Release Of Trehalose Research Articles

Enhanced adaptation to salinity stress in lentil seedlings through the use of trehalose-functionalized silica nanoparticles (TSiNPs): Exploring silica-sugar absorption and oxidative balance

Silica nanoparticles (SiNPs) confer better growth and development of plants under salinity stress. Moreover, the surface-functionalization of SiNPs with bioactive molecules is expected to enhance its efficacy. The present study thus aimed to modify the surface of SiNPs, by attaching a bioactive molecule (trehalose) to obtain TSiNPs. The successful surface functionalization was confirmed using FTIR, XRD, and EDS. The spherical shape and amorphous nature of the nanoparticles were confirmed using SEM. The TEM image analysis revealed that the size of SiNPs and TSiNPs ranged between 20-50 nm and 200–250 nm, respectively. A novel bioassay experiment designed to study the release of silica and trehalose from nanoparticles elucidated that the TSiNPs improved the release and uptake of silica. Also, trehalose uptake significantly improved after 72 h of application due to enhanced release of trehalose from TSiNPs. Further, this study also aimed to investigate the potential benefits of SiNPs and TSiNPs in promoting the growth and development of plants under salinity stress. In this context, the nanoparticles were applied to the saline-stressed (0, 200, 300 mM) lentil seedlings for the in-planta experiments. The results revealed that both SiNPs and TSiNPs improved the growth of seedlings (shoot, and root length), ionic balance (K+/Na+ ratio), and osmolyte status (sugars, proline, glycine betaine, trehalose). Additionally, increased antioxidant enzyme activities helped scavenge ROS (H2O2, O2.-) generated in NaCl-stressed seedlings, ultimately improving the membrane integrity (by reducing MDA and EL). However, the TSiNPs exhibited a much-enhanced activity in stress alleviation compared to the SiNPs.

Regulation of a Trehalose-Specific Facilitated Transporter (TRET) by Insulin and Adipokinetic Hormone in Rhodnius prolixus, a Vector of Chagas Disease.

Using the blood-sucking kissing bug, Rhodnius prolixus as an experimental model, we have studied the involvement of insulin-like peptides (ILPs) and adipokinetic hormone (AKH) signaling in carbohydrate metabolism, focusing on the regulation of the trehalose-specific facilitated transporter (Rhopr-TRET), particularly in the ovaries. We find that trehalose stores in ovaries increase after feeding, synchronously with the beginning of vitellogenesis, but that the transcript expression of enzymes involved in trehalose synthesis show no changes between unfed and blood-fed animals. However, an eightfold increase in Rhopr-TRET transcript expression is observed in the ovaries post-blood meal. In vivo and ex vivo assays using exogenous insulins and Rhopr-AKH, reveal that Rhopr-TRET is up-regulated in ovaries by both peptide families. In accordance with these results, when ILP and AKH signaling cascades are impaired using RNA interference, Rhopr-TRET transcript is down-regulated. In addition, trehalose injection induces an up-regulation of Rhopr-TRET transcript expression and suggests an activation of insulin signaling. Overall, the results support the hypothesis of a direct trehalose uptake by ovaries from the hemolymph through Rhopr-TRET, regulated by ILP and/or AKH. We also show that Rhopr-TRET may work cooperatively with AKH signaling to support the release of trehalose from the ovaries into the hemolymph during the unfed (starved) condition. In conclusion, the results indicate that in females of R. prolixus, trehalose metabolism and its hormonal regulation by ILP and AKH play critical roles in adapting to different nutritional conditions and physiological states.

D-glucose overflow metabolism in an evolutionary engineered high-performance D-xylose consuming Saccharomyces cerevisiae strain.

