Male reproduction is affected by RNA interference of period and timeless in the desert locust Schistocerca gregaria

In all living organisms, behavior, metabolism and physiology are under the regulation of a circadian clock. The molecular machinery of this clock has been conserved throughout the animal kingdom. Besides regulating the circadian timing of a variety of processes through a central oscillating mechanism in the brain, these circadian clock genes were found to have a function in peripheral tissues in different insects. Here, we provide evidence that the circadian clock genes period (per) and timeless (tim) have a role in the male locust reproduction. A knockdown of either of the two genes has no effect on male sexual maturation or behavior, but progeny output in their untreated female copulation partners is affected. Indeed, the fertilization rates of the eggs are lower for females with a per or tim RNAi copulation partner as compared to the eggs deposited by females that mated with a control male. As the sperm content of the seminal vesicles is higher in per or tim knockdown males, we suggest that this phenotype could be caused by a disturbance of the circadian regulated sperm transfer in the male reproductive organs, or an insufficient maturation of the sperm after release from the testes.     

Effects of copulation temperature on female reproductive output and longevity in the wolf spider Pardosa astrigera (Araneae: Lycosidae)

The copulation duration of male wolf spider Pardosa astrigera, was significantly influenced by environmental temperature, as had been found in some insect species. Therefore, temperature during male courtship and copulation may influence the amount of sperm and seminal fluids transferred during copulation, which in turn could influence female fitness. In order to test this hypothesis, we subjected pairs of male and female P. astrigera to five temperature groups from 16 to 32 °C at an interval of 4 °C, and investigated whether and to what extent the various temperatures during male courtship and copulation influenced female reproductive output and female adult longevity under controlled laboratory conditions. With the increase of copulation temperature, females were more likely to lay egg sacs. The total egg sacs and lifetime fecundity of female were positively influenced by copulation temperature, whereas female lifetime spiderlings and adult longevity were independent of copulation temperature.     

Functional Characterization of the Short Neuropeptide F Receptor in the Desert Locust,Schistocerca gregaria

Whereas short neuropeptide F (sNPF) has already been reported to stimulate feeding behaviour in a variety of insect species, the opposite effect was observed in the desert locust. In the present study, we cloned a G protein-coupled receptor (GPCR) cDNA from the desert locust, Schistocerca gregaria. Cell-based functional analysis of this receptor indicated that it is activated by both known isoforms of Schgr-sNPF in a concentration dependent manner, with EC₅₀ values in the nanomolar range. This Schgr-sNPF receptor constitutes the first functionally characterized peptide GPCR in locusts. The in vivo effects of the sNPF signalling pathway on the regulation of feeding in locusts were further studied by knocking down the newly identified Schgr-sNPF receptor by means of RNA interference, as well as by means of peptide injection studies. While injection of sNPF caused an inhibitory effect on food uptake in the desert locust, knocking down the corresponding peptide receptor resulted in an increase of total food uptake when compared to control animals. This is the first comprehensive study in which a clearly negative correlation is described between the sNPF signalling pathway and feeding, prompting a reconsideration of the diverse roles of sNPFs in the physiology of insects.     

Olfactory bulbectomy-induced changes in the reproductive behaviour of the south Indian gerbil, Tatera indica cuvieri

Male South Indian gerbils (T. indica cuvieri), both adults and weanlings, were olfactory bulbectomized (OBX) and changes in male reproductive behaviour were assessed. Consequent to OBX, majority of the adult males failed to ejaculate, and courtship behaviour has also been considerably reduced. All OBX weanlings were rendered incapable of ejaculation. However, maturational parameters, and organ weights (testes, epididymis and seminal vesicle) remained unchanged in OBX males.     

Male Seminal Fluid Substances Affect Sperm Competition Success and Female Reproductive Behavior in a Seed Beetle

