960, 1963). The adult cephalic salivary gland produces an oily substance involved in wax manipulation and mouthpart lubrication (Simpson et al. 1968). Salivary glands show a gradual increase in activity until their production peaks at about 15-25 days, after which they rapidly decline with the shift to guarding and foraging activities (Winston 1987).
Worker honey bees dissolve dry sugar, clean their queen’s body, and possibly also soften or lubricate materials being chewed, with the watery secretion of their labial glands. The small amount of oil produced by these glands accumulates in them and what little is discharged adheres to the tongue hairs and is not mixed with the food; its function is obscure (Simpson 1960).
The thoracic gland of the worker, the largest exocrine gland of the honey bee, was investigated by dissection, light microscopy, scanning and transmission electron microscopy (Schönitzer and Seifert 1990). The glands are paired and each lateral half consists of two parts, a smaller external and a larger internal lobe. The lobes are composed of densely packed secretory tubes and ducts, the tubes of which often show ramifications. A reservoir is packed within the anterior medial part of the gland. The secretory tubes are composed of two types of cells, secretory cells, which are most frequent, and parietal cells. Secretory cells are characterized by a basal labyrinth (an intricate structure of interconnecting passages), abundant rough endoplasmic reticulum, dark secretory vesicles, light vesicles of different sizes, and apical microvilli (microscopic cellular membrane protrusions that increase the surface area of cells). Parietal cells are smaller and have a characteristically lobed nucleus and no secretory vesicles. Between the cells there are intercellular canaliculi (a narrow canal or tubular passage). In the center of each tube there is an extracellular space with a central cuticular channel. The abundance of rough endoplasmic reticulum and the rare occurrence of smooth endoplasmic reticulum implies a saliva with proteins but rarely with pheromones. Between the secretory tubes there are frequently neuronal profiles which are partly in contact with the secretory cells. Thus, they demonstrate that this gland is controlled by the nervous system, in contrast to findings of previous investigators. The axonal endings contain dark neurosecretory vesicles as well as light synaptic vesicles. Large parts of the glands are surrounded by a thin tissue sheath which has a smooth surface towards the secretory tubes and shows irregular protrusions towards the outer side. This sheath is considered to be a tracheal air sac, and due to its large extension is probably of importance for the hemolymph flow through the thorax.
Fujita et al. (2010) performed a proteomic analysis of the honey bee salivary system. Although most (31-35) of the major proteins from the post-cerebral gland and thoracic gland were housekeeping proteins, the spot intensities for aldolase (enzyme that helps break down certain sugars to produce energy) and cetyl-CoA acyltransferase 2 (enzyme involved with fat metabolism) were stronger in the post-cerebral salivary glands than in the thoracic glands in the 2-dimensional gel electrophoresis. Immunoblotting confirmed that the expression of these proteins was stronger in the post-cerebral than in the thoracic glands, whereas expression was almost not detectable in the hypopharyngeal gland, suggesting that carbohydrate metabolism is enhanced in the post-cerebral salivary gland. In addition, imaginal disc growth factor 4 (IDGF4) was synthesized in the honey bee salivary system. Immunoblotting indicated IDGF4 expression was very strong in the post-cerebral gland, moderate in the thoracic gland and very weak in the hypopharyngeal gland. A considerable amount of IDGF4 was detected in royal jelly, while less was detected in honey, strongly suggesting that the salivary system secretes IDGF4 into royal jelly and honey. This growth factor might therefore affect the growth and physiology of the other colony members.
The molecular basis of how salivary glands fulfill their biological duty is not fully understood. Proteomics and phosphoproteomics of cephalic salivary glands and thoracic salivary glands were compared between normal and single-cohort honey bee colonies. Single-cohort colonies are artificially established colonies that are composed entirely of the same aged young bees, without appropriate aged nurses and foragers. This allows one to investigate the functional flexibility of salivary glands of workers by comparison with the same aged workers of their parent colonies (normal colonies). Of 113 and 64 differentially regulated proteins and phosphoproteins, 86 and 33 were identified, respectively. The salivary glands require a wide spectrum of proteins to support their multifaceted functions and ensure normal social management of the colony. Changes of protein expression and phosphoproteins are key role players (Feng et al. 2013). The post-cerebral salivary gland triggers labor transition from in-hive work to foraging activities via the regulation of juvenile hormone and ethyl oleate levels. The stronger expression of proteins involved in carbohydrate and energy metabolism, protein folding, protein metabolism, cellular homeostasis and cytoskeleton in the thoracic salivary gland, supports the gland to efficiently enhance honey processing by synthesis and secretion of saliva into nectar.
