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1749-6632 (Electronic) 0077-8923; The evolution of communication is a fundamental biological problem. The genetic control of the signal and its reception must be tightly coadapted, especially in interindividual sexual communication. However, there is very little experimental evidence for tight genetic linkage connecting the emission of a signal and its reception. In Drosophila melanogaster, desat1 is the first known gene that simultaneously affects the emission and the perception of sex pheromones. Our experiments show that both aspects of pheromonal communication (the emission and the perception of sex pheromones) depend on distinct genetic control and may result from tissue-specific expression of different transcripts, all coding for the same desaturase. Therefore, and given the high conservation of its coding region, the pleiotropic activity of the desat1 gene may have arisen from an evolutionary process that shaped its regulatory regions.

0012-1606 (Print) Journal Article Research Support, Non-U.S. Gov't; Drosophila thoracic muscles are comprised of both direct flight muscles (DFMs) and indirect flight muscles (IFMs). The IFMs can be further subdivided into dorsolongitudinal muscles (DLMs) and dorsoventral muscles (DVMs). The correct patterning of each category of muscles requires the coordination of specific executive regulatory programs. DFM development requires key regulatory genes such as cut (ct) and apterous (ap), whereas IFM development requires vestigial (vg). Using a new vg(null) mutant, we report that a total absence of vg leads to DLM degeneration through an apoptotic process and to a total absence of DVMs in the adult. We show that vg and scalloped (sd), the only known VG transcriptional coactivator, are coexpressed during IFM development. Moreover, we observed an ectopic expression of ct and ap, two markers of DFM development, in developing IFMs of vg(null) pupae. In addition, in vg(null) adult flies, degenerating DLMs express twist (twi) ectopically. We provide evidence that ap ectopic expression can induce per se ectopic twi expression and muscle degeneration. All these data seem to indicate that, in the absence of vg, the IFM developmental program switches into the DFM developmental program. Moreover, we were able to rescue the muscle phenotype of vg(null) flies by using the activity of ap promoter to drive VG expression. Thus, vg appears to be a key regulatory gene of IFM development.

Overexpression of the UAS-tra transgene in Drosophila melanogaster females led to the complete elimination of their cuticular pheromones. According to current models of Drosophila behavior, these flies should induce no courtship. In fact, they are still attractive to conspecific males. Three classes of stimuli are shown to induce courtship, with different effects on male behavior: ( i ) known pheromones produced by control females, ( ii ) stimuli produced by living control and transgenic flies, and ( iii ) as-yet-undetermined pheromones present on both control and transgenic flies. Only the latter class of pheromones are required for mating. They appear to represent a layer of ancestral attractive substances present in D. melanogaster and its sibling species; known cuticular pheromones modulate this attractivity positively or negatively. The absence of inhibitory pheromones leads to high levels of interspecific mating, suggesting an important role for these cuticular hydrocarbons in isolation between species.

0965-1748 (Print) Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, Non-P.H.S.; Proteins of the third instar larval cuticle of Drosophila melanogaster, LCP5-LCP9, were purified and their N-terminal sequences determined. Three of these proteins (LCP5, 6, and 8) were found to be encoded by two multicopy genes previously mapped to the gene cluster at 65A 5-6 on the left arm of the third chromosome. The analysis of the patterns of developmental expression of the 8 distinct genes at this site showed that all but two were expressed during larval life. The patterns fell into three groups: one where expression was all through larval life, one where expression was primarily in the third instar, and one only during the production of the adult cuticle. One duplicated gene was not expressed in the Canton S strain at any time from the embryo to adult ecdysis. These findings indicate that there is not a unique set of cuticle proteins in the third versus the first and second instar larval cuticles and indicates that overlapping gene sets in several different gene clusters encode the proteins of the cuticle of different developmental stages.

0167-7063 (Print) Journal Article Research Support, Non-U.S. Gov't; Sites in the brain that show functional, sexual dimorphism in courtship behavior have been mapped at high resolution in male/female mosaics of Drosophila melanogaster. The sex mosaics were produced by enhancer-trap expression of GAL4 driving the female-spliced form of the transformer gene (tra), revealing sites in the dorsal brain, lateral protocerebrum, suboesophageal, thoracic and abdominal ganglia, and suggesting the importance of cross-talk between these regions in the implementation of the courtship sequence.

Courtship dominance in male Drosophila occurs when a male directs high levels of courtship towards another male, who remains passive [1]. We investigated the cues that shape this effect and report here that it is induced by the perception of adult male cuticular hydrocarbons during a critical period.

International audience; Amitermes evuncifer (Termitidae, Termitinae) is a pest termite in several countries of Africa, damaging crops and buildings. The trail-following pheromone of this termite was isolated from worker sternal glands and identified by GC-MS after solid phase microextraction (SPME). The pheromone is composed of (3Z,67,8E)-dodeca-3,6,8-trien-1-ol (dodecatrienol) and neocembrene acting in synergy, neocembrene being 5-10 times more abundant than dodecatrienol. Dodecatrienol alone is able to elicit trail-following in workers, whereas neocembrene alone is inactive. However, the association of both components induces a significantly higher trail-following activity. Multicomponent trail-following pheromones were only known, until now, in the Termitidae Nasutitermitinae, and in the Rhinotermitidae Prorhinotermes.

1749-6632 (Electronic) 0077-8923 (Linking) Journal Article Review

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OBJECTIVE—Insulin secretion involves complex events in which the mitochondria play a pivotal role in the generation of signals that couple glucose detection to insulin secretion. Studies on the mitochondrial generation of reactive oxygen species (ROS) generally focus on chronic nutrient exposure. Here, we investigate whether transient mitochondrial ROS production linked to glucose-induced increased respiration might act as a signal for monitoring insulin secretion. RESEARCH DESIGN AND METHODS—ROS production in response to glucose was investigated in freshly isolated rat islets. ROS effects were studied using a pharmacological approach and calcium imaging. RESULTS—Transient glucose increase from 5.5 to 16.7 mmol/l stimulated ROS generation, which was reversed by antioxidants. Insulin secretion was dose dependently blunted by antioxidants and highly correlated with ROS levels. The incapacity of β-cells to secrete insulin in response to glucose with antioxidants was associated with a decrease in ROS production and in contrast to the maintenance of high levels of ATP and NADH. Then, we investigated the mitochondrial origin of ROS (mROS) as the triggering signal. Insulin release was mimicked by the mitochondrial-complex blockers, antimycin and rotenone, that generate mROS. The adding of antioxidants to mitochondrial blockers or to glucose was used to lower mROS reversed insulin secretion. Finally, calcium imaging on perifused islets using glucose stimulation or mitochondrial blockers revealed that calcium mobilization was completely reversed using the antioxidant trolox and that it was of extracellular origin. No toxic effects were present using these pharmacological approaches. CONCLUSIONS—Altogether, these complementary results demonstrate that mROS production is a necessary stimulus for glucose-induced insulin secretion.

