Peter W. Sorensen

Functional Identification of a Goldfish Odorant Receptor

The vertebrate olfactory system utilizes odorant receptors to receive and discriminate thousands of different chemical stimuli. An understanding of how these receptors encode information about an odorants molecular structure requires a characterization of their ligand specificities. We employed an expression cloning strategy to identify a goldfish odorant receptor that is activated by amino acids-potent odorants for fish. Structure-activity analysis indicates that the receptor is preferentially tuned to recognize basic amino acids. The receptor is a member of a multigene family of G protein-coupled receptors, sharing sequence similarities with the calcium sensing, metabotropic glutamate, and V2R class of vomeronasal receptors. The ligand tuning properties of the goldfish amino acid odorant receptor provide information for unraveling the molecular mechanisms underlying olfactory coding.

Hormonal and pheromonal control of spawning behavior in the goldfish

Species that employ sexual reproduction must synchronize gamete maturity with behavior within and between genders. Teleost fishes solve this challenge by using reproductive hormones both as endogenous signals to synchronize sexual behavior with gamete maturation, and as exogenous signals (pheromones) to synchronize spawning interactions between fish. This dual role of hormonal products is best understood in the goldfish, an external fertilizer with a promiscuous mating system. Female gonadal growth and vitellogenesis is stimulated by 17β-estradiol (E2) which also evokes release of a recrudescent pheromone. At the completion of vitellogenesis, ovarian E2 production drops and plasma testosterone increases, sensitizing the female gonadotropin II (luteinizing hormone; LH) system to environmental cues (temperature, spawning substrate, pheromones). These cues eventually trigger a LH surge that alters steroidogenesic pathways to favor the production of progestins including 17,20β-dihydroxy-4-pregnen-3-one (17,20β-P). Plasma 17,20β-P stimulates oocyte maturation but is also released to the water along with sulfated 17,20β-P and androstenedione to serve as a preovulatory pheromone. This pheromone stimulates male behavior, LH release, and sperm production. At the time of ovulation, females become sexually active in response to prostaglandin F2α (PGF2α) synthesized in the oviduct. PGF2α and its metabolites are released as a postovulatory pheromone that induces male spawning behavior which further increases male LH and sperm production. Androgenic hormones are required for male behavior and LH release. Although goldfish are gonochorists, hormone treatments can induce heterotypical functions in adults. Similar findings in other fish demonstrate that a sexually bipotential brain is not restricted to hermaphroditic fishes.

Brief review of fish pheromones and discussion of their possible uses in the control of non‐indigenous teleost fishes

Abstract Most species of fish rely on pheromones (chemical signals released by conspecifics) to mediate social behaviours. Three categories of pheromones can be discerned based on their function: anti‐predator cues, social cues, and reproductive cues. Each of these categories comprises pheromones that can induce “primer” effects (developmental and/or endocrinological changes) and/or “releaser” effects (strong behavioural changes). A handful of fish pheromones have been chemically identified and all are remarkably potent. Almost all are metabolic products whose production is seemingly unspecialised, insofar as they are not synthesised by specialised structures. Importantly, their potency and specificity makes them ideal candidates for use in control of both threatened (native) and unwanted (non‐indigenous) fish species. As has been so for insect control and now sea lamprey control in the North American Great Lakes, these cues could be used in many ways as part of integrated control programmes for invasive teleost fishes. Ideally, these programmes would be designed to simultaneously exploit multiple weaknesses in species’ life histories while being fully cognisant of stock‐recruitment relationships. Generally the approach would be to use a variety of pheromones to supplement and increase the efficiencies of other control strategies including the application of poisons or fish with genetic modification, trapping for removal or sterilisation, and barriers to prevent spread. Integrated pest control using pheromones appears especially practical for the common carp (Cyprinus carpio) which appears to use many of the same cues as the goldfish (Carassius auratus) and for which half a dozen pheromones have already been identified.

