Activity of the prothoracic gland and its sensitivity to prothoracicotropic hormone in the penultimate and last-larval instar of Bombyx mori

The time course of secretion of ecdysone in vitro by the prothoracic glands of Bombyx mori was studied through the penultimate and last-larval instars. Ecdysone was produced by the glands in high amounts by the penultimate instar at 72 and 84 h while the glands in the last instar exhibited a high activity over 4 days around the time of gut purge and thereafter. The glands in the penultimate instar produced ecdysone at a low level throughout the instar before the sharp peak of activity, when they became inactive and remained so for the first 3 days of the last instar after when they regained secretory activity. Sensitivity of the glands to prothoracicotropic hormone varied in accord with the changes in their secretory activity. Inactive glands were not stimulated by 22K-prothoracicotropic hormone. In addition, glands with maximal activity in the penultimate instar were insensitive to 22K-prothoracicotropic hormone. These results suggest that the prothoracic glands in the penultimate and last-instar larvae are physiologically different.     

ROLE OF ADENOSINE 3'', 5''-MONOPHOSPHATE IN THE REGULATION OF CIRCADIAN OSCILLATION OF SEROTONIN N-ACETYLTRANSFERASE ACTIVITY IN CULTURED CHICKEN PINEAL GLAND

—When pineal glands of 10–12-day-old chicks were organ-cultured in darkness, serotonin N-acetyltransferase activity was low during the daytime, increased at midnight and then decreased to the daytime level the next morning. The pattern of increase and decrease of enzyme activity in cultured pineal glands was comparable to the circadian rhythm of N-acetyltransferase activity in vivo. When pineal glands were kept at a low temperature for 5 h prior to culture, the phase of autonomous rhythm of enzyme activity was delayed. When chicken pineal glands were cultured during the daytime for 6 h, derivatives of adenosine 3′, 5′-monophosphate (cyclic AMP), cholera toxin, a high concentration of KCl and phosphodiesterase inhibitors increased N-acetyltransferase activity 3–7-fold, indicating an involvement of cyclic AMP in the regulation of N-acetyltransferase activity in chicken pineal gland as has been shown in rat pineal gland. When pineal glands were cultured at night in darkness, cholera toxin or a high KCl did not enhance the night-time increase of the enzyme activity. Derivatives of cyclic AMP or phosphodiesterase inhibitors enhanced the autonomous night-time increase of N-acetyltransferase activity in an additive or more than additive manner in cultured pineal glands. These observations suggest that adenylate cyclase of pinealocytes is inactive during daytime, but is activated at night in darkness, which is transduced to the synthesis of N-acetyltransferase molecules. Catecholamines suppressed the basal level and the nocturnal increase of N-acetyltransferase activity via α-adrenergic receptor. The nocturnal increase of enzyme activity was prevented by cycloheximide or actinomycin D. Cocaine, which stabilizes cell membrane potential or light exposure, blocked the nighttime increase of N-acetyltransferase activity in cultured chicken pineal glands.     

Characteristics of sialidase in the rat salivary glands

Using 4-methylumbelliferyl-N-acetylneuraminic acid (4MU-NeuAc) as substrate, we measured sialidase activity in the salivary glands and other organs of the rat. The pH optima of salivary gland sialidase were between 4.0 and 4.5, which were similar to those of the enzyme in the brain, liver and kidney. Among the salivary glands, the submandibular one showed the highest sialidase activity followed by the parotid and the sublingual glands. However, sialidase activity in these glands was lower when compared with the activity in the brain, liver and kidney. From the subcellular distribution study, salivary gland sialidase was found to be mainly localized in the lysosomes. The pH optima of the lysosomal sialidase of the salivary glands were between 4.0 and 4.5; and Km values for 4MU-NeuAc approximately 0.09 mmol/l. In the submandibular and parotid glands, a soluble sialidase with a different pH optimum (5.5) and Km value (0.25 mmol/l) was also detected.     

Secretory behaviour of hypertrophic and hyperplastic salivary gland

Synopsis The enzyme content and the secretory behaviour of normal rat salivary glands were compared with these properties in glands made hypertrophic and hyperplastic by the chronic administration of isoproterenol. The enlarged glands displayed reductions in the concentrations of ribonuclease, deoxyribonuclease and amylase. The secretory behaviourin vivo was similar for all enzymes in both types of glands, but the enlarged glands secreted a lower percentage of their contentin vitro. The reduction in amylase activity was shown by immunological techniques to be due to a reduction in the number of enzyme molecules. The reduction in ribonuclease activity was not due to changes in the level of ribonuclease inhibitors.     