Male seminal fluid proteins are known to affect female reproductive behavior and physiology by reducing mating receptivity and by increasing egg production rates. Such substances are also though to increase the competitive fertilization success of males, but the empirical foundation for this tenet is restricted. Here, we examined the effects of injections of size-fractioned protein extracts from male reproductive organs on both male competitive fertilization success (i.e., P2 in double mating experiments) and female reproduction in the seed beetle Callosobruchus maculatus. We found that extracts of male seminal vesicles and ejaculatory ducts increased competitive fertilization success when males mated with females 1 day after the females’ initial mating, while extracts from accessory glands and testes increased competitive fertilization success when males mated with females 2 days after the females’ initial mating. Moreover, different size fractions of seminal fluid proteins had distinct and partly antagonistic effects on male competitive fertilization success. Collectively, our experiments show that several different seminal fluid proteins, deriving from different parts in the male reproductive tract and of different molecular weight, affect male competitive fertilization success in C. maculatus. Our results highlight the diverse effects of seminal fluid proteins and show that the function of such proteins can be contingent upon female mating status. We also document effects of different size fractions on female mating receptivity and egg laying rates, which can serve as a basis for future efforts to identify the molecular identity of seminal fluid proteins and their function in this model species.     

The circadian clock genes affect reproductive capacity in the desert locust Schistocerca gregaria

The circadian clocks govern many metabolic and behavioral processes in an organism. In insects, these clocks and their molecular machinery have been found to influence reproduction in many different ways. Reproductive behavior including courtship, copulation and egg deposition, is under strong influence of the daily rhythm. At the molecular level, the individual clock components also have their role in normal progress of oogenesis and spermatogenesis. In this study on the desert locust Schistocerca gregaria, three circadian clock genes were identified and their expression profiles were determined. High expression was predominantly found in reproductive tissues. Similar daily expression profiles were found for period (per) and timeless (tim), while the clock (clk) mRNA level is higher 12 h before the first per and tim peak. A knockdown of either per or tim resulted in a significant decrease in the progeny produced by dsRNA treated females confirming the role of clock genes in reproduction and providing evidence that both PER and TIM are needed in the ovaries for egg development. Since the knockdown of clk is lethal for the desert locust, its function remains yet to be elucidated.     

The Effect of Limb Loss on the Courtship and Mating Behavior of the Wolf Spider Pardosa milvina (Araneae: Lycosidae)

Limb loss is common in the wolf spider Pardosa milvina, appearing in nearly one third of adult males but occurring less frequently among adult females and juveniles. Since males wave their first pair of legs during courtship displays, the reproductive consequences of limb loss may be significant. We measured the courtship and mating effects of the loss of one, two, or four legs among adult male P. milvina. Missing one or two legs did not significantly reduce a male's ability to mate, but missing four legs was detrimental to mating success, reduced both courtship intensity and copulation duration, and increased cannibalism frequency. Results suggest behavioral flexibility in compensating for limited leg loss and a defensive function of the anteriormost legs to thwart female cannibalism attempts.     

Copulation Song in Drosophila: Do Females Sing to Change Male Ejaculate Allocation and Incite Postcopulatory Mate Choice?

Drosophila males sing a courtship song to achieve copulations with females. Females were recently found to sing a distinct song during copulation, which depends on male seminal fluid transfer and delays female remating. Here, it is hypothesized that female copulation song is a signal directed at the copulating male and changes ejaculate allocation. This may alter female remating and sperm usage, and thereby affect postcopulatory mate choice. Mechanisms of how female copulation song is elicited, how males respond to copulation song, and how remating is modulated, are considered. The potential adaptive value of female signaling during copulation is discussed with reference to vertebrate copulation calls and their proposed function in eliciting mate guarding. Female copulation song may be widespread within the Drosophila genus. This newly discovered behavior opens many interesting avenues for future research, including investigation of how sexually dimorphic neuronal circuits mediate communication between nervous system and reproductive organs.     

Pheromonal inhibition of male courtship behaviour in the brown tree snake, Boiga irregularis: a mechanism for the rejection of potential mates

Pheromones play a central role in coordinating the events leading up to copulation in snakes. We report here a novel pheromone system in the brown tree snake in which females release a pheromone that inhibits male courtship behaviour. In a previous study, we made observations of female brown tree snakes releasing cloacal secretions (CS) during courtship that appeared to cause courting males to cease courtship. All snakes have glands that release CS through ducts located along the cloacal orifice. Although CS have been studied for many years, their function in the mediation of snake behaviour has not been experimentally well determined. We examined the role of CS in the reproductive behaviour of male and female brown tree snakes. We conducted four experiments to test the effect of both male and female CS on brown tree snake behaviour under two behavioural contexts, courtship and male-male ritualized combat. Within each experiment, we compared the effects of CS to a control. Female CS caused a decrease in the time that males spent courting females and a decrease in the intensity of courtship compared with the control treatment. Male CS did not, however, affect the time that males spent displaying courtship or the intensity of that courtship. Neither male nor female CS had significant effects on male ritualized combat behaviour, including time that males spent in combat or the intensity of combat behaviours displayed. Furthermore, neither female nor male CS had an effect on female courtship versus controls. The inhibition of brown tree snake reproductive behaviours is specific to female CS inhibiting male courtship behaviour. This pheromone acts in concert with the female sex pheromone to regulate the events leading to copulation.Copyright 2003 Published by Elsevier Science Ltd on behalf of The Association for the Study of Animal Behaviour.      