Neural processing, perception, and behavioral responses to natural chemical stimuli by fish and crustaceans

This manuscript reviews the chemical ecology of two of the major aquatic animal models, fish and crustaceans, in the study of chemoreception. By necessity, it is restricted in scope, with most emphasis placed on teleost fish and decapod crustaceans. First, we describe the nature of the chemical world perceived by fish and crustaceans, giving examples of the abilities of these animals to analyze complex natural odors. Fish and crustaceans share the same environments and have evolved some similar chemosensory features: the ability to detect and discern mixtures of small metabolites in highly variable backgrounds and to use this information to identify food, mates, predators, and habitat. Next, we give examples of the molecular nature of some of these natural products, including a description of methodologies used to identify them. Both fish and crustaceans use their olfactory and gustatory systems to detect amino acids, amines, and nucleotides, among many other compounds, while fish olfactory systems also detect mixtures of sex steroids and prostaglandins with high specificity and sensitivity. Third, we discuss the importance of plasticity in chemical sensing by fish and crustaceans. Finally, we conclude with a description of how natural chemical stimuli are processed by chemosensory systems. In both fishes and crustaceans, the olfactory system is especially adept at mixture discrimination, while gustation is well suited to facilitate precise localization and ingestion of food. The behaviors of both fish and crustaceans can be defined by the chemical worlds in which they live and the abilities of their nervous systems to detect and identify specific features in their domains. An understanding of these worlds and the sensory systems that provide the animals with information about them provides insight into the chemical ecology of these species.

Environmental estrogens suppress hormones, behavior, and reproductive fitness in male fathead minnows

This study explored the possibility that environmental estrogens in sewage effluent may reduce the reproductive fitness of adult male fish by suppressing their reproductive behaviors, including their ability to compete for nests and females. Male fathead minnows (Pimephales promelas) were exposed for three weeks to either blank control, effluent released by a sewage treatment plant (STPE), waterborne estradiol (E2), or a synthetic androgen (methyltestosterone [MT]). Afterward, fish were placed with females and a nest, and their behavior was monitored for 5 d in either the presence or the absence of a competing (unexposed control) male. Males exposed to either the STPE or E2 (approximately 50 ng/L, a level chosen to mimic the estrogenic content of the STPE) had elevated levels of circulating vitellogenin (p < 0.05) and lower levels of 11-ketotestosterone (KT; p < 0.05). Nearly all STPE- and E2-exposed males spawned successfully in the absence of a competing male, but in both cases, exposed males suffered nearly total reproductive failure when they had to compete. Conversely, males exposed to MT (approximately 50 ng/L) outcompeted control males. Behavioral observations suggested that subtle differences in agonistic behaviors, typically associated with circulating androgens (i.e., KT), were responsible. We speculate that male fish exposed to estrogenic effluent in the field are less likely to reproduce successfully within large populations of wild fish, thereby causing abnormal and potentially detrimental patterns of gene flow within those populations.

The Chemical Ecology and Potential Application of the Sea Lamprey Migratory Pheromone

A variety of studies has established that migratory adult sea lampreys (Petromyzon marinus) locate rivers for spawning using a potent bile acid-derived pheromone released by larval conspecifics and perhaps other lamprey species. Adult sea lampreys are caught in the greatest numbers in those tributaries of the Great Lakes which contain large populations of larval sea lamprey, and removing larvae from these tributaries makes them less attractive than neighboring rivers which still have larvae. Stream selection appears to be mediated by odorous cues because occluding the olfactory systems of adults severely reduces their ability to locate streams. Experiments using laboratory mazes confirm that adult sea lampreys are attracted to very low dilutions of stream water, especially when it is collected from streams that contain larval lamprey. Tests of larval holding water find it to be highly attractive and suggest that a single larva activates almost 4,000 L of river water an hour. Finally, biochemical studies combined with electrophysiological recording from the sea lamprey olfactory system and behavioral experiments demonstrate that the migratory pheromone released by larvae is comprised of the unique lamprey bile acid, petromyzonol sulfate and at least one unknown compound. The precise importance of this mixture and its specific role(s) in mediating stream finding behavior and upstream migration has yet to be resolved. Together, these studies demonstrate that this pheromone is a potent stimulant of adult sea lamprey migratory behavior which might be useful in sea lamprey control. Three possibilities for its use in streams as well as six pheromone formulations are discussed.

