Mating enhances the probability of winning aggressive encounters in male lobster cockroaches

In the present study, we report that contact with isolated female antenna significantly increases both the pheromone 3-hydroxy-2-butanone (3H-2B) release and the hemolymph JH III level in all examined aggressive posture-adopting (AP) and NP (non-AP-adopting) socially naïve males, with significantly faster concomitant pre-mating wing-raising behavior in AP as compared to NP males. 3H-2B release and JH III level were significantly increased after mating in both AP and NP males. A positive correlation was observed between mating experience and dominant status. Furthermore, mated-AP males initiated fights more rapidly and fought for a significantly longer duration than mated-NP males; retention with the paired female for 24 h did not affect this increase. JH III level and 3H-2B release were significantly increased in dominant males as compared to subordinates. These results suggest that prior mating experience in invertebrates may enhance aggression in subsequent male–male encounters, with accompanying physiological (hormone and pheromone) responses.     

Pheromone, juvenile hormone, and social status in the male lobster cockroach Nauphoeta cinerea

In this study, the major pheromone component, 3‐hydroxy‐2‐butanone (3H‐2B), released by dominants was measured during early scotophase. Both the JH III titer in the hemolymph and the 3H‐2B content of the sternal glands of the dominants and subordinates were then measured during late scotophase and late photophase. These investigations were performed on encounter days 1, 2, 3, 5, 7, 9, 12, and 20. The results showed that, for non‐aggressive posture (AP)‐adopting socially naïve males (SNMs), both the 3H‐2B release and the hemolymph JH III titer were maintained at a low level. Once a fight occurred, 3H‐2B release was raised significantly in the AP‐adopting dominants, but not in non‐AP‐adopting subordinates, and remained raised throughout the entire experimental period. At 30 min after the first encounter, the hemolymph JH III titer was significantly increased in dominants, but not in subordinates. A significantly higher hemolymph JH III titer was observed in dominants during late scotophase on days 3, 5, 12, and 20 and during late photophase on days 3, 5, and 20. After fighting, the sternal gland 3H‐2B content of the dominants or subordinates was significantly lower than in SNMs. In dominants, the sternal gland 3H‐2B content during late scotophase was significantly lower than that during late photophase in the first 9 domination days, while, in the subordinates, the 3H‐2B content during late scotophase was either similar to, or significantly higher than, that in late photophase. In the dominants, 3H‐2B release and JH III titer were positively correlated. In rank switchers, the switched social status was positively correlated with both 3H‐2B release and JH III titer. Comparison of 3H‐2B release and JH III titer in 1‐time, 3‐time, or 5‐time dominants showed that, although winning significantly increased both 3H‐2B release and JH III titer, there is no significant difference in 3H‐2B release between 3‐ and 5‐time winners, while the JH III titer was most significantly increased in the 3‐time winners. The possible relationship between pheromone release, JH III titer, and social status is discussed. Arch. Insect Biochem. Physiol. 2008. © 2008 Wiley‐Liss, Inc.     

Juvenile hormone and the ontogeny of cockroach aggression

Our previous study [Kou et al., 2008. Juvenile hormone levels are increased in winners of cockroach fights. Horm. Behav. 52, 252–260] showed that the basic principle of the challenge hypothesis (hormone levels can respond to social stimuli to modulate aggression in vertebrates] could be applied to juvenile hormone (JH) levels and aggression in the lobster cockroach Nauphoeta cinerea. In that study, 80- to 85-day-old socially naïve males were used, as fighting is much more easily initiated in these older animals than in younger males, and JH III levels in the dominant were found to be significantly increased after an encounter compared to before the encounter and were significantly higher than those in the subordinates. In N. cinerea, newly emerged males usually show no aggressiveness towards each other and aggression is only initiated after several days of close contact. To investigate the development of aggression from an early age, in the present study, newly emerged males were paired to investigate the relationship between JH levels and aggression. The results showed that injection of JH III significantly increased the probability of the young males being fight winners. In each age group in which aggression was initiated, the dominants had significantly higher JH levels than either the subordinates or the same aged non-fighters. JH injection of subordinates on the day of rank establishment had no effect on the probability of rank switch. These results indicate that, (i) in newly emerged male pairs, JH plays a decisive role in rank establishment and the fact that dominant status is significantly associated with a higher JH titer and subordinate status with a lower JH titer is consistent with the basic principle of the challenge hypothesis, and (ii) after rank establishment, the lack of effect of JH treatment on rank change is consistent with the idea of “social inertia” in vertebrates.     