Figure 3. Alimentary canal of the honey bee larva. Silk Gland (Salivary Gland) (skGld), Malpighian tubules, proctodaeum (Proc), ventriculus (Vent), stomodaeum (Stom) and anus (An). (Snodgrass 1956).
The salivary secretions of adult females are functionally different from larvae. In the honey bee larva, the post-cerebral salivary glands are not yet developed. The thoracic salivary glands are represented by a pair of long, slender, tubes extending back to the sixth abdominal segment, which are also the larval silk glands (Snodgrass 1956). The silk glands run a convoluted course through the body beneath the gut and above the nerve cord and join together in the head to open at the spinneret situated on the combined labium and hypopharynx (Figure 3). These glands are the forerunners of the thoracic salivary glands of the adult, into which they develop during pupal metamorphosis (Morse and Hooper 1985). The silk glands of the bee larva attain their highest development just at the time the larva is ready to spin its cocoon. When finished, the glands rapidly degenerate during the first part of the pupal period: the cells lose their outlines and become irregular masses of vacuolated protoplasm, in which the nuclei break up into fragments that quickly degenerate (Snodgrass 1956).
In the salivary glands of the larval honey bee, silk is produced in the 5th larval instar (developmental stage between each molt) ( Silva-Zacarin et al. 2007). According to Silva and Silva de Moraes (2002) and Silva-Zacarin et al. (2003), at the beginning of the 5th instar, the salivary gland lumen (inside space of a tubular structure) is filled with a homogeneous secretion that is synthesized before silk production. In the mid-5th instar, these glands produce silk proteins that are released from the secretory cells as a homogeneous substance, which subsequently polymerizes in the lumen to form silk fibrils. By the end of the 5th instar, the gland lumen is filled with a compact fibrillar secretion. When the silk is released to spin the cocoon at the end of the 5th instar, the secretion that remains in the lumen loses its compaction and its macromolecular organization. Ultrastructural analysis of the larval salivary glands has revealed that after the peak of protein synthesis, the secretory portion of the gland undergoes histolysis (the breaking down of tissues) following construction of the cocoon, within the larva during metamorphosis.
Le Conte et al. (2006) found that the larval salivary glands (silk glands) have at least one additional function prior to silk production; the release of a brood pheromone composed of 10 fatty acid esters. This study showed that the salivary glands of the larvae can be considered the reservoir for the fatty acid esters that constitute brood pheromone. A blend of ten fatty acid methyl and ethyl esters produced by larvae have been shown to possess pheromone releaser effects, like the capping of the cells (Le Conte et al. 1994), and the recognition of the larval age and needs (Slessor et al. 2005). The esters also have primer effects stimulating hypopharyngeal glands of nurses (Mohammedi et al. 1996) or inhibiting ovary development of the workers (Mohammedi et al. 1998). The full blend of 10 esters (brood pheromone), modulates the behavioral development of young bees and stimulates workers for pollen foraging (Slessor et al. 2005).
No traces of esters were detected on the posterior part of the larvae. They were detected only in cuticular rinses of the anterior section of the larvae. Sections of the larval mouthparts of the head contained traces of the esters suggesting that ester secretion occurs at or near the mouthparts. The digestive tract and the salivary glands are the only two structures known to be connected to the mouth of the larvae. No esters were detected in extracts of the digestive tract. The salivary gland extracts contained significantly more ethyl and methyl esters than extracts from the rest of the larval body. The total amount of esters per larva was significantly different in the salivary glands (321±87 ng) compared to the rest of the larvae (87±23 ng). Dissection of larvae has revealed that the salivary gland is the reservoir for the brood pheromone esters (Leconte et al. 2006).
References
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Clarence Collison is an Emeritus Professor of Entomology and Department Head Emeritus of Entomology and Plant Pathology at Mississippi State University, Mississippi State, MS.