1939-327X (Electronic) Journal article; Objective : Recent data demonstrate that glucose sensing in different tissues is initiated by an intracellular redox-signaling pathway in physiological conditions. However, the relevance of such a mechanism in metabolic disease is not known. The aim of the present study was to determine whether brain-glucose hypersensitivity present in obese Zucker rat is related to an alteration in redox signaling. Research design and Methods: Brain glucose sensing alteration was investigated in vivo through the evaluation of electrical activity in arcuate nucleus, changes in ROS levels, and hypothalamic glucose-induced insulin secretion. In basal conditions, modifications of redox state and mitochondrial function were assessed through oxidized glutathione, glutathione peroxidase, manganese superoxide dismutase, aconitase activities and mitochondrial respiration. Results : Hypothalamic hypersensitivity to glucose was characterized by enhanced electrical activity of the arcuate nucleus and increased insulin secretion at a low glucose concentration, which does not produce such an effect in normal rats. It was associated with 1) increased ROS levels in response to this low glucose load, 2) constitutive oxidized environment coupled with lower antioxidant enzyme activity at both the cellular and mitochondrial level, and 3) over-expression of several mitochondrial subunits of the respiratory chain coupled with a global dysfunction in mitochondrial activity. Moreover, pharmacological restoration of the glutathione hypothalamic redox state by reduced-glutathione infusion in the third ventricle fully reversed the cerebral hypersensitivity to glucose. Conclusions : Altogether, these data demonstrate that obese Zucker rats' impaired hypothalamic regulation in terms of glucose sensing is linked to an abnormal redox signaling, which originates from mitochondria dysfunction.

0363-6143 (Print) Comparative Study In Vitro Journal Article Research Support, N.I.H., Extramural; Fasting increases neuropeptide Y (NPY) expression, peptide levels, and the excitability of NPY-expressing neurons in the hypothalamic arcuate (ARC) nucleus. A subpopulation of ARC-NPY neurons ( approximately 40%) are glucose-inhibited (GI)-type glucose-sensing neurons. Hence, they depolarize in response to decreased glucose. Because fasting enhances NPY neurotransmission, we propose that during fasting, GI neurons depolarize in response to smaller decreases in glucose. This increased excitation in response to glucose decreases would increase NPY-GI neuronal excitability and enhance NPY neurotransmission. Using an in vitro hypothalamic explant system, we show that fasting enhances NPY release in response to decreased glucose concentration. By measuring relative changes in membrane potential using a membrane potential-sensitive dye, we demonstrate that during fasting, a smaller decrease in glucose depolarizes NPY-GI neurons. Furthermore, incubation in low (0.7 mM) glucose enhanced while leptin (10 nM) blocked depolarization of GI neurons in response to decreased glucose. Fasting, leptin, and glucose-induced changes in NPY-GI neuron glucose sensing were mediated by 5'-AMP-activated protein kinase (AMPK). We conclude that during energy sufficiency, leptin reduces the ability of NPY-GI neurons to sense decreased glucose. However, after a fast, decreased leptin and glucose activate AMPK in NPY-GI neurons. As a result, NPY-GI neurons become depolarized in response to smaller glucose fluctuations. Increased excitation of NPY-GI neurons enhances NPY release. NPY, in turn, shifts energy homeostasis toward increased food intake and decreased energy expenditure to restore energy balance.

1939-327X (Electronic) Journal Article Research Support, Non-U.S. Gov't; OBJECTIVE: Insulin plays an important role in the hypothalamic control of energy balance, especially by reducing food intake. Emerging data point to a pivotal role of reactive oxygen species (ROS) in energy homeostasis regulation, but their involvement in the anorexigenic effect of insulin is unknown. Furthermore, ROS signal derived from NADPH oxidase activation is required for physiological insulin effects in peripheral cells. In this study, we investigated the involvement of hypothalamic ROS and NADPH oxidase in the feeding behavior regulation by insulin. RESEARCH DESIGN AND METHODS: We first measured hypothalamic ROS levels and food intake after acute intracerebroventricular injection of insulin. Second, effect of pretreatment with a ROS scavenger or an NADPH oxidase inhibitor was evaluated. Third, we examined the consequences of two nutritional conditions of central insulin unresponsiveness (fasting or short-term high-fat diet) on the ability of insulin to modify ROS level and food intake. RESULTS: In normal chow-fed mice, insulin inhibited food intake. At the same dose, insulin rapidly and transiently increased hypothalamic ROS levels by 36%. The pharmacological suppression of this insulin-stimulated ROS elevation, either by antioxidant or by an NADPH oxidase inhibitor, abolished the anorexigenic effect of insulin. Finally, in fasted and short-term high-fat diet-fed mice, insulin did not promote elevation of ROS level and food intake inhibition, likely because of an increase in hypothalamic diet-induced antioxidant defense systems. CONCLUSIONS: A hypothalamic ROS increase through NADPH oxidase is required for the anorexigenic effect of insulin.