Methyl farnesoate and juvenile hormone III in normal and precocene treated embryos of the ovoviviparous cockroach Nauphoeta cinerea

Summary

Titers of juvenile hormone III and methyl farnesoate, its unepoxidized precursor, were measured throughout embryonic development using a gas-chromatographic method and it was revealed that both substances are undetectable before dorsal closure. Thereafter they both reach similarly high concentrations (800 ng/g) until a few days before hatching, when their titers begin to decrease. Application of the precocene analogue, 7-ethoxy-6-methoxy-2,2-dimethyl-chromene, to egg-cases at dorsal closure results in very low or undetectable titers of juvenile hormone III, depending on the dose applied, whereas methyl farnesoate is seen to reach high levels similar to those seen in the controls. The severe developmental disturbances observed suggest that juvenile hormone III is very important for normal development and formation of the 1st instar larva.     

Regulation of sex pheromone production in the male Nauphoeta cinerea cockroach: Role of brain extracts,corpora allata�(CA), and juvenile hormone (JH)

The neuroendocrine mechanisms underlying pheromone regulation in cockroaches are unclear because of a lack of physiological and chemical data. The present report describes experiments designed to determine the role of the brain, corpora allata, and juvenile hormone III in the production of sex pheromone by male Nauphoeta cinerea cockroaches. The levels of two sex pheromone components, i.e., acetoin and 2‐methylthiazolidine, were measured by gas chromatographic analysis of sternal gland extracts obtained from individual males. Allatectomy or decapitation performed up to 2 to 3 days after imaginal molt caused a decrease in sex phermone levels. Conversely decapitation or allatectomy performed after 3 to 4 days post‐eclosion had almost no effect on sex pheromone levels. Injection of JHIII into allatectomized and decapitated males stimulated pheromone production while injection of brain extract had no effect. These results indicate that JHIII is involved in the differentiation of sternal glands and that regulation via pheromone biosynthesis activating neuropeptide (PBAN) does not occur in N. Cinerea cockroaches. Arch. Insect Biochem. Physiol. 40:165–172, 1999. © 1999 Wiley‐Liss, Inc.     

Solubilization, identification, and localization of vitellogenin-binding sites in follicles of the cockroach, Nauphoeta cinerea

Binding sites for vitellogenin were solubilized and analyzed either with a filter assay or with ligand blotting. We tested sodium dodecylsulfate (SDS), Chaps, octyl-beta-D-glucoside, and sodium deoxycholate and found SDS and sodium deoxycholate to be most effective in solubilizing both high and low molecular weight binding sites. In the filter assay the sodium deoxycholate extracts, but not the SDS extracts, maintained binding activity after dilution of the solubilizer below its critical micellar concentration. In ligand blotting we consistently observed, in vitellogenic follicles, binding sites with an apparent Mr of approximately 200,000, 35,000, and three closely spaced bands between 14,000 and 20,000. Binding of vitellogenin to all binding sites was suppressed in the presence of the drug suramin. Analysis of corpora lutea and oothecae as well as of ovariole sheath, follicle cell/basal lamina, and oocyte plasma membrane preparations showed that the 35 and 14-20 kDa binding sites are located in the outer follicle compartments, and the 200 kDa binding site in the oocyte plasma membrane. In the latter we occasionally also observed binding sites with an apparent Mr of approximately 150,000, 95,000, 67,000 and 30,000, particularly at stages after ovulation. The 35 and 14-20 kDa binding sites, as visualized in stained gels and in ligand blotting, are rather abundant and were also seen in several other male and female tissues of Nauphoeta and even in other species. They also bound other 14C-labeled hemolymph proteins and thus appear to be rather unspecific. As binding analysis with nonsolubilized and sodium deoxycholate-solubilized membranes revealed that the quantity of vitellogenin bound by binding sites of the outer follicle compartments was low, it is conceivable that the abundance of the 14-20 kDa and 35 kDa binding sites in ligand blotting is merely an effect of SDS and does not reflect the in vivo situation. We suppose that the 200 kDa binding site of the oocyte plasma membrane represents the vitellogenin receptor involved in endocytosis.     