Alteration of Ca2+‐ATPase activity in the homogenate,plasma membrane and microsomes of the salivary glands of streptozotocin‐induced diabetic rats

Diabetes has been implicated in the dryness of the mouth, loss of taste sensation, sialosis, and other disorders of the oral cavity, by impairment of the salivary glands. The aim of the present study was to examine the plasma membrane, microsomal, and homogenate Ca²⁺‐ATPase activity in the rat submandibular and parotid salivary glands of streptozotocin‐induced diabetes. We have also examined the influence of the acidosis state on this parameter. Diabetes was induced by an intraperitoneal injection of streptozotocin and acidosis was induced by daily injection of NH₄Cl. At 15 and 30 days after diabetes induction, the animals were euthanized and the submandibular and parotid salivary glands were removed and analyzed. Ca²⁺‐ATPase (total, independent, and dependent) was determined in the homogenate, microsomal, and plasma membranes of the salivary glands of diabetic and control rats. Calcium concentration was also determined in the glands and showed to be higher in the diabetic animals. Ca²⁺‐ATPase activity was found to be reduced in all cell fractions studied in the diabetic animals compared with control. Similar results were obtained for the submandibular salivary glands of acidotic animals; however in the parotid salivary glands it was found an increase in the enzyme activity. Copyright © 2009 John Wiley & Sons, Ltd.     

Developmental changes in the activities of prolinase and prolidase in rat salivary glands,and the effect of thyroxine administration

Summary As the salivary glands are interesting tissues to study proliferation, we studied the activities of prolinase and prolidase using Pro-Ala and Pro-Hyp as substrates, respectively, in developing rat salivary glands between day 1 and week 10 after birth. Developmental changes of prolinase activity in the submandibular and sublingual glands were similar to those in the parotid gland, which steadily increased and reached the adult level by 20–25 days after birth. However, the changes in the activity of prolidase in the submandibular and sublingual glands were different from those in the parotid gland: the activity in the parotid gland slowly increased with maturation and reached a maximum level on day 30, but the activity in the submandibular and sublingual glands continuously increased with maturation. When thyroxine was injected every two days from day 1 to day 19, both enzyme activities were induced precociously in the parotid gland but not in the submandibular and sublingual glands. On the study of regional distribution in rat tissues, the correlation coefficient between prolinase and prolidase activities was high in the peripheral but not high in the brain regions.These results indicate that the physiological roles of prolinase and prolidase are very similar but not the same.     

Immunohistochemical and ultrastructural investigation of acinar cells in submandibular and sublingual glands of rats fed a liquid diet

In atrophic parotid glands induced by liquid diet, acinar cell apoptosis is increased while proliferative activity is reduced. This study aimed to clarify how liquid diet affects submandibular and sublingual glands, including acinar cell apoptosis and proliferation. Seven-week-old male Wistar rats were fed either a liquid (experimental group) or pellet diet (control group) from 3 to 21 days, respectively. Submandibular and sublingual glands were weighed and examined histologically, ultrastructurally, and immunohistochemically using antibodies to cleaved caspase-3 (Casp-3) and 5-bromo-2′-deoxyuridine (BrdU). Weights of submandibular and sublingual gland from the experimental group were not significantly different from controls at any time point. Histological and ultrastructural characteristics of experimental acinar cells in both glands were normal. Acinar cells in control and experimental submandibular glands were positively stained with periodic acid Schiff (PAS) and weakly stained by alcian blue (AB). In control and experimental sublingual glands, mucous acinar cells were PAS-positive and strongly AB-positive. Although Casp-3- and BrdU-positive acinar cells were identified in both glands in the experimental group, their labeling indices were not significantly different from controls. In conclusion, liquid diet in rats does not induce atrophic alterations to acinar cells, including apoptosis and proliferative activity in submandibular and sublingual glands.     