ABSTRACTCo-consumption of D-xylose and D-glucose by Saccharomyces cerevisiae is essential for cost-efficient cellulosic bioethanol production. There is a need for improved sugar conversion rates to minimize fermentation times. Previously, we have employed evolutionary engineering to enhance D-xylose transport and metabolism in the presence of D-glucose in a xylose-fermenting S. cerevisiae strain devoid of hexokinases. Re-introduction of Hxk2 in the high performance xylose-consuming strains restored D-glucose utilization during D-xylose/D-glucose co-metabolism, but at rates lower than the non-evolved strain. In the absence of D-xylose, D-glucose consumption was similar to the parental strain. The evolved strains accumulated trehalose-6-phosphate during sugar co-metabolism, and showed an increased expression of trehalose pathway genes. Upon the deletion of TSL1, trehalose-6-phosphate levels were decreased and D-glucose consumption and growth on mixed sugars was improved. The data suggest that D-glucose/D-xylose co-consumption in high-performance D-xylose consuming strains causes the glycolytic flux to saturate. Excess D-glucose is phosphorylated enters the trehalose pathway resulting in glucose recycling and energy dissipation, accumulation of trehalose-6-phosphate which inhibits the hexokinase activity, and release of trehalose into the medium.

Nanomedicine enables autophagy-enhanced cancer-cell ferroptosis

Ferroptosis and autophagy, playing significant roles in tumor treatment, are two typical forms of the programmed cell death. However, the rational combination of ferroptosis and autophagy for synergistic tumor therapy is still highly challenging. Herein, we report on an intriguing nanomedicine strategy for achieving autophagy-enhanced ferroptosis on efficiently combating cancer, which was based on the construction of trehalose-loaded mSiO2@MnOx-mPEG (TreMMM) nanoparticles with satisfactory biocompatibility. The nanoparticles are endowed with high glutathione (GSH) consumption efficiency, thereby inducing cancer-cell ferroptosis via inactivating glutathione peroxidases 4 (GPX4). Subsequently, the TreMMM degradation due to the GSH depletion and pH sensitivity contributed to the trehalose release for inducing autophagy, promoting/enhancing ferroptosis by NCOA4-mediated degradation of ferritin. A substantial in vitro and in vivo antitumor effect was achieved by such an intriguing autophagy-enhanced ferroptosis. Therefore, the rational combination of GSH-consumption-induced ferroptosis and trehalose-induced autophagy by nanomedicine design provides an alternative but effective strategy for tumor treatment.

Trehalose as an osmolyte in Candidatus Accumulibacter phosphatis

Candidatus Accumulibacter phosphatis is an important microorganism for enhanced biological phosphorus removal (EBPR). In a previous study, we found a remarkable flexibility regarding salinity, since this same microorganism could thrive in both freshwater- and seawater-based environments, but the mechanism for the tolerance to saline conditions remained unknown. Here, we identified and described the role of trehalose as an osmolyte in Ca. Accumulibacter phosphatis. A freshwater-adapted culture was exposed to a single batch cycle of hyperosmotic and hypo-osmotic shock, which led to the release of trehalose up to 5.34 mg trehalose/g volatile suspended solids (VSS). Long-term adaptation to 30% seawater-based medium in a sequencing batch reactor (SBR) gave a stable operation with complete anaerobic uptake of acetate and propionate along with phosphate release of 0.73 Pmol/Cmol, and complete aerobic uptake of phosphate. Microbial analysis showed Ca. Accumulibacter phosphatis clade I as the dominant organism in both the freshwater- and seawater-adapted cultures (> 90% presence). Exposure of the seawater-adapted culture to a single batch cycle of hyperosmotic incubation and hypo-osmotic shock led to an increase in trehalose release upon hypo-osmotic shock when higher salinity is used for the hyperosmotic incubation. Maximum trehalose release upon hypo-osmotic shock was achieved after hyperosmotic incubation with 3× salinity increase relative to the salinity in the SBR adaptation reactor, resulting in the release of 11.9 mg trehalose/g VSS. Genome analysis shows the possibility of Ca. Accumulibacter phosphatis to convert glycogen into trehalose by the presence of treX, treY, and treZ genes. Addition of trehalose to the reactor led to its consumption, both during anaerobic and aerobic phases. These results indicate the flexibility of the metabolism of Ca. Accumulibacter phosphatis towards variations in salinity.Key points• Trehalose is identified as an osmolyte in Candidatus Accumulibacter phosphatis.• Ca. Accumulibacter phosphatis can convert glycogen into trehalose.• Ca. Accumulibacter phosphatis clade I is present and active in both seawater and freshwater.