1872-678X (Electronic) Journal Article Research Support, Non-U.S. Gov't; Different roles of mitochondria in brain function according to brain area are now clearly emerging. Unfortunately, no technique is yet described to investigate mitochondria function in specific brain area. In this article, we provide a complete description of a procedure to analyze the mitochondrial function in rat brain biopsies. Our two-step method consists in a saponin permeabilization of fresh brain tissues in combination with high-resolution respirometry to acquire the integrated respiratory rate of the biopsy. In the first part, we carefully checked the mitochondria integrity after permeabilization, defined experimental conditions to determine the respiratory control ratio (RCR), and tested the reproducibility of this technique. In the second part, we applied our method to test its sensitivity. As a result, this method was sensitive enough to reveal region specificity of mitochondrial respiration within the brain. Moreover, we detected physiopathological modulation of the mitochondrial function in the hypothalamus. Thus this new technique that takes all cell types into account, and does not discard or select any mitochondria sub-population is very suitable to analyze the integrated mitochondrial respiration of brain biopsies.

1525-2191 (Electronic) Journal Article; Obesity is associated with low-grade inflammation and leukocyte infiltration in white adipose tissue (WAT) and is linked to diabetic complications. Semicarbazide-sensitive amine oxidase, also known as vascular adhesion protein-1 (SSAO/VAP-1), is a membrane protein that is highly expressed in adipocytes and is also present on the endothelial cell surface where it is involved in leukocyte extravasation. We studied fat deposition and leukocyte infiltration in WAT of mice with a null mutation in the amine oxidase copper-containing-3 (AOC3) gene encoding SSAO/VAP-1. Both epididymal and inguinal WATs were larger in 6-month-old AOC3-KO males than in age-matched wild-type controls. However, WAT from AOC3-KO mice contained lower CD45 mRNA levels and fewer CD45(+) leukocytes. Subpopulation analyses revealed a diminished infiltration of WAT by T cells, macrophages, natural killer, and natural killer T cells. A decrease in leukocyte content in WAT was also detected in female AOC3-KO mice as early as 2 months of age, whereas increased fat mass was evident by 6 months of age. Reduced CD45(+) populations in WAT of AOC3-KO mice was not rescued by human SSAO/VAP-1 expression on adipocytes under the control of aP2, suggesting the importance of vascular AOC3 in leukocyte entrance into fat. Our results indicate that SSAO/VAP-1 is instrumental for the presence of leukocytes in WAT. Therefore, AOC3-KO mice present a unique model of mild obesity, characterized by increased WAT devoid of low-grade inflammation.

1525-2191 (Electronic) Journal Article Research Support, Non-U.S. Gov't; Treatment of cancer using radiation can be significantly compromised by the development of severe acute and late damage to normal tissue. Treatments that either reduce the risk and severity of damage or that facilitate the healing of radiation injuries are being developed, including autologous adipose tissue grafts to repair tissue defects or involutional disorders that result from tumor resection. Adipose tissue is specialized in energy storage and contains different cell types, including preadipocytes, which could be used for autologous transplantation. It has long been considered a poorly proliferative connective tissue; however, the acute effects of ionizing radiation on adipose tissue have not been investigated. Therefore, the aim of this study was to characterize the alterations induced in adipose tissue by total body irradiation. A severe decrease in proliferating cells, as well as a significant increase in apoptotic cells, was observed in vivo in inguinal fat pads following irradiation. Additionally, irradiation altered the hematopoietic population. Decreases in the proliferation and differentiation capacities of non-hematopoietic progenitors were also observed following irradiation. Together, these data demonstrate that subcutaneous adipose tissue is very sensitive to irradiation, leading to a profound alteration of its developmental potential. This damage could also alter the reconstructive properties of adipose tissue and, therefore, calls into question its use in autologous fat transfer following radiotherapy.

1262-3636 Sp. Iss. 1

International audience; Seven termite species (Isoptera) from five families were tested for disease susceptibility against the entomopathogenic fungus Metarhizium anisopliae using a standard protocol: Mastotermes darwiniensis (Mastotermitidae), Hodotermopsis sjoestedti (termopsidae), Hodotermes mossambicus (Hodotermitidae), Kalotermes flavicollis (Kalotermitidae), Reticulitermes flavipes and Prorhinotermes canalifrons (Rhinotermitidae), and Nasutitermes voeltzkowi (termitidae). Our results showed a large diversity in disease susceptibility against M. anisopliae among the different species tested and we suggest that the evolution of disease resistance mechanisms in Isoptera may be influenced by the selective pressure of the nesting ecology of each species.

1873-2933 (Electronic) Journal Article; OBJECTIVES: This work aims to evaluate the resistance of mononuclear cells to oxidative stress using a "KRL" test, formerly utilized to evaluate the resistance of erythrocyte to free radicals. METHODS: The "KRL" test evaluates the resistance to lysis of cells treated by free radicals generated under standardized conditions. RESULTS: We defined new analytical parameters (level of radical production, time course, number of cells) to obtain an accurate assay determining the resistance to oxidative stress of mononuclear cells, in comparison to that of erythrocytes. This test allows the evaluation of change in the redox state of mononuclear cells (improved by an antioxidant mix or deteriorated by antimycin A-induced mitochondrial radical overproduction). Interestingly, our data show that the sensitivity of mononuclear cells to oxidative stress is not correlated with the susceptibility of erythrocytes to oxidative stress. CONCLUSIONS: The quantification of the susceptibility of mononuclear cells to oxidative stress gives additional information (in addition to erythrocyte resistance) and could be helpful for patients with chronic inflammation.

Wolbachia-like microorganisms have been implicated in unidirectional cytoplasmic incompatibility between strains of Drosophila simulans. Reduced egg eclosion occurs when females from uninfected strains (type W) are crossed with males from infected strains (type R). Here we characterize a third incompatibility type (type S) which is also correlated with the presence of Wolbachia-like microorganisms. Despite the fact that the symbionts cannot be morphologically distinguished, we observed complete bidirectional incompatibility between R and S strains. This indicates that the determinants of incompatibility are different in the two infected types. S/W incompatibility is unidirectional and similar to R/W incompatibility. A worldwide survey of D. simulans strains showed that type S incompatibility was found only in insular populations which harbor the mitochondrial type SiI. Both W and R types were found among mainland and island populations harboring the worldwide mitochondrial type SiII. Type S incompatibility could be involved in the reinforcement of the geographical isolation of SiI populations.