Degradation of juvenile hormone and methylation of juvenile hormone acid by corpora cardiaca–corpora allata of the cockroach,Nauphoeta cinerea: I. Biochemical aspects

Corpora cardiaca-corpora allata (CC-CA) from vitellogenic females of Nauphoeta cinerea degraded, in vitro, racemic and (10R)-juvenile hormone III (JH III) at a rate of 249 pmol/CC-CA/h and 786 pmol/CC-CA/h, respectively. The major metabolite formed was JH III acid, together with some highly polar products. CC-CA homogenates degraded racemic JH III to a small extent, whereas (10R)-JH III was degraded efficiently to JH III acid. No highly polar products were formed by CC-CA homogenates. When CC-CA were incubated with racemic JH III acid, some of this substance was degraded to highly polar products, and a minor part was methylated to JH III. CC degraded very little JH III acid and did not methylate it to JH III. CC-CA homogenates methylated JH III acid very efficiently; we measured an apparent K_max of 37.8 μM and a V_max of 1,260 pmol/4 h/ CC-CA equivalent. The addition of JH III acid to CC-CA in vitro increased the rate of biosynthesis of JH III, as determined by measuring incorporation of methyl[¹⁴C]methionine into JH III. These data indicate that the metabolite JH III acid can enter the CA and be methylated to JH III.     

The effect of ventral nerve cord severance and male castration on female mating behavior, clutch size, and maternal care in the ring-legged earwig

Mating is critical for the expression of oviposition and maternal care in the earwig, Euborellia annulipes; additionally, mating diminishes receptivity to additional mating and promotes a decline in juvenile hormone synthesis at the end of the gonadotrophic cycle (in contrast to most insect species wherein mating stimulates juvenile hormone production). We report here that severance of the ventral nerve cord of virgin females similarly promoted egg deposition and maternal care of eggs, diminished mating receptivity, and elicited a timely decline in juvenile hormone biosynthesis. Mating of intact females to adult males that were castrated as larvae did not abolish oviposition; however, clutch size was reduced, and no eggs developed. Such castrated males had smaller seminal vesicles than did intact males, presumably attributable to lack of sperm in castrated males. In contrast, mating of intact females to males castrated on day 1 of adult life did not reduce clutch size compared with those of sham-operated animals and did not abolish fertilization; in fact, these castrated males produced viable offspring after six matings. These results are consistent with the notion that ventral nerve cord severance mimicked mating in intact animals. Following mating, the ventral nerve cord likely is a conduit to release the brain from inhibiting oviposition and maternal care. The presence of sperm in the spermatheca is not necessary for release of this inhibition but may modulate clutch size.     

Effects of decapitation and ovariectomy on the regulation of juvenile hormone synthesis in the cockroach, Diploptera punctata

Corpora allata from Diploptera punctata females at adult ecdysis or at the end of the last-larval stadium, when implanted into decapitated females, underwent a cycle of juvenile hormone synthesis similar in timing and magnitude to that of glands implanted into control animals which had been starved and allatectomized. Starvation did not alter the cycle in rates of juvenile hormone synthesis of sham-operated animals.Decapitation of ovariectomized animals resulted in no cycle in rates of juvenile hormone synthesis by implanted adult corpora allata; however, implantation of an ovary along with the corpora allata into decapitated, ovariectomized hosts resulted in a cycle of juvenile hormone synthesis. In control animals, which retained their heads but were starved and allatectomized as well as ovariectomized, the implanted corpora allata showed a cycle of juvenile hormone synthesis only when implanted with an ovary. The maximal rates of juvenile hormone synthesis by the corpora allata in both experimental and control conditions were lower than normal, likely due to the repeated trauma of surgery. However, at no time from eclosion to the end of the first gonotrophic period was the brain necessary for the cyclic response of the corpora allata to the presence of the ovary.     