Trehalose-Rich, Degradable Hydrogels Designed for Trehalose Release under Physiologically Relevant Conditions

Trehalose, a natural disaccharide, is primarily known for its ability to protect proteins from inactivation and denaturation caused by a variety of stress conditions. Furthermore, over the past few years, it has emerged as a promising therapeutic candidate for treatment of neurodegenerative diseases. Herein, we examine the attachment of trehalose to polymers for release under selected physiologically relevant conditions. The proposed strategies are evaluated specifically using hydrogels undergoing simultaneous degradation during trehalose release. These materials are fabricated via copolymerization of the appropriate acrylamide-type monomers with polymerizable trehalose esters or benzylidene acetals. This provides trehalose release in a slightly alkaline (i.e., pH 7.4) or mildly acidic (i.e., pH 5.0) environment, respectively. Using this method materials containing up to 51.7 wt% of trehalose are obtained. The presented results provide a solid basis for future studies on polymeric materials intended for trehalose release in biological systems.

Cold-Responsive Nanoparticle Enables Intracellular Delivery and Rapid Release of Trehalose for Organic-Solvent-Free Cryopreservation.

Conventional cryopreservation of mammalian cells requires the use of toxic organic solvents (e.g., dimethyl sulfoxide) as cryoprotectants. Consequently, the cryopreserved cells must undergo a tedious washing procedure to remove the organic solvents for their further applications in cell-based medicine, and many of the precious cells may be lost or killed during the procedure. Trehalose has been explored as a nontoxic alternative to traditional cryoprotectants. However, mammalian cells do not synthesize trehalose or express trehalose transporters in their membranes, and the lack of an approach for the efficient intracellular delivery of trehalose has been a major hurdle for its use in cell cryopreservation. In this study, a cold-responsive polymer (poly(N-isopropylacrylamide-co-butyl acrylate)) is utilized to synthesize nanoparticles for the encapsulation and intracellular delivery of trehalose. The trehalose-laden nanoparticles can be efficiently taken up by mammalian cells. The nanoparticles quickly and irreversibly disassemble upon cold treatment, enabling the controlled and rapid release of trehalose from the nanoparticles inside cells. The latter is confirmed by an evident increase in cell volume upon cold treatment. This rapid cold-triggered intracellular release of trehalose is crucial to developing a fast protocol to cryopreserve cells using trehalose. Cells with intracellular trehalose delivered using the nanoparticles show comparable postcryopreservation viability compared to that of cells treated with DMSO, eliminating the need for the tedious and cell-damaging washing procedure required for using the DMSO-cryopreserved cells in vivo. This cold-responsive nanoparticle may greatly facilitate the use of trehalose as a nontoxic cryoprotectant for banking cells and tissues to meet their high demand by modern cell-based medicine.

High-efficiency expression of Sulfolobus acidocaldarius maltooligosyl trehalose trehalohydrolase in Escherichia coli through host strain and induction strategy optimization

Maltooligosyl trehalose trehalohydrolase (MTHase, EC 3.2.1.141) catalyzes the release of trehalose, a novel food ingredient, by splitting the α-1,4-glucosidic linkage adjacent to the α-1,1-glucosidic linkage of maltooligosyl trehalose. However, the high-yield preparation of recombinant MTHase has not yet been reported. In this study, a codon-optimized synthetic gene encoding Sulfolobus acidocaldarius MTHase was expressed in Escherichia coli. In initial expression experiments conducted using pET-24a (+) and E. coli BL21 (DE3), the MTHase activity was 10.4U/mL and a large amount of the expression product formed inclusion bodies. The familiar strategies, including addition of additives, co-expression with molecular chaperones, and expression with a fusion partner, failed to enhance soluble MTHase expression. Considering the intermolecular disulfide bond of MTHase, expression was investigated using a system comprising plasmid pET-32a (+) and host E. coli Origami (DE3), which is conducive to cytoplasmic disulfide bond formation. The MTHase activity increased to 55.0U/mL, a 5.3-fold increase. Optimization of the induction conditions in a 3-L fermentor showed that when the lactose was fed at 0.2g/L/h beginning at an OD600 of 40 and the induction temperature was maintained at 30°C, the MTHase activity reached a maximum of 204.6U/mL. This is the first report describing a systematic effort to obtain high-efficiency MTHase production. The high yield obtained using this process provides the basis for the industrial-scale production of trehalose. This report is also expected to be valuable in the production of other enzymes containing disulfide bonds.