International audience; Cystine-glutamate transporters import cystine into cells for glutathione synthesis and protection from oxidative stress, but also export significant amounts of glutamate. Increasing evidence suggests that 'ambient extracellular glutamate' secreted by cystine-glutamate transporters in the nervous system modulates glutamatergic synapse strength and behavior. To date, the only cystine-glutamate transporter mutants examined behaviorally are Drosophila genderblind mutants. These animals contain loss-of-function mutations in the 'genderblind' gene, which encodes an xCT subunit essential for cystine-glutamate transporter function. Genderblind was named based on a mutant courtship phenotype: male genderblind mutants are attracted to normally aversive male pheromones and thus court and attempt to copulate with both male and female partners equally. However, genderblind protein is expressed in many parts of the fly brain and thus might be expected to also regulate other behaviors, including behaviors not related to male courtship or chemosensation. Here, we show that genderblind mutants display faster recovery and increased negative geotaxis after strong mechanical stimuli (e.g., they climb faster and farther after vial banging). This phenotype is displayed by both males and females, consistent with strong genderblind expression in both sexes.

1529-2401 (Electronic) Journal Article Research Support, Non-U.S. Gov't; Sexual behavior between males is observed in many species, but the biological factors involved are poorly known. In mammals, manipulation of dopamine has revealed the role of this neuromodulator on male sexual behavior. We used genetic and pharmacological approaches to manipulate the dopamine level in dopaminergic cells in Drosophila and investigated the consequence of this manipulation on male-male courtship behavior. Males with increased dopamine level showed enhanced propensity to court other males but did not change their courtship toward virgin females, general olfactory response, general gustatory response, or locomotor activity. Our results indicate that the high intensity of male-male interaction shown by these manipulated males was related to their altered sensory perception of other males.

1097-6256 (Print) Journal Article Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't; Mate choice is an evolutionarily critical decision that requires the detection of multiple sex-specific signals followed by central integration of these signals to direct appropriate behavior. The mechanisms controlling mate choice remain poorly understood. Here, we show that the glial amino-acid transporter genderblind controls whether Drosophila melanogaster males will attempt to mate with other males. Genderblind (gb) mutant males showed no alteration in heterosexual courtship or copulation, but were attracted to normally unappealing male species-specific chemosensory cues. As a result, genderblind mutant males courted and attempted to copulate with other Drosophila males. This homosexual behavior could be induced within hours using inducible RNAi, suggesting that genderblind controls nervous system function rather than its development. Consistent with this, and indicating that glial genderblind regulates ambient extracellular glutamate to suppress glutamatergic synapse strength in vivo, homosexual behavior could be turned on and off by altering glutamatergic transmission pharmacologically and/or genetically.

1873-5495 (Electronic) Journal Article Research Support, U.S. Gov't, Non-P.H.S.; The colleterial glands of insects are organs associated with the female genital apparatus. In cockroaches, these glands produce secretions that cover two parallel rows of eggs during oviposition, and in oviparous species, these secretions become the tanned, sculpted, rigid outer casing of the ootheca. The goal of this study was to compare the gross anatomy of the colleterial glands and the ultrastructure of their component tubules in the phylogenetically significant genera Cryptocercus (Blattaria) and Mastotermes (Isoptera). Recent studies indicate that cockroaches in the genus Cryptocercus are the sister group of termites, and Mastotermes is the only termite known to produce a cockroach-like ootheca. One additional oviparous cockroach, Therea, and two additional termites, Zootermopsis and Pseudacanthotermes, were also examined. As in other cockroaches, the colleterial glands of Cryptocercus and Therea are asymmetrical, with a well developed bipartite left gland and a smaller right gland. In the termites Mastotermes, Zootermopsis, and Pseudacanthotermes, the colleterial glands are composed of a well-developed, paired, anterior gland and a small posterior gland; histological staining and cytological evidence suggest that these are homologues of the left and the right colleterial glands of cockroaches, respectively. At the ultrastructural level, colleterial gland tubules are made of cells belonging to a modified class 1 type cell in the cockroaches, in Mastotermes, and in Zootermopsis; the latter lays its eggs singly, without a surrounding ootheca-like structure. In the advanced termite Pseudacanthotermes, the tubules are made of secretory units belonging to the class 3 cell type. This study demonstrates that the cytological characteristics of colleterial glands in basal termites are similar to those of cockroaches, whether the termite secretes an oothecal casing that covers two parallel rows of eggs, as in Mastotermes, or lays its eggs singly, as in Zootermopsis. The function of colleterial glands in non-mastotermitid termites is unknown.

1873-5495 (Electronic) Comparative Study Journal Article; Thirty-nine species belonging to different families of termites are studied to give a comprehensive view of the evolution of the sternal glands. Several modifications occurring at cuticular and cytological levels are described in neuter castes. The outer epicuticle is always pierced by epicuticular pores. In advanced termites the epicuticular filaments greatly increase in number and length creating a thick layer. The pore canals gradually enlarge while the cuticle changes into a lattice structure lining an extracellular space in which the secretion is stored. Two classes of cells are present in basal termites (Mastotermitidae, Hodotermitidae, Termopsidae and Kalotermitidae) but their glandular structures greatly differ between families. A more complex organization with three classes of cells is found in the Serritermitidae and Rhinotermitidae. A regressive evolution occurs in the Termitidae where only two classes of cells are present. A dual nervous control (campaniform sensilla and neurosecretory fibers) is found in lower termites, except for the Hodotermitidae which have mechanosensory bristles. In the other families, neurosecretory fibers are lacking. A comparison with phylogenetic data is given. A more versatile role of sternal glands in neuter castes is hypothesized.