Female presence affects male behavior and testosterone levels in the European starling (Sturnus vulgaris)

In this study, we confronted individually housed male European starlings (Sturnus vulgaris) with a female conspecific for 60 min to study the consequences on behavior and plasma testosterone (T) concentrations. Control males experienced a similar procedure, the only difference being that they were tested in the absence of a female. Female presence significantly affected both behavior and plasma T levels of male starlings. Experimental males spent significantly more time singing in the nest box and flew significantly more into the nest box with green nesting material during female presentations than during control periods. Control males never showed these mate attraction behaviors. In total 5 of the 16 experimental males did not respond behaviorally to the female stimulus bird (NR males). In contrast to T levels of control males, plasma T concentrations of both experimental males that did respond to the female (R males) and of NR males (which only perceived the female stimulus) were positively influenced by female presentation. The time spent singing in the nest box by experimental males (R and NR males combined) during female presence tended to be positively correlated with changes in plasma T levels. Finally, before introduction of a female, plasma T levels of R males were significantly higher than those of NR males indicating that individually housed males respond to the presence of a female conspecific by increasing their mate attraction behaviors only when a threshold plasma T concentration has been reached.     

Fatherhood, pairbonding and testosterone in the Philippines

In species with a high level of paternal care, including humans, testosterone (T) is believed to help mediate the trade-off between parenting and mating effort. This hypothesis is supported by the observation of lower T in pairbonded men or fathers compared to single, non-fathers; however, prior work has highlighted population variation in the association between T and pairbonding or fatherhood status. Here we evaluate this hypothesis in a large (n = 890), representative birth cohort of young men (age range 20.5–22.5 years) living in Cebu City, the Philippines. Bioavailable T was measured in saliva collected prior to bed and immediately upon waking the following morning. Plasma T and luteinizing hormone (LH) were measured in morning plasma samples. In this sample, 20% of men were pairbonded, defined as living with a partner or married, 13% were fathers, and roughly half of fathers reported involvement in childcare. Pairbonded men had significantly lower T at both times of day. Unlike in other populations, this relationship was accounted for entirely by fatherhood status: among the large sub-sample of non-fathers, mean T was nearly identical among pairbonded and single men. There was a strong association between self-reported involvement in childcare and lower evening T, supporting the idea that the evening nadir in T is related to social interactions across the day. Similar relationships were found for total plasma T and LH, suggesting that these relationships are coordinated by centrally-mediated changes in LH secretion. The relatively modest T difference in relation to fatherhood at Cebu, in comparison to other studies, may reflect a lower level of paternal involvement in childcare activities in this population. Our findings using a large, well-characterized birth cohort support the hypothesized role of T as a mediator of mating and parenting effort in humans, while contributing evidence for cultural variation in the relative importance of pairbonding and fathering to these relationships.     

Influences of octopamine and juvenile hormone on locomotor behavior and <Emphasis Type="Italic">period</Emphasis> gene expression in the honeybee, <Emphasis Type="Italic">Apis mellifera</Emphasis>

Octopamine (OA) and juvenile hormone (JH) are implicated in the regulation of age-based division of labor in the honeybee, Apis mellifera. We tested the hypothesis that these two neuroendocrine signals influence task-associated plasticity in circadian and diurnal rhythms, and in brain expression of the clock gene period (per). Treatment with OA, OA antagonist (epinastine), or both, did not affect the age at onset of circadian rhythmicity or the free running period in constant darkness (DD). Young bees orally treated with OA in light–dark (LD) illumination regime for 6 days followed by DD showed reduced alpha (the period between the daily onset and offset of activity) during the first 4 days in LD and the first 4 days in DD. Oral treatment with OA, epinastine, or both, but not manipulations of JH levels, caused increased average daily levels and aberrant patterns of brain per mRNA oscillation in young bees. These results suggest that OA and JH do not influence the development or function of the central pacemaker but rather that OA influences the brain expression of a clock gene and characteristics of locomotor behavior that are not thought to be under direct control of the circadian pacemaker. Electronic supplementary material Supplementary material is available in the online version of this article at and is accessible for authorized users.     