Live for the moment—Adaptations in the male genital system of a sexually cannibalistic spider (Theridiidae, Araneae)

Monogyny in spiders culminates in extreme traits, like dramatic male self-sacrifice and emasculation of the male by the female during copulation. Here we show that monogynous males can be highly adapted for this fatal sexual behaviour. Dwarf males of the one-palped theridiid spider Tidarren argo, which are cannibalised immediately after the insertion of their single copulatory organ, stop spermiogenesis when reaching adulthood. Their testes atrophy, which might economise the energy expenditures of these males. We also found that the amount of seminal fluid produced is stored in an enlarged seminal vesicle until the single sperm induction takes place. The volume of the seminal vesicle is similar to the sperm droplet taken up into the copulatory organ (palpal organ). Sperm uptake takes much longer than in related species most likely due to the large amount of seminal fluid. As shown by histological observations males are able to fill one of the paired female sperm storage organs during copulation thereby presumably impeding subsequent charging by rival males.     

New reproductive anomalies in fruitless-mutant Drosophila males: extreme lengthening of mating durations and infertility correlated with defective serotonergic innervation of reproductive organs

Several features of male reproductive behavior are under the neural control of fruitless (fru) in Drosophila melanogaster. This gene is known to influence courtship steps prior to mating, due to the absence of attempted copulation in the behavioral repertoire of most types of fru-mutant males. However, certain combinations of fru mutations allow for fertility. By analyzing such matings and their consequences, we uncovered two striking defects: mating times up to four times the normal average duration of copulation; and frequent infertility, regardless of the time of mating by a given transheterozygous fru-mutant male. The lengthened copulation times may be connected with fru-induced defects in the formation of a male-specific abdominal muscle. Production of sperm and certain seminal fluid proteins are normal in these fru mutants. However, analysis of postmating qualities of females that copulated with transheterozygous mutants strongly implied defects in the ability of these males to transfer sperm and seminal fluids. Such abnormalities may be associated with certain serotonergic neurons in the abdominal ganglion in which production of 5HT is regulated by fru. These cells send processes to contractile muscles of the male's internal sex organs; such projection patterns are aberrant in the semifertile fru mutants. Therefore, the reproductive functions regulated by fruitless are expanded in their scope, encompassing not only the earliest stages of courtship behavior along with almost all subsequent steps in the behavioral sequence, but also more than one component of the culminating events.     

Male displays and female preferences in the courtship of a gregarious cricket

The courtship of males of the gregarious cricket Amphiacusta maya involves a variety of signals. The quantitative aspects of both successful and unsuccessful courtship sequences were examined to determine whether certain aspects of male displays were correlated with female mating preferences regardless of which male performed them. Although variability among males was high for most courtship components measured, I found no evidence of female choice with respect to the courtship variables studied. About 35% of the duration of each male courtship sequence is devoted to chirping, but there were no differences in either the likelihood of copulation or the latency to copulation between normal males and experimentally silenced males. The possibility that intrinsic differences in male quality explained the variability in courtship duration was examined with a two-way analysis of variance. The variance in courtship duration was attributable to variance among females, not to variance among males. Thus the courtship behaviour of male A. maya is variable enough to allow females to exert stabilizing or disruptive selection on displays, but there is no evidence that females use the available information.     

The Influence of Pedipalp Autotomy on the Courtship and Mating Behavior of Pardosa milvina (Araneae: Lycosidae)

Male Pardosa milvina wolf spiders use their pedipalps both for copulation and courtship. Pedipalp loss is significantly more common among adult males compared to females. We measured the courtship and mating effects associated with the loss of one or both pedipalps among adult male P. milvina. Pedipalp loss significantly reduced courtship intensity, but had no influence on mounting success. Intact males were less likely to be cannibalized and suffered fewer predatory attacks by females than autotomized males. Loss of the left pedipalp resulted in significantly less intense courtship, higher female aggression levels, and delayed onset of courtship whereas loss of the right pedipalp minimally affected male and female behavior relative to intact males. Pedipalp displays may function in reducing female aggression rather than increasing female receptivity.     