0098-0331 (Print) Journal Article; In the framework of an evolutionary study, trail pheromones have been studied in the most basal extant termite, Mastotermes darwiniensis (Mastotermitidae), and two other basal termites, the Termopsidae Porotermes adamsoni (Porotermitinae) and Stolotermes victoriensis (Stolotermitinae). Although workers of M. darwiniensis do not walk in single file while exploring a new environment under experimental conditions and are unable to follow artificial trails in 'open field' experiments, they do secrete a trail-following pheromone from their sternal glands. This unique behavior might reflect a primitive function of communication of the sternal gland. The major component of the pheromone appears to be the same in the three basal species: the norsesquiterpene alcohol (E)-2,6,10-trimethyl-5,9-undecadien-1-ol. This represents a new chemical category of trail-following pheromones for termites. The quantity of pheromone was estimated as 20 pg/individual in M. darwiniensis, 700 pg/individual in P. adamsoni, and 4 pg/individual in S. victoriensis. The activity threshold was 1 ng/cm in M. darwiniensis and 10 pg/cm in P. adamsoni. In M. darwiniensis, the trail pheromone was secreted by sternal gland 4 and to a lesser degree by sternal gland 3, sternal gland 5 being almost inactive. This study highlighted phylogenetic relationships between the Mastotermitidae and two subfamilies of the Termopsidae, the Porotermitinae and the Stolotermitinae. Furthermore, it indicated a heterogeneity within the Termopsidae, with Porotermitinae and Stolotermitinae on one hand, and Termopsinae on the other. Finally, Mastotermitidae and Termopsidae, with C14 trail pheromones, are clearly separated from the Kalotermitidae, Rhinotermitidae, and Termitidae that secrete C12 or C20 trail pheromones.

1095-6433 doi: DOI: 10.1016/j.cbpa.2007.01.155

0012-1797 (Print) Journal Article Research Support, Non-U.S. Gov't; The ability for the brain to sense peripheral fuel availability is mainly accomplished within the hypothalamus, which detects ongoing systemic nutrients and adjusts food intake and peripheral metabolism as needed. Here, we hypothesized that mitochondrial reactive oxygen species (ROS) could trigger sensing of nutrients within the hypothalamus. For this purpose, we induced acute hypertriglyceridemia in rats and examined the function of mitochondria in the hypothalamus. Hypertriglyceridemia led to a rapid increase in the mitochondrial respiration in the ventral hypothalamus together with a transient production of ROS. Cerebral inhibition of fatty acids-CoA mitochondrial uptake prevented the hypertriglyceridemia-stimulated ROS production, indicating that ROS derived from mitochondrial metabolism. The hypertriglyceridemia-stimulated ROS production was associated with change in the intracellular redox state without any noxious cytotoxic effects, suggesting that ROS function acutely as signaling molecules. Moreover, cerebral inhibition of hypertriglyceridemia-stimulated ROS production fully abolished the satiety related to the hypertriglyceridemia, suggesting that hypothalamic ROS production was required to restrain food intake during hypertriglyceridemia. Finally, we found that fasting disrupted the hypertriglyceridemia-stimulated ROS production, indicating that the redox mechanism of brain nutrient sensing could be modulated under physiological conditions. Altogether, these findings support the role of mitochondrial ROS as molecular actors implied in brain nutrient sensing.

0001-8244 (Print) Journal Article Research Support, Non-U.S. Gov't; Brain maturation, a developmental process influenced by both endogenous and environmental factors, can affect sexual behavior. In vertebrates and invertebrates, sexual maturation is under the influence of hormones and neuromodulators, but the role of developmental genes in this process is still poorly understood. We report that prospero (pros), a gene crucial for nervous system development, can change the age of onset of sexual behavior in Drosophila melanogaster males: adult males carrying a single copy of several pros mutations court females and mate at a younger age than control males. However, these pros mutations had no effect on female sexual receptivity and did not alter other male phenotypes related to mating behavior. The Pros protein was detected in several brain and sensory structures of immature adult males, some of which are normally involved in the regulation of male specific behaviors. Our data suggest that the altered pros expression affects the age of onset of male mating behavior.

0012-1606 (Print) Comparative Study Journal Article Research Support, Non-U.S. Gov't; Adaptive animal behaviors depend upon the precise development of the nervous system that underlies them. In Drosophila melanogaster, the pan-neural prospero gene (pros), is involved in various aspects of neurogenesis including cell cycle control, axonal outgrowth, neuronal and glial cell differentiation. As these results have been generally obtained with null pros mutants inducing embryonic lethality, the role of pros during later development remains poorly known. Using several pros-Voila (prosV) alleles, that induce multiple developmental and behavioral anomalies in the larva and in adult, we explored the relationship between these phenotypes and the variation of pros expression in 5 different neural regions during pre-imaginal development. We found that the quantity of pros mRNA spliced variants and of Pros protein varied between these alleles in a tissue-specific and developmental way. Moreover, in prosV1 and prosV13 alleles, the respective decrease or increase of pros expression, affected (i) neuronal and glial cell composition, (ii) cell proliferation and death and (iii) axonal-dendritic outgrowth in a stage and cellular context dependant way. The various phenotypic consequences induced during development, related to more or less subtle differences in gene expression, indicate that Pros level needs a precise and specific adjustment in each neural organ to allow its proper function.

Sex pheromones are chemical signals frequently required for mate choice, but their reciprocal role on mate preference has rarely been shown in both sexes. InDrosophila melanogasterflies, the predominant cuticular hydrocarbons (CHs) are sexually dimorphic: only females produce 7,11-dienes, whereas 7-tricosene (7-T) is the principal male CH. Males generally prefer females with 7,11-dienes, but the role of 7-T on female behaviour remains unclear. With perfumed males, control females mated faster and more often with males carrying increased levels of 7-T showing that this CH acts as a chemical stimulant forD. melanogasterfemales. Control females—but not antenna-less females—could detect small variation of 7-T. Finally, our finding thatdesat1mutant female showed altered response towards 7-T provides an additional role for this gene which affects the production and the perception of pheromones involved in mate choice, in both sexes.