Enhanced osteogenic activity and anti-inflammatory properties of Lenti-BMP-2-loaded TiO₂ nanotube layers fabricated by lyophilization following trehalose addition.

To enhance biocompatibility and osseointegration between titanium implants and surrounding bone tissue, numerous efforts have been made to modify the surface topography and composition of Ti implants. In this paper, Lenti-BMP-2-loaded TiO2 nanotube coatings were fabricated by lyophilization in the presence of trehalose to functionalize the surface. We characterized TiO2 nanotube layers in terms of the following: surface morphology; Lenti-BMP-2 and trehalose release; their ability to induce osteogenesis, proliferation, and anti-inflammation in vitro; and osseointegration in vivo. The anodized TiO2 nanotube surfaces exhibited an amorphous glassy matrix perpendicular to the Ti surface. Both Lenti-BMP-2 and trehalose showed sustained release over the course of 8 days. Results from real-time quantitative polymerase chain reaction studies demonstrated that lyophilized Lenti-BMP-2/TiO2 nanotubes constructed with trehalose (Lyo-Tre-Lenti-BMP-2) significantly promoted osteogenic differentiation of bone marrow stromal cells but not their proliferation. In addition, Lyo-Tre-Lenti-BMP-2 nanotubes effectively inhibited lipopolysaccharide-induced interleukin-1β and tumor necrosis factor-α production. In vivo, the formulation also promoted osseointegration. This study presents a promising new method for surface-modifying biomedical Ti-based implants to simultaneously enhance their osteogenic potential and anti-inflammatory properties, which can better satisfy clinical needs.

Interaction between Trehalose and MTHase from Sulfolobus solfataricus Studied by Theoretical Computation and Site-Directed Mutagenesis

Maltooligosyltrehalose trehalohydrolase (MTHase) catalyzes the release of trehalose by cleaving the α-1,4-glucosidic linkage next to the α-1,1-linked terminal disaccharide of maltooligosyltrehalose. Computer simulation using the hydrogen bond analysis, free energy decomposition, and computational alanine scanning were employed to investigate the interaction between maltooligosyltrehalose and the enzyme. The same residues that were chosen for theoretical investigation were also studied by site-directed mutagenesis and enzyme kinetic analysis. The importance of residues determined either experimentally or computed theoretically were in good accord with each other. It was found that residues Y155, D156, and W218 of subsites -2 and -3 of the enzyme might play an important role in interacting with the ligand. The theoretically constructed structure of the enzyme-ligand complex was further validated through an ab initio quantum chemical calculation using the Gaussian09 package. The activation energy computed from this latter study was very similar to those reported in literatures for the same type of hydrolysis reactions.

RNA interference unveils functions of the hypertrehalosemic hormone on cyclic fluctuation of hemolymph trehalose and oviposition in the virgin female Blattella germanica

Hypertrehalosemic hormone (HTH) is a neuropeptide within the adipokinetic hormone (AKH) family that induces a release of trehalose from fat body into hemolymph in a number of insects. In this study, we first showed that female adult German cockroach, Blattella germanica, displayed a cyclic fluctuation of hemolymph trehalose levels correlated to the maturation of oocytes in the reproductive cycle. After cloning the HTH cDNA from the German cockroach (Blage-HTH), expression studies indicated that Blage-HTH mRNA showed the cyclic changes during the first reproductive cycle, where peak values occurred in 8-day-old virgin female cockroaches, which were going to produce oothecae. The functions of Blage-HTH were studied using RNA interference (RNAi) to knockdown its expression. Adult virgin females of B. germanica injected with Blage-HTH dsRNA increased hemolymph trehalose levels in the late period of vitellogenesis more slowly than control. Furthermore, RNAi of Blage-HTH delayed oviposition time and some (10%) individuals did not produce the first ootheca until 15 days after eclosion, whereas the control group produced ootheca before 9 days in all cases.