Structural and functional organization of the photosystems in spinach chloroplasts. Antenna size,relative electron-transport capacity,and chlorophyll composition

The structural and functional organization of the spinach chloroplast photosystems (PS) I, II_α and II_β was investigated. Sensitive absorbance difference spectrophotometry in the ultraviolet (?A₃₂₀) and red (?A₇₀₀) regions of the spectrum provided information on the relative concentration of PS II and PS I reaction centers. The kinetic analysis of PS II and PS I photochemistry under continuous weak excitation provided information on the number (N) of chlorophyll (Chl) molecules transferring excitation energy to PS II_α, PS II_β and PS I. Spinach chloroplasts contained almost twice as many PS II reaction centers compared to PS I reaction centers. The number N_α of chlorophyll (Chl) molecules associated with PS II_α was 234, while N_β = 100 and N_{PS I} = 210. Thus, the functional photosynthetic unit size of PS II reaction centers was different from that of PS I reaction centers. The relative electron-transport capacity of PS II was significantly greater than that of PS I. Hence, under light-limiting green excitation when both Chl a and Chl b molecules are excited equally, the limiting factor in the overall electron-transfer reaction was the turnover of PS I. The Chl composition of PS I, PS II_α and PS II_β was analyzed on the basis of a core Chl a reaction center complex component and a Chl $\text{a}\text{b-}\text{LHC}$ component. There is a dissimilar Chl $\text{a}\text{b-}\text{LHC}$ composition in the three photosystems with 77% of total Chl b associated with PS II_α only. The results indicate that PS II_α, located in the membrane of the grana partition region, is poised to receive excitation from a wider spectral window than PS II_β and PS I.     

Effects of 20-hydroxyecdysone and other hormones on egg development, and identification of a vitellin-binding protein in the ovary of the tick, Amblyomma hebraeum

Partially fed adult female Amblyomma hebraeum ticks were injected with 20-hydroxyecdysone (20E; up to 43 microg/g body weight (bw)), juvenile hormone III (JH III; up to 100 microg/g bw), bovine insulin (up to 2000 mU/g bw), or triiodothyronine (up to 200 ng/g bw) in an attempt to stimulate vitellogenesis. Of these, only 20E stimulated synthesis and release of vitellogenin (Vg). Immunoblot analysis revealed that Vg-synthesis occurred in the fat body. However, consistent with earlier observations suggesting that a distinct signal may be required for Vg-uptake, there was no significant Vg-uptake by oocytes of partially fed, 20E-treated ticks. Because Vg-uptake commonly occurs via receptor-mediated endocytosis (i.e., a specific Vg-receptor), we attempted to identify a vitellin (Vt)-binding protein in ovaries of engorged female ticks. A single 86 kDa Vt-binding protein was identified, even under reducing conditions (2-mercaptoethanol), by a ligand-blotting technique. Sodium salt of suramin (5 mM) inhibited binding of Vt to the 86 kDa protein. However, this protein was also detected in ovaries from small partially fed ticks (50-100 mg), suggesting that the inability of 20E to stimulate Vg-uptake in partially fed ticks may not have been due to the absence of a Vg-receptor.     

Relative contribution of cell contact pattern, specific PKC isoforms and gap junctional communication in tight junction assembly in the mouse early embryo

In mouse early development, cell contact patterns regulate the spatial organization and segregation of inner cell mass (ICM) and trophectoderm epithelium (TE) during blastocyst morphogenesis. Progressive membrane assembly of tight junctional (TJ) proteins in the differentiating TE during cleavage is upregulated by cell contact asymmetry (outside position) and suppressed within the ICM by cell contact symmetry (inside position). This is reversible, and immunosurgical isolation of the ICM induces upregulation of TJ assembly in a sequence that broadly mimics that occurring during blastocyst formation. The mechanism relating cell contact pattern and TJ assembly was investigated in the ICM model with respect to PKC-mediated signaling and gap junctional communication. Our results indicate that complete cell contact asymmetry is required for TJ biogenesis and acts upstream of PKC-mediated signaling. Specific inhibition of two PKC isoforms, PKCdelta and zeta, revealed that both PKC activities are required for membrane assembly of ZO-2 TJ protein, while only PKCzeta activity is involved in regulating ZO-1alpha+ membrane assembly, suggesting different mechanisms for individual TJ proteins. Gap junctional communication had no apparent influence on either TJ formation or PKC signaling but was itself affected by changes of cell contact patterns. Our data suggest that the dynamics of cell contact patterns coordinate the spatial organization of TJ formation via specific PKC signaling pathways during blastocyst biogenesis.     