0003-3472 doi: DOI: 10.1016/j.anbehav.2006.01.027; Nearly all studies of Drosophila melanogaster courtship have focused exclusively on male behaviour. Female precopulatory behaviour is often relegated to ‘accept' or ‘reject' mating, and how female behaviour interacts with that of males remains largely unknown. Using a video-computing approach, we measured 10 behavioural states and 22 elementary behaviours that occur during the precopulatory phase of Canton-S (Cs) flies. Male and female behaviours were studied under a variety of social conditions. This allowed us to identify which one(s) is relevant to courtship or to general activity. Our analysis showed that the courtship repertoire was not constant over time, as female and male behaviours varied during courtship, and that virgin Cs females showed several behavioural sequences whose frequency, latency and total duration could predict their mating success. The most striking index of females' willingness to copulate was the way they extruded the ovipositor: a partial extrusion stimulated male courtship, whereas total extrusion inhibited it. Furthermore, females that mated preened their abdomen more frequently than did females that did not mate. We also observed that male sexual activity was strongly increased when the female emitted a tiny volatile droplet at the tip of the ovipositor. We hypothesized that partial ovipositor extrusion, droplet emission and abdominal preening are related behaviours that result in the active dispersion of compounds spread on the female's abdomen. Such a detailed picture of the precopulatory behaviours should allow better characterization of the sensory stimuli exchanged during courtship in Drosophila.

1379-1176 (Print) Journal Article; In order to complete previous studies conducted on Blattella germanica, three insecticides from different groups were evaluated: boric acid, an inorganic compound, benfuracarb, a carbamate, and halofenozide, a non-steroidal ecdysone agonist. Boric acid (8.20%, LD50) and benfuracarb (2%, LD50) were incorporated into the diet and orally administrated to newly emerged adults of both sexes, while halofenozide (0.33%, LD50) was applied topically. Hydrocarbons extracts was made on bidistilled pentane from control and treated series sampled 6 days following treatment. Extracts was analyzed by gas chromatography. Data showed that cuticular profiles of control and treated series were qualitatively similar with thirteen major compounds; however, significant quantitative differences were noted. Boric acid seemed to feminize the cuticular profile in males with a significant reduction of the two first cuticular compounds detected. Halofenozide and benfuracarb reduced cuticular compounds in both sexes.

0048-3575 doi: DOI: 10.1016/j.pestbp.2005.05.002; Oral toxicity of boric acid, an inorganic insecticide, was evaluated on German cockroach, Blattella germanica L. (Dictyoptera, Blattellidae). Newly emerged adults were exposed to various concentrations of boric acid incorporated into the diet. Results showed that treated insects exhibited toxic symptoms with a dose-dependent mortality. Histological study of midgut revealed alterations in the epithelial cells and a signiWcant increase in the epithelium thickness. In a second series of experiments, the compound was investigated on the activities of acetylcholinesterase (AChE) and glutathione S-transferases (GSTs). Data showed that the compound induced GSTs and reduced the activity of AChE. From this experiment, it may be concluded that ingested boric acid caused death of insects perhaps ultimately by starvation via alterations of the midgut. Additionally, the compound seemed to present a neurotoxic action as evidenced by the symptoms of poisoning and the reduction in AchE activity.

1567-133X (Print) Journal Article Research Support, Non-U.S. Gov't; During Drosophila embryonic development, various morphogenetic processes require the remodeling of the extracellular matrix. In a previous study, we have identified and characterized a cDNA encoding a novel putative extracellular matrix protein named tenebrin, in the beetle Tenebrio molitor. Here, we examine the expression of the Drosophila ortholog, referred to as Tenectin (Tnc), during embryonic development. Tnc is expressed in the majority of tissues of neuroectodermic origin such as hindgut, foregut, tracheal system, anal plate, and CNS. In the CNS, the Tnc transcript is restricted to a few cells, whereas the protein is located in the dorsal part of the axonal tracts. In the hindgut and the trachea, Tnc protein is expressed on the apical pole of the cells. Tnc is an extracellular matrix protein secreted in a polarized way in different organs of Drosophila embryos.

1526-954X (Print) Journal Article Research Support, Non-U.S. Gov't; The regulatory sequences of the Drosophila ACP65A cuticle gene were analyzed in vivo in transgenic flies, using both fusion genes constructs and transposase-mediated deletions within a P element containing ACP65A regulatory sequences fused to the lacZ gene (deletion scanning). The sequences located between -594 and +161 are sufficient to confer both temporal and spatial expression specificities, indicating the presence of tissue-specific enhancers and response elements to hormone-induced factors. In addition, timing of expression and tissue-specificity appear to be controlled by distinct cis-regulatory elements, which suggests the existence of independent hormonal and tissue-specific signaling pathways. Gain and loss of function studies also implicate DHR38, the Drosophila homolog of the vertebrate NGFI-B-type nuclear receptors, as an important activator of the ACP65A gene.

0001-8244 (Print) Journal Article Review; I review the recent literature on cuticular hydrocarbons (CHs) in Drosophila. First, the major structural features of CHs are examined in a variety of species with regard to phylogeny. The genetic bases of the CH variation between and within species have been investigated with some of the genes involved characterized and manipulated. The effect of non-genetic factors as temperature, food and development is also examined with regard to CH production. Using a model involving the stimulating or the inhibiting role of CHs, it is possible to speculate on the mechanisms of CH perception and on the role(s) that these substances could play on sexual isolation and on the evolution of pheromonal communication.

All tests involved a pair of 5-day-old male and female (intact or decapitated) flies. Females were ‘homotypic’ (same species and strain as the tested male: D. melanogaster, Cs strain; D. mauritiana, 163.1 strain; D. simulans, Seychelles strain), ‘desat1 non-perfumed’ (D. melanogaster desat1 mutant), ‘perfumed’ (desat1 with transfer of Cs females pheromones), or ‘Cs’ (D. melanogaster control strain). Data shown are the frequencies of courtship (with both intact and decapitated females) and of mating (with intact females), within a 1 h observation period and were calculated from the total number of tested pairs (shown in brackets). D. mauritiana males courted (χ2 = 16.81, P < 0.001) and mated (χ2 = 8.83, P < 0.01) more frequently with intact desat1 non

0092-8674 (Print) Journal Article Research Support, Non-U.S. Gov't; A critical determinant of insect body size is the time at which the larva stops feeding and initiates wandering in preparation for metamorphosis. No genes have been identified that regulate growth by contributing to this key developmental decision to terminate feeding. We show here that mutations in the DHR4 orphan nuclear receptor result in larvae that precociously leave the food to form premature prepupae, resulting in abbreviated larval development that translates directly into smaller and lighter animals. In addition, we show that DHR4 plays a central role in the genetic cascades triggered by the steroid hormone ecdysone at the onset of metamorphosis, acting as both a repressor of the early ecdysone-induced regulatory genes and an inducer of the betaFTZ-F1 midprepupal competence factor. We propose that DHR4 coordinates growth and maturation in Drosophila by mediating endocrine responses to the attainment of critical weight during larval development.