Effects of some inhibitors of DNA synthesis and repair upon the cycle of serotonin N-acetyltransferase activity in cultured chick pineal glands

When chick pineal glands were cultured in the dark with aphidicolin from midphotoperiod, the increase of serotonin N-acetyltransferase (NAT) activity was stimulated and the time of peak NAT activity was advanced. The peak level of NAT activity was also reached sooner on the 2nd day of culture. The increase of NAT activity was also stimulated in glands cultured under diurnal illumination, but the time of peak activity was not advanced. Effects with glands explanted into culture in the dark at other times were smaller and the time of peak NAT activity was not changed. Cytosine arabinoside and dideoxythymidine also stimulated the increase of NAT activity and advanced the time of peak activity with glands cultured in the dark from midphotoperiod. 3-Aminobenzamide markedly stimulated the increase of NAT activity both in the dark and under diurnal lighting when pineal glands were explanted into culture at mid- or late photoperiod. In contrast, with glands in culture from earlier in the photoperiod, aminobenzamide had no effect upon the increase of NAT activity up to the peak level found with control glands. Thereafter results were variable. Effects of cordycepin upon development of NAT activity were similar to those of 3-aminobenzamide but less marked. Incorporation of thymidine into acid-insoluble material in the dark was very markedly inhibited by aphidicolin, cytosine arabinoside, and dideoxythymidine, but only slightly by cordycepin. Aminobenzamide strongly inhibited incorporation by glands cultured from midphotoperiod, but had little effect with glands in culture from near the end of the photoperiod. We adopt the working hypothesis that excision repair of DNA may be a major component in the mechanism of the chick pineal clock.     

Melatonin in the lacrimal gland: first demonstration and experimental manipulation

NAT, HIOMT and melatonin are described in the extra-orbital lacrimal glands. The extra-orbital lacrimal glands of female Syrian hamsters contain higher NAT activity and melatonin levels than those in male glands, while male glands have higher HIOMT activity. Castration did not change melatonin in the lacrimal glands, although NAT and HIOMT activities were altered. The exposure of female hamsters to light in the morning (0600h) was associated with a reduction in both NAT activity and melatonin levels. Porphyrins were not detected in the lacrimal glands of either male or female hamsters.     

Changes in dopamine-sensitive adenylate cyclase activity in salivary glands of female lone star ticks,Amblyomma americanum (L.), during feeding

The activity of dopamine-sensitive adenylate cyclase in feeding female ixodid tick salivary glands was dependent on the state of tick feeding. Activity was significantly greater than the “basal” activity in salivary glands from ticks at all stages of tick feeding. Enzyme activity was not detected in the glands of unfed females. Enzyme activity reached a peak in glands of ticks weighing approx. 200 mg then declined as ticks increased in weight beyond 200 mg to repletion. Replete ticks (detached from the host for 12–24 hr) had similar levels of basal and dopamine-stimulated adenylate cyclase activity as that measured in salivary glands of high weight (>200 mg) ticks. Enzyme activity was 19–62% less in glands from ticks feeding on hosts that had been parasitized 2–4 months earlier by lone star ticks.     

Protein metabolism by the salivary glands and other organs of the larva of the blowfly, Calliphora erythrocephala

Protein metabolism in salivary glands, gut, haemolymph, and fat body during the last larval instar of the blowfly, Calliphora erythrocephala, has been investigated. In salivary glands, protein release, protein synthesis, amylase, and pepsin-like protease activity were maximal in 6 day larvae, this being at a time when the larvae had finished feeding. All these functions declined in glands from the rounded-off white puparial stage (R.O.) while acid phosphatase activity rose throughout the third instar to a maximum at the R.O. stage, Glands from 6 and 7 day larvae released protein which on disk gel electrophoresis separated into four minor bands and two major bands one of the latter possessing protease activity.In the gut, pepsin-like protease activity was maximal in 4 day larvae after which it fell rapidly thus following the feeding pattern of the larva in contrast to that in the salivary glands which did not.In vitro experiments showed that protease was released from 6 day glands through the basal membrane of the cells and not via the duct. A pepsin-like protease was also found in the haemolymph and fat body, the activity in the fat body rising rapidly during the latter part of the third instar, a rise which is attributed to the fat body sequestering protease from the haemolymph. Acid phosphatase activity in the fat body was maximal in 5 day larvae indicating that this enzyme was synthesized early in the third instar. It was shown that fat body sequestered ¹⁴C-labelled protein synthesized by and released from the salivary glands, most of the ¹⁴C activity being associated with a 600 g precipitable, acid-phosphatase rich fraction.It is proposed that in late third instar larvae the salivary glands function as glands of internal secretion, releasing protease into the haemolymph, which is then sequestered by the fat body (and perhaps other tissues) and is subsequently used in the lysis of the tissues at the time of metamorphosis.     