Trehalose-recycling ABC transporter LpqY-SugA-SugB-SugC is essential for virulence of Mycobacterium tuberculosis

Mycobacterium tuberculosis (Mtb) is an exclusively human pathogen that proliferates within phagosomes of host phagocytes. Host lipids are believed to provide the major carbon and energy sources for Mtb, with only limited availability of carbohydrates. There is an apparent paradox because five putative carbohydrate uptake permeases are present in Mtb, but there are essentially no host carbohydrates inside phagosomes. Nevertheless, carbohydrate transporters have been implicated in Mtb pathogenesis, suggesting that acquisition of host sugars is important during some stages of infection. Here we show, however, that the LpqY-SugA-SugB-SugC ATP-binding cassette transporter is highly specific for uptake of the disaccharide trehalose, a sugar not present in mammals, thus refuting a role in nutrient acquisition from the host. Trehalose release is known to occur as a byproduct of the biosynthesis of the mycolic acid cell envelope by Mtb's antigen 85 complex. The antigen 85 complex constitutes a group of extracellular mycolyl transferases, which transfer the lipid moiety of the glycolipid trehalose monomycolate (TMM) to arabinogalactan or another molecule of TMM, yielding trehalose dimycolate. These reactions also lead to the concomitant extracellular release of the trehalose moiety of TMM. We found that the LpqY-SugA-SugB-SugC ATP-binding cassette transporter is a recycling system mediating the retrograde transport of released trehalose. Perturbations in trehalose recycling strongly impaired virulence of Mtb. This study reveals an unexpected accessory component involved in the formation of the mycolic acid cell envelope in mycobacteria and provides a previously unknown role for sugar transporters in bacterial pathogenesis.

The trehalose transporter 1 gene sequence is conserved in insects and encodes proteins with different kinetic properties involved in trehalose import into peripheral tissues

We recently cloned a trehalose transporter gene ( Tret1) that contributes to anhydrobiosis induction in the sleeping chironomid Polypedilum vanderplanki Hinton. Because trehalose is the main haemolymph sugar in most insects, they might possess Tret1 orthologs involved in maintaining haemolymph trehalose levels. We cloned Tret1 orthologs from four species in three insect orders. The similarities of the amino acid sequence to TRET1 in P. vanderplanki were 58.5–80.4%. Phylogenetic analysis suggested the Tret1 sequences were conserved in insects. The Xenopus oocyte expression system showed apparent differences in the K m and V max values for trehalose transport activity among the six proteins encoded by the corresponding orthologs. The TRET1 orthologs of Anopheles gambiae ( K m: 45.74 ± 3.58 mM) and Bombyx mori (71.58 ± 6.45 mM) showed low trehalose affinity, whereas those of Apis mellifera (9.42 ± 2.37 mM) and Drosophila melanogaster (10.94 ± 7.70 mM) showed high affinity. This difference in kinetics might be reflected in the haemolymph trehalose:glucose ratio of each species. Tret1 was expressed not only in the fat body but also in muscle and testis. These findings suggest that insect TRET1 is responsible for the release of trehalose from the fat body and the incorporation of trehalose into other tissues that require a carbon source, thereby regulating trehalose levels in the haemolymph.

The encapsulation and intracellular delivery of trehalose using a thermally responsivenanocapsule

The thermally responsive wall permeability of an empty core–shell structuredPluronic nanocapsule (together with its temperature dependent size andsurface charge) was successfully utilized for encapsulation, intracellulardelivery, and controlled release of trehalose, a highly hydrophilic small (MW = 342 D) molecule (a disaccharide of glucose) that is exceptional for long-termstabilization of biologicals (particularly at ambient temperatures). It was foundthat trehalose can be physically encapsulated in the nanocapsule using asoaking–freeze-drying–heating procedure. The nanocapsule is capable of physicallywithholding trehalose with negligible release in hours for cellular uptake at37 °C when its wall permeability is low. A quick release of the encapsulated sugarcan be achieved by thermally cycling the nanocapsule between 37 and22 °C (or lower). A significant amount of trehalose (up to 0.3 M) can be delivered into NIH 3T3fibroblasts by incubating the cells with the trehalose-encapsulated nanocapsules at37 °C for 40 min. Moreover, cytotoxicity of the nanocapsule for the purpose of intracellulardelivery of trehalose was found to be negligible. Altogether, the thermally responsivenanocapsule is effective for intracellular delivery of trehalose, which is critical for thelong-term stabilization of mammalian cells at ambient temperatures and the eventualsuccess of modern cell-based medicine.