0962-8452 (Print) Comparative Study Journal Article Research Support, Non-U.S. Gov't; The evolution of communication is a fundamental biological problem. The genetic control of the signal and its reception must be tightly coadapted, especially in inter-individual sexual communication. However, there is very little experimental evidence for tight genetic linkage connecting the emission of a signal and its reception. We found that a single genomic transposon inserted in the desatl gene of Drosophila melanogaster simultaneously affected the emission and the perception of sex-specific signals. This mutation greatly decreased the production of unsaturated hydrocarbons on the cuticle of mature flies of both sexes. These substances represent the sex pheromones necessary for mate discrimination: control males could not discriminate the sex of mutant desatl flies. Moreover, mutant males were unable to discriminate the sex pheromones of con-trol flies. Expression of desatl was found in the peripheral tissues that produce and detect sex pheromones.Excision of the transposon rescued both the production and discrimination phenotypes, but the two effects did not always coincide. This indicates that the emission and perception of pheromones are coded by differ-ent products of the same gene, reflecting the pleiotropic activity of desatl .

Behaviour depends (a) on genes that specify the neural and non-neural elements involved in the perception of and responses to sensory stimuli and (b) on experience that can modulate the fine development of these elements. We exposed transgenic and control Drosophila melanogaster males, and their hybrids, to male siblings during adult development and measured the contribution of genes and of experience to their courtship behaviour. The transgene CheB42a specifically targets male gustatory sensillae and alters the perception of male inhibitory pheromones which leads to frequent male–male interactions. The age at which social experience occurred and the genotype of tester males induced a variable effect on the intensity of male homo- and heterosexual courtship. The strong interaction between the effects of genes and of social experience reveals the plasticity of the apparently stereotyped elements involved in male courtship behaviour. Finally, a high intensity of homosexual courtship was found only in males that simultaneously carried a mutation in their white gene and the CheB42a transgene.

0022-0949 (Print) Comparative Study Journal Article Research Support, Non-U.S. Gov't; Social interaction with conspecifics can influence the developing brain and behaviour of the exposed animal. This experience can involve the exchange and retention of visual, chemical, acoustic and tactile signals. When several Drosophila melanogaster male flies are associated with mated females in the presence of food, they show frequent aggressive interactions. To measure the role of social experience on male-male interaction, two tester males - naive or exposed to sibling(s) during a variable period of their adult development - were confronted in the absence of female and food. The two males displayed homosexual courtship and aggressive behaviours, the frequency, intensity and directionality of which varied according to their experience. The effect of social experience was greatly enhanced between transgenic males partially defective for pheromonal perception, indicating that male inhibitory pheromones are normally used to repress male-male interaction.

0022–0795/04/0181–147; This study investigated the ability of insulin and of insect insulin-like peptides (ILPs) to stimulate ovarian steroidogenesis in the blowfly Phormia regina. Bovine insulin was active on ovaries isolated in vitro, which showed an age-dependent sensitivity; this peptide progressively stimulated steroidogenesis in ovaries isolated from the third day after adult molt, but not in younger ones, and had maximal activity after the fifth day. This stimulatory effect was observed equally from females reared in the presence or in the absence of males, excluding a regulatory effect of mating. The mode of action of insulin in blowflies did not involve cAMP, but triggered a specific and well-conserved transduction cascade. In particular, a peroxovanadium compound, known to activate specifically the insulin receptor in mammals, also stimulated blowfly ovarian steroidogenesis in vitro. Conversely, chemicals known to inhibit the mammalian insulin receptor or downstream elements of its signaling pathway, such as LY294002, an inhibitor of phosphatidylinositol 3-kinase (PI3K), were able to prevent the steroidogenic action of bovine insulin on fly ovaries. Extracts from the median neurosecretory cells (MNCs) of blowfly brains, which are known to contain endogenous ILPs, stimulated ovarian steroidogenesis very efficiently and were also sensitive to inhibition by LY294002. These experiments indicated the involvement of PI3K in the mode of action of MNC extracts and substantiated that their endogenous ILPs are involved in the regulation of ovarian steroidogenesis. This conclusion was corroborated by the effects of synthetic bombyxin II, an ILP originating from silkworm MNCs, which also stimulated steroidogenesis in isolated blowfly ovaries. Altogether, these data suggest that insulinlike neurohormones from MNCs play a crucial role as steroidogenic gonadotropins in female flies

The detection of changes in glucose level constitutes the first step of the control of glucose homeostasis. Glucose sensors are therefore expected to be present in different parts of the body and particularly in the central nervous system. Some studies have already attempted to determine glucose-sensitive cerebral structures either after a glucoprivic stimulus or after prolonged hyperglycaemia. By analogy to beta cells, it was postulated that the glucose sensors in the brain could involve GLUT2, glucokinase and/or ATP-sensitive K + channels. Surprisingly, GLUT2 was mainly found in astrocytes. Thus, the aims of the present investigation were to determine, in awake rats: (i) the hypothalamic areas that respond to acute hyperglycaemic condition induced by an intracarotid injection of glucose and (ii) the involvement of astrocytes in glucose-sensing by the use of a glial drug, methionine sulfoximine. Rats were given intracarotid injections of glucose solution to trigger a transient insulin secretion without change in peripheral glycaemia, thus involving only central nervous regulation. Hypothalamic activation was determined by immunodetection of the immediate early gene c-fos protein. Acute glucose injection induces significant activation of arcuate and paraventricular nuclei. This stimulation mainly affects neurones in both nuclei, but also astrocytes in the former as illustrated by double immunohistochemistry (Fos and neuronal nuclei or glial fibrillary acidic protein). After specific impairment of astrocyte metabolism by methionine sulfoximine, cerebral activation disappears in the arcuate nucleus, correlated with the lack of cerebral glucose-induced insulin secretion. Therefore, arcuate and paraventricular hypothalamic nuclei are able to detect acute cerebral hyperglycaemia, leading to a peripheral stimulation of insulin secretion. Arcuate nucleus and more especially astrocytes in this nucleus play a pivotal role in glucose-sensing.