Femoral glands of the lizard, Crotaphytus collaris

The anatomy of the femoral glands in an iguanid lizard, Crotaphytus collaris collaris (Say), is described. The 48 lizards (including three embryos) from which glands were examined were obtained throughout their season of activity at one locality in Kansas. In animals of both sexes the glands lie in a linear series on the ventral aspect of each thigh. They are composed of branching tubes and tubules of epidermal and dermal origin. The row of femoral pores is the only external manifestation of the glands. Post hatching, the glands of males increase in size and complexity; little onto-genetic change occurs in the glands of females. The relative length of the glands appears to vary seasonally in adult males suggesting variation in their activity. The greatest relative sizes occur in the breeding season. At times a stratum corneum, continuous with the stratum corneum of the skin, occurs in the duct of the gland internal to part of the secretion plug. Formation of the stratum corneum seems to be initiated in the autumn prior to hibernation, and the stratum corneum removes the outer part of the secretion plug in the next ecdysis; meanwhile, production of a new secretion plug is initiated. The anatomy of the femoral glands in Crotaphytus is similar to that of the described glands of other species of lizards.     

A cytochemical study of the leaf-gland enzymes of insectivorous plants of the genus Pinguicula

Summary Cytochemical methods have been used to study the distribution of acid phosphatase, esterase, ribonuclease, amylase and protease activity in the stimulated and unstimulated leaf glands of Pinguicula grandiflora, P. vulgaris, P. lusitanica, and P. caudata. Two gland types are present, stalked and sessile. The stalked glands bear a muco-polysaccharide secretion droplet, and are concerned with capture of the prey; the sessile glands are specialised for digestion. In unstimulated glands of both classes, acid phosphatase, esterase and ribonuclease activity is associated with the anticlinal walls of the head cells, which have a characteristic spongy inner surface, comparable with that of transfer cells. Acid phosphatase and esterase activity was also detected in the vacuoles of the head cells of the sessile glands. Substrate film tests showed that amylase is readily released from the stalked glands but not from the sessile ones, while in contrast proteolytic activity is mainly associated with the sessile glands.On stimulation by suitable nitrogenous materials, the glands begin to sectete fluid onto the leaf surface within 1 hr. During the process the enzymes held in the spongy walls are discharged, and activity is also lost from the intracellular sites in the sessile glands.Digestion on the leaf surface and resorption of the products has been followed autoradiographically after feeding of ¹⁴C-labelled protein. Within 2 hr, digestion products enter the leaf, and move towards the margin in the vascular system. Movement out of the leaf begins within 12 hr. Microautoradiographs showed a concentration of products around the bases of the sessile glands and in the cells of the gland head, showing that these glands are involved in resorption as well as secretion.A possible mechanism of gland function is discussed.     

Structural Aspects of the Salt Glands of the Plumbaginaceae

Faraday, C. D. and Thomson, W. W. 1986. Structural aspects ofthe salt glands of the Plumbaginaceae.—J. exp. Bot. 37:461–470. The epidermal salt-secreting glands of 11 species from six differentgenera within the Plumbaginaceae were examined Gland ultrastructurewas considered with respect to species, secretory activity,and secretory product. All mature glands had a similar ultrastructure.Cytoplasmically dense secretory cells contained a full complementof organelles and structures which included numerous mitochondriaand few plastids. Reconstruction of serial paradermal sectionsthrough entire glands revealed that each gland cell generallycontained one or two vacuoles with a convoluted tonoplast inboth secreting and non-secreting states. The absence of numerousvacuoles and vesicles during secretory activity suggested thation secretion was by a transmembrane pathway rather than bya vesicle-mediated pathway. Key words: Salt glands, ultrastructure, Plumbaginaceae     

Phase-Shifting of Cycles in Level of Serotonin N-Acetyltransferase Activity and Cyclic GMP Content of Cultured Chick Pineal Glands by Methotrexate

Methotrexate at 1 microM stimulated increase of serotonin N-acetyltransferase (NAT) activity in chick pineal glands cultured under each of three conditions of illumination. The peak of the circadian rhythm in NAT activity and the "spike" in content of cyclic GMP were both advanced in pineal glands cultured in the dark from midphotoperiod. In contrast, the time of peak NAT activity in glands cultured in the dark from late photoperiod was unaffected. In addition, methotrexate did not affect times of reaching maximum NAT activities in glands cultured from midphotoperiod in the light or under diurnal illumination. Doubling the concentration of methotrexate also eliminated the lag phase in increase of NAT activity in glands cultured in the dark. However, at a concentration of 5 microM methotrexate the curve depicting increase of NAT activity was biphasic, and neither time nor level of peak NAT activity differed from those of control glands. Results of attempts to demonstrate persistent effects of exposure to methotrexate were inconclusive.     