Evolutionary Rate Covariation Identifies New Members of a Protein Network Required for Drosophila melanogaster Female Post-Mating Responses

Seminal fluid proteins transferred from males to females during copulation are required for full fertility and can exert dramatic effects on female physiology and behavior. In Drosophila melanogaster, the seminal protein sex peptide (SP) affects mated females by increasing egg production and decreasing receptivity to courtship. These behavioral changes persist for several days because SP binds to sperm that are stored in the female. SP is then gradually released, allowing it to interact with its female-expressed receptor. The binding of SP to sperm requires five additional seminal proteins, which act together in a network. Hundreds of uncharacterized male and female proteins have been identified in this species, but individually screening each protein for network function would present a logistical challenge. To prioritize the screening of these proteins for involvement in the SP network, we used a comparative genomic method to identify candidate proteins whose evolutionary rates across the Drosophila phylogeny co-vary with those of the SP network proteins. Subsequent functional testing of 18 co-varying candidates by RNA interference identified three male seminal proteins and three female reproductive tract proteins that are each required for the long-term persistence of SP responses in females. Molecular genetic analysis showed the three new male proteins are required for the transfer of other network proteins to females and for SP to become bound to sperm that are stored in mated females. The three female proteins, in contrast, act downstream of SP binding and sperm storage. These findings expand the number of seminal proteins required for SP''s actions in the female and show that multiple female proteins are necessary for the SP response. Furthermore, our functional analyses demonstrate that evolutionary rate covariation is a valuable predictive tool for identifying candidate members of interacting protein networks.     

Reproductive toxicity assessment of lasofoxifene, a selective estrogen receptor modulator (SERM), in male rats

BACKGROUND: Lasofoxifene is a nonsteroidal selective estrogen receptor modulator (SERM) with greater than 100-fold selectivity against all other steroid receptors and is a potentially superior treatment for postmenopausal osteoporosis. The purpose of this study was to evaluate the effects of lasofoxifene on male reproduction in rats in light of the known effects of estrogen modulating compounds on male reproductive ability. METHODS: Lasofoxifene was administered to adult male rats at doses of 0.1, 1, 10, and 100 mg/kg for 66-70 consecutive days. After 28 days of dosing, male rats were cohabited with untreated female rats. Female rats were euthanized on gestation day 14 and a uterine examination was carried out for evaluation of reproductive parameters and embryo viability. Male rats were euthanized after 66-70 days of dosing and epididymal sperm motility and concentration were assayed. The testes, epididymides, prostate, and seminal vesicles were weighed and microscopically examined. RESULTS: The duration of cohabitation was increased for 100 mg/kg males by 0.7 days. The number of males copulating and the number of implantation sites produced per copulation were reduced in the 10 and 100 mg/kg groups. Weights of the seminal vesicles and epididymides were reduced for all groups, although the testes weight and epididymal sperm motility and concentration were not affected by treatment. There were no microscopic findings in the male reproductive tissues. CONCLUSION: The changes in male fertility and reproductive tissue weights after exposure to lasofoxifene are consistent with those previously described for estrogen receptor-modulating compounds.     

Inhibition of sexual maturation by a urinary pheromone in male prairie deer mice

Male prairie deer mice (Peromyscus maniculatus bairdii) were treated from weaning until 8 weeks of age with chemical stimuli from conspecifics or with control substances. Growth of the testes and seminal vesicles was retarded in males that were reared in contact with soiled bedding material transferred from cages of adult males. No additional inhibition resulted from the physical presence of an adult male either continuously or for 1 hr per day. Application of urine collected from adult males to the noses of young males retarded seminal vesicle growth. Removal of the olfactory bulbs of males at 3 weeks of age blocked the inhibitory influence of urine on sexual maturation. Exposure to urine from adult females did not alter the growth of the reproductive organs in young males. The ability of a male deer mouse to retard the sexual maturation of young male conspecifics (Bediz, G. M., and Whitsett, J. M., 1979, J. Comp. Physiol. Psychol.93, 493–500) appears to be a consequence of chemical stimuli excreted in its urine.