Identification of the Essential Catalytic Residues and Selectivity-Related Residues of Maltooligosyltrehalose Trehalohydrolase from the Thermophilic Archaeon Sulfolobus solfataricus ATCC 35092

Maltooligosyltrehalose trehalohydrolase (MTHase) catalyzes the release of trehalose by cleaving the alpha-1,4-glucosidic linkage next to the alpha-1,1-linked terminal disaccharide of maltooligosyltrehalose. Mutations at residues D255, E286, and D380 were constructed to identify the essential catalytic residues of MTHase, while mutations at residues W218, A259, Y328, F355, and R356 were constructed to identify selectivity-related residues of the enzyme. The specific activities of the purified D255A, E286A, and D380A MTHases were only 0.15, 0.09 and 0.01%, respectively, of that of wild-type MTHase, suggesting that these three residues are essential catalytic residues. Compared with wild-type MTHase, A259S, Y328F, F355Y, and R356K MTHases had increased selectivity ratios, which were defined as the ratios of the catalytic efficiencies for glucose formation to those for trehalose formation in the hydrolysis of maltooligosaccharides and maltooligosyltrehaloses, respectively, while W218A and W218F MTHases had decreased selectivity ratios. When starch digestion was carried out at 75 degrees C and wild-type and mutant MTHases were, respectively, used with isoamylase and maltooligosyltrehalose synthase (MTSase), the ratios of initial rates of glucose formation to those of trehalose formation were inversely correlated to the peak trehalose yields.

A comparative study of the metabolic effects of hypertrehalosemic hormone and 1,2,3,4,5,6-hexachlorocyclohexane (γ-HCH) in the American cockroach, Periplaneta americana

Several effects of hypertrehalosemic hormone (Peram-HTH) have been compared with the action of 1,2,3,4,5,6-hexachlorocyclohexane (γ-HCH) in the cockroach, Periplaneta americana. The data show that both agents stimulate respiration while maintaining an R.Q. of approximately 1. The action of γ-HCH differs from that of Peram-HTH-I in that it promotes oxidation of large amounts of soluble carbohydrate. Recent studies show that Peram-HTH-I has an equally potent stimulatory effect on lipid deposition in the fat body and on the release of trehalose from the same tissue. It is of interest that γ-HCH not only increases triacylglycerol accumulation in the fatγ body but that this effect is more pronounced if the γ-HCH is applied to the ventral surface of the mesothorax. γ-HCH also depleted whole body glycoprotein and glycolipid whereas Peram-HTH-I had no effect on glycoprotein but increased that of glycolipid. It is clear that γ-HCH causes complex disruptions to the metabolism of the insect, many of which are related to the release of hormones. The study supports the idea that certain actions of γ-HCH are mediated through the release of Peram-HTH-I.

Hypertrehalosaemic neuropeptides decrease levels of the glycolytic signal fructose 2,6-bisphosphate in cockroach fat body

In cockroach fat body, trehalogenesis and glycolysis compete for glucose phosphates as common substrates. During trehalogenesis, glycolysis is inhibited, although the mechanism responsible for this is not known. Incubation of the isolated fat body from the Argentine cockroach Blaptica dubia with an extract of the corpora cardiaca containing as little as 0.005 gland equivalents ml-1 of incubation medium increased the release of trehalose (anthrone-positive material) from the tissue by more than 100 %. The content of the glycolytic activator fructose 2,6-bisphosphate was decreased by up to 50 %. A decapeptide was isolated from the corpora cardiaca of B. dubia and shown to be identical to the naturally occurring Blaberus discoidalis hypertrehalosaemic peptide (Bld HrTH), which was also found in the corpora cardiaca. Synthetic Bld HrTH at 2 nmol l-1 and above increased trehalose production and decreased the content of fructose 2,6-bisphosphate to the same extent as did corpus cardiacum extract. The octapeptides Periplaneta americana cardioacceleratory hormones I and II (Pea CAH-I and Pea CAH-II) also had a significant effect on both parameters. Fructose 2,6-bisphosphate is a potent activator of phosphofructokinase from cockroach fat body if the enzyme is assayed at near-physiological concentrations of substrates and effectors. It is suggested that, because of the decrease in fructose 2,6-bisphosphate levels in the fat body, the activity of the key glycolytic enzyme phosphofructokinase is diminished. This can explain the inhibition of glycolytic flux by hypertrehalosaemic peptides which alters the balance of glucose metabolism in favour of trehalose formation.