1379-1176 (Print) Journal Article Research Support, Non-U.S. Gov't; Blatta germanica is the more prevalent cockroach species in Algeria. In the present study, we tested the effect on reproduction in B. germanica of two insect growth regulators, RH-0345, a benzoylhydrazine analogue that mimics the action of 20-hydroxyecdysone, and methoprene, one of the most commercially important juvenile hormone analogues, and a novel carbamate insecticide, benfuracarb. The compounds were applied topically (10 and 20 microg/insect for RH-0345, and 1 and 10 microg/insect for methoprene) or orally administrated (at 2% for benfuracarb) on newly emerged females and evaluated on reproductive events during the adult life (2, 4 and 6 days). Treatment with RH-0345 and benfuracarb reduced significantly the number of oocytes, the size and the volume of the basal oocyte during the experimental period. Methoprene distorted the ovarian development since it caused a significant reduction in the number of oocytes at 2, 4 and 6 days for the two tested doses, and an increase in oocyte size at 2, 4 and 6 days with 1 microg and a decrease with 10 microg. In a second series of experiments, the effects of these compounds were assayed on the ovarian proteins. Data from biochemical analysis revealed that RH-0345 and benfuracarb reduced the ovarian amounts of proteins, while treatment with methoprene increased it during the sexual maturation.

0022-0949 (Print) Comparative Study Journal Article Research Support, Non-U.S. Gov't; Sex pheromones are chemical signals used for mate attraction and discrimination in many invertebrate species. These compounds are often complex mixtures with different components having different effects. We tested live Drosophila melanogaster mutant female flies genetically depleted for unsaturated cuticular hydrocarbons, which were then perfumed with these substances to measure their influence on various aspects of reproduction. Female pheromones of the control Cs strain enhanced female attractivity, copulation duration and tended to decrease the number of female progeny of mutant females mated with Cs males, but no dose-dependent effect was found. Cs and variant males showed different response to Cs female pheromone, suggesting a strain-specific coadaptation of female and male characters. The fact that female pheromones induced reciprocal effects on the frequency of the genes contributed by females and males suggests that these substances regulate coevolutionary processes between the sexes.

0362-2525 (Print) Comparative Study Journal Article; In female alates of Macrotermes annandalei, two types of abdominal glands are involved in the secretion of sex pheromone. Tergal glands are found at the anterior margin of tergites 6-10 and posterior sternal glands (PSGs) are located at the anterior margin of sternites 6-7. The cytological features of both types of glands are quite similar. The fine structural organization of PSGs is studied more precisely and described for the first time. The glandular cuticle is pitted with narrow apertures corresponding to the openings of numerous subcuticular pouches. Several Class 3 glandular units open in each pouch. One canal cell and one secretory cell make an individual glandular unit. The canal cell is enlarged apically and is connected with the other canal cells to form a common pouch. Based on the structural features found in these glands, we propose a common secretory process for PSGs and tergal glands. During the physiological maturation of alates inside the nest, secretory vesicles amass in the cytoplasm of secretory cells, while large intercellular spaces collapse the cuticular pouches. At the time of dispersal flight, pouches are filled with the content of secretory vesicles while intercellular spaces are sharply reduced. After calling behavior, no secretion remains in the glands and pouches collapse again, while secretory cells are drastically reduced in size. The structure and the secretory processes of PSGs and tergal glands are compared to those of abdominal sexual glands known in termites.

0028-1042 (Print) Journal Article; The diunsaturated C12 alcohol (Z,Z)-dodeca-3,6-dien-1-ol (dodecadienol) has been characterized by GC-MS and FTIR as a novel releaser pheromone in termites. This alcohol identified in Ancistrotermes pakistanicus (Termitidae, Macrotermitinae) possesses a double pheromonal function which again illustrates the chemical parsimony of termites compared with other social insects. In workers, dodecadienol elicits trail-following at a very low concentration (activity threshold at 0.1 pg/cm of trail); in male alates it induces trail-following at a low concentration (1-10 pg/cm) and sexual attraction at a higher concentration (about 1 ng). Traces of the monounsaturated C12 alcohol (Z)-dodec-3-en-1-ol (dodecenol), known as a trail pheromone of several Macrotermitinae, were also found in the sternal gland extracts of A. pakistanicus, although only dodecadienol was present at the surface of the sternal gland. Workers of A. pakistanicus are not sensitive to dodecenol, but they are as sensitive to dodecatrienol as to dodecadienol. However, in the study area (Vietnam), A. pakistanicus is living in sympatry only with those Macrotermitinae using dodecenol as a trail pheromone, the foraging populations therefore being well isolated through their respective trail pheromones. The presence of three types of unsaturated C12 alcohols as releaser pheromones in the only Macrotermitinae subfamily is discussed, and a possible biosynthetic pathway from linoleic acid is proposed for dodecadienol.

0949-944X (Print) Journal Article Research Support, Non-U.S. Gov't; We used differential display to isolate epidermis cDNAs corresponding to juvenile-hormone analog-regulated mRNA from the beetle Tenebrio molitor. One of them encodes a putative extracellular matrix (ECM) protein, named Tenebrin. Indeed, the deduced protein sequence contains ECM typical features like the presence of a signal peptide, internal repeats, a RGD tripeptide sequence motif known to bind integrins and von Willebrand factor type c domains involved in protein-protein interactions. Northern blot analysis reveals a single transcript of about 11 kb with an expression pattern correlated to 20-hydroxyecdysone fluctuations during metamorphosis. In vivo injections of exogenous 20-hydroxyecdysone alone or combined with cycloheximide show that Tenebrin expression is directly induced by this hormone. Methoprene (a juvenile hormone analog) application experiments show that Tenebrin expression is rapidly induced by this analog. This gene is still up-regulated in the presence of protein synthesis inhibitor but, in these conditions, the mRNA induction level is not maximal.

0376-6357 (Print) Journal article