Structure-activity relationships in corpora allata of the cockroach Diploptera punctata: roles of mating and the ovary

Morphometric studies were made on corpora allata of the cockroach Diploptera punctata from animals in which increasing gland size is not coupled to hormone synthesis (ovariectomized mated females; last-instar larvae) and in which gland size is coupled to hormone synthesis (normal mated and virgin females; penultimate-instar larvae). Cell number, gland volume, and juvenile hormone synthesis were measured. From electron micrographs, nuclear, cytoplasmic, and extracellular volumes; and cell membrane area were calculated; and fine structure described. Low-activity glands of ovariectomized mated females resembled high-activity glands from mated females in high cell number, large overall and cytoplasmic volume, and low nuclear-cytoplasmic ratio; they differed in having organelles typical of low-activity glands, mitochondria with dense matrices and large whorls of smooth endoplasmic reticulum. Inactive lastinstar larval glands resembled mated ovariectomized, female glands in increased cell number and organelles characteristic of inactive glands; however, their nuclearcytoplasmic volume ratio was much higher. Penultimate cytoplasmic volume ratio was much higher. Penultimate larval glands with high activity per cell resembled active glands of normal mated females. Ovariectomy did not change morphometric parameters of virgin female glands; thus mating results in increase in size of adult female glands whereas the growing ovary is needed for changes in mitochondria and endoplasmic reticulum associated with high juvenile hormone synthesis.     

Evidence for amylase in avian salivary glands

Four glands of the house sparrow, chicken and turkey were examined histologically and for their content of amylase. These were the external and intermediate mandibular glands, the maxillary gland and glandula anguli oris of the sparrow and the anterior and posterior mandibular, maxillary and anguli oris glands of the chicken and turkey. Amylase was determined by a starch substrate slide method and by biochemical assay. General morphology and mucopolysaccharide staining are described. All four glands of the sparrow demonstrated significant amylolytic activity by the assay. In the external mandibular and anguli oris glands this activity could be traced to mucous and seromucous cells of origin by means of the starch substrate slide procedure. None of the glands of the chicken or turkey displayed significant amylolytic activity.     

Activation of prothoracic glands by brains in vitro

The effects of brains from both diapausing and non-diapausing Mamestra brassicae pupae on the prothoracic glands from pupae of the same condition were studied by observations of the morphological changes and bioassay of the prothoracic glands in vitro.It was ascertained that the active brains intensified the hormonal activity of prothoracic glands from younger diapausing pupae more than those from older pupae. Further, these results coincided with the fact that the prothoracic glands from brainless pupae were more difficult to activate by active brains the longer the time after the glands had been extirpated.The brains from both younger and older diapausing M. brassicae pupae were able to activate co-cultured inactive prothoracic glands in vitro. These results suggest that even the brain from diapausing pupae of M. brassicae can synthesize and release the prothoracic gland activating hormone in vitro.     

Input and output signals in a model neural system: The regulation of melatonin production in the pineal gland

Summary The production of melatonin has been studied using organ cultures of pineal glands incubated with methionine-methyl-³H. Melatonin-O-methyl-³H was extracted from cultured pineal glands and incubation media, and the activity of N-acetyltransferase was measured. This is the first of two enzymes necessary for the conversion of serotonin to melatonin in the pineal. The treatment of pineal glands with norepinephrine or dibutyryl cyclic AMP increased the release of melatonin-O-methyl-³H into the incubation media and the concentration of melatonin-O-methyl-³H in the glands. These treatments also resulted in the stimulation of N-acetyltransferase, as compared to untreated glands. The transduction of neural information to biochemical, signals which regulate the melatonin pathway appears to involve the release of norepinephrine, which stimulates N-acetyltransferase activity through an adenyl cyclase-cyclic AMP mechanism, as evidenced by these and other studies discussed. In the present study the effects of harmine were studied. This hallucinogen is known to inhibit monoamine oxidase and stimulate melatonin production. Harmine was observed to stimulate N-acetyltransferase. This observation raises the possibility that an important action of this psychotropic drug may be on mechanisms which convert neural activity into biochemical events.