Ketone components in the contact sex pheromone of the white-spotted longicorn beetle, Anoplophora malasiaca, and pheromonal activity of synthetic ketones

Two active fractions were found during the isolation of contact sex pheromone of female elytra of the white‐spotted longicorn beetle, Anoplophora malasiaca (Thomson) (Coleoptera: Cerambycidae), in addition to fraction of hydrocarbons that had previously been identified. One fraction was essential to evoke a series of precopulatory behaviors of males toward a glass dummy when coated together with the hydrocarbon blend. The other fraction enhanced this activity when added to the mixture. From the latter synergistic fraction, we isolated five novel compounds and identified them as 10‐heptacosanone, (Z)‐18‐heptacosen‐10‐one, (18Z,21Z)‐heptacosa‐18,21‐dien‐10‐one, (18Z,21Z,24Z)‐heptacosa‐18,21,24‐trien‐10‐one, and 12‐heptacosanone by GC‐MS and NMR analyses. A blend of four of these synthetic ketones, without 12‐heptacosanone, in the ratio and concentration found in female elytra extract (250 : 400 : 1000 : 180 ng FE^?1) showed greater synergistic effect than the natural fraction containing the ketones. This effect was canceled out by further addition of 12‐heptacosanone (100 ng FE^?1), which was still comparable to the effect of the natural ketone fraction.     

Correlation of oxygen consumption, cytochrome c oxidase, and cytochrome c oxidase subunit I gene expression in the termination of larval diapause in the bamboo borer, Omphisa fuscidentalis

The moth Omphisa fuscidentalis (Lepidoptera, Pyralidae) is a univoltine insect with a larval diapause period lasting up to 9 months. We studied changes in O(2) consumption in conjunction with cytochrome c oxidase activity and cytochrome c oxidase subunit I (cox1) gene expression. O(2) consumption changed within a day, showing a supradian rhythm with a ca.12-h cycle at 25 degrees C. During the first two-thirds of the diapause period, from October to March, O(2) consumption was constant until January and then increased by March. Topical application of methoprene, a juvenile hormone analog (JHA), to diapausing larvae terminated the diapause and was associated with an increase in O(2) consumption rate at diapause termination. In JHA-treated larvae, cytochrome c oxidase activity in fat bodies was high at the beginning of the prepupal period and highest at pupation. cox1 expression in fat bodies displayed a transient peak 8 days after JHA application and peaked in the prepupal period. Taken together, our results show that the break of diapause by JHA is associated with the activation of cox1, bringing about an increase in cytochrome c oxidase activity, followed by an increase in O(2) consumption rate.     

Genomic structure, polymorphism and expression analysis of the growth hormone (GH) gene in female and male Half-smooth tongue sole (Cynoglossus semilaevis)

Growth hormone (GH) is a polypeptide which is an important regulator of development and somatic growth in teleosts, and may be associated with the mechanisms which drive sexual growth dimorphism in the Half-smooth tongue sole (Cynoglossus semilaevis). In this study, the full length gh cDNA was cloned from C. semilaevis by homology cloning and the rapid amplification of cDNA ends polymerase chain reaction (RACE-PCR). The full-length gh cDNA is 826 bp and contains an open reading frame (ORF) of 603 bp encoding a protein of 200 amino acids (AA). The precursor of gh consists of a 17 amino-acid signal peptide followed by a 183 amino-acid mature polypeptide. GH gene sequences obtained from female and male adults consist of 3428 bp and 3371 bp, respectively, each of which includes six exons and five introns, and the difference in the GH gene size was mainly caused by the microsatellites. When 14 tissues from females, normal males and extra-large male adults were analyzed for sex-specific tissue expression, the gh mRNA was found to be predominantly expressed in the pituitary, and the expression levels in females were 3.6 times as much as those in normal males, while the mRNA expression in extra-large males was 1.7 times as much as those in normal males. Sex differences in gh mRNA expression during development were also examined by using a full-sib family of C. semilaevis, and the gh mRNA was detected at all of the 12 time points sampled from 10 to 380 days-old. A significant increase in gh mRNA was detected starting in 80 day old fish and was then followed by a drop to very low levels starting at 230 day old fish. Differential expression indicated that the gh expression level in females was significantly higher than males (P < 0.01) at all of the stages except for 10 days-old. Two microsatellite loci were identified in the second intron of the GH gene. Using these two polymorphic markers to genotype 224 individuals, there was no significant difference between the females and males in the Bohai Sea, the Yellow Sea and the hatchery samples.