Evidence that free fatty acids in trophocytes of Periplaneta americana fat body may be regulated by the activity of phospholipase A 2 and cyclooxygenase

Previous studies have shown that palmitic, stearic, oleic and linoleic acid levels in trophocytes prepared from the fat body of male Periplaneta americana are increased following treatment of the cells with hypertrehalosemic hormone (HTH). Melittin, an activator of phospholipase A 2, mimicked the action of HTH by increasing the free fatty acid content in a concentration-dependent manner. The increase caused by HTH could be eliminated by pretreatment of the trophocytes with 1 mM 4′-bromophenacyl bromide (BPB), an inhibitor of phospholipase A 2. BPB also decreases the concentration of free fatty acids in trophocytes not treated with HTH but by a smaller margin. Nordihydroguaiaretic acid (NDGA) and indomethacin, inhibitors of lipoxygenase and cyclooxygenase, respectively, eliminated the increase in free fatty acids evoked by HTH. In the absence of HTH both inhibitors increased the free fatty acid content of the trophocytes, an effect consistent with the known mode of action of these agents. None of the inhibitors tested, all of which blocked HTH activated trehalose synthesis, prevented activation of phosphorylase by HTH. This is taken as evidence that other downstream sites are also important in the regulation of trehalose production by the fat body. It is suggested that the increase in free fatty acids evoked by HTH, or metabolites of those fatty acids, may regulate the synthesis and release of trehalose from the trophocytes because of potential effects on trehalose phosphate synthase, trehalose 6-phosphate phosphatase, and the trehalose transport mechanism in the trophocyte membrane.

Octopamine in insects: neurotransmitter, neurohormone, and neuromodulator

The biogenic amine octopamine is widely distributed within the nervous system of invertebrates. This review examines the role of octopamine in insects and shows that octopamine can act as a neurotransmitter, a neurohormone, and a neuromodulator. Examples of its neurotransmitter role are found in the firefly lantern and also possibly in the glandular lobe of the corpora cardiaca of locusts. Octopamine is also present in the haemolymph of cockroaches and locusts where it functions as a neurohormone, controlling the release of trehalose and lipid from fat body. In locusts, an identified octopaminergic neuron innervates the extensor-tibiae muscle of the hind legs. This neuron inhibits a myogenic rhythm of contraction and relaxation found in a proximal bundle of muscle fibers, and modulates the activity expressed by motoneurons which innervate the extensor-tibiae muscle. Octopamine is, therefore, a neuromodulator in this system. The functional significance of octopamine in insects is discussed with special reference to its association with general arousal phenomena.

Regulation of carbohydrate mobilization and utilization in Leucophaea maderae

Hormonal control of the phosphorylase system, metabolism of radioactive substrates, and trehalose synthesis were examined in the fat body. Fat body phosphorylase is maximally activated within 10 min after being exposed to corpus cardiacum extracts and is sensitive to very low concentrations of the extract. The concentration of phosphorylase-activating factor in the gland appears to be related to changes in glycogen content during oögenesis. The hormone elicits an accelerated rate of trehalose release, an increase in oxygen consumption, and a decrease in the respiratory quotient. The rate of oxidation of glycose to CO 2 is decreased, but that of acetate and palmitate are both increased in fat body incubated with corpus cardiacum extracts. A hypothesis considering the new metabolic effects of the hormone suggests that its regulatory action is effective at a number of different sites.