Evolution and Specialization of Fish Hormonal Pheromones

Teleost fish commonly release steroid and prostaglandin hormones and their metabolites to the water, where some function as water-borne odorants that induce specific physiological and/or behavioral reproductive responses in conspecifics. In this paper, we evaluate processes responsible for the evolution and specialization of these hormonal pheromones and give examples. Several new definitions are coined. The topic is complex owing to the evolutionary age and great diversity of fish species. We hypothesize that the evolution of fish hormonal pheromones has been influenced by two types of factors, factors intrinsic to the pheromonal function (e.g. factors which directly determine the nature of pheromones while being themselves influenced by how well the pheromones function), and extrinsic factors which are not subject to conspecific feedback. Both types of factors may influence receivers and/or donors of pheromonal stimuli in independent manners. We further hypothesize that hormonal pheromones originated through’ spying,’ in which the receiver benefits by detecting a hormonal cue and in which neither the donor nor its cue is specialized. However, we also suggest that in many instances the receiver’s response will benefit the donor, thereby driving specialization of pheromone production-a phenomenon we term’ communication.’ Whereas evolutionary pressures to increase the efficiency of cue detection may lead to an increase in the number of stimuli detected, sexual selection acting on the donor may cause species to increase the efficiency of signal production. Donors might achieve this by’ amplifying’ signal output and/or increasing the biological relevance by optimizing its information content through’ signal elaboration.’ In some instances hormonal pheromones also will be modified by extrinsic factors, a subset of which may lead to the evolution of species-specificity.

Evidence that petromyzontid lampreys employ a common migratory pheromone that is partially comprised of bile acids.

This study examined whether the larval pheromone employed by adult sea lamprey (Petromyzon marinus) to locate spawning streams and known to be at least partially comprised of bile acids is also employed by other lamprey species. Both production and release of lamprey-specific bile acids, and sensitivity to them were examined in a wide variety of species. High pressure liquid chromatography and electrospray ionization/mass spectrometry (ESI-MS) found gallbladders from 10 species of European and North American lamprey to contain large quantities of petromyzonol sulfate (PS) together with much smaller quantities of allocholic acid (ACA) and petromyzonol (P). Evaluation of holding waters from three of these species using ESI-MS found all to contain large quantities of PS and lesser quantities of ACA in similar ratios. Electro-olfactogram recording from the olfactory systems of three parasitic lamprey species found all to detect PS and ACA with high sensitivity. Behavioral studies using migratory adult sea lamprey found them to be attracted to the odors of heterospecific larvae as well as conspecific larvae, both of which contained similar amounts of PS and ACA. Finally, adult silver lampreys (Ichthyomyzon unicuspis) were also found to be attracted to the odor of larval sea lamprey. Together, these results demonstrate that PS and ACA are commonly produced and released by larval petromyzontid lampreys and likely used as part of a common evolutionarily conserved pheromone. This scenario is reasonable because lampreys share similar larval and spawning habitat requirements, and their larvae derive no apparent benefit from producing compounds that serve as an attractant for adults.

Hormonal pheromones in fish

Fish commonly exhibit reproductive responses to waterborne steroids, prostaglandins (PGs), and their metabolites (hormonal pheromones) – a logical result of a long evolutionary history spent in a medium that often reduces visual information but readily exposes the olfactory organ to information-rich chemicals released by conspecifics. Although fish from major taxa, including carps, catfishes, salmon, and gobies, respond with great sensitivity and specificity to a variety of hormonal pheromones, in no species are hormonal pheromones well understood. The phenomenon appears best characterized in goldfish, where steroid and PG pheromones released sequentially by periovulatory females dramatically affect male behavior, physiology, and reproductive fitness. Goldfish steroidal pheromones appear to involve only specializations of the receiving individual, a situation which these authors have termed chemical spying, whereas in other fish, where the sending individual is also specialized, hormonal pheromones evidently function in true communication. By demonstrating that sex steroids and PGs synchronize reproductive functions not only by endogenous hormonal actions but also by exogenous pheromonal actions, the relatively recent discovery of hormonal pheromones has considerably expanded our concepts of fish reproductive function while providing valuable model systems for future study of olfactory function and pheromone evolution.

The three steroidal components of the goldfish preovulatory pheromone signal evoke different behaviors in males.

The goldfish sex pheromone system is the best understood among the teleost fishes. Pheromones in this species are unspecialized hormonal products, which are released in ratios that vary with reproductive status. This study examined behavioral responses of male goldfish to three steroidal components of the female preovulatory pheromone: 17,20beta-dihydroxy-4-pregnen-3-one (1720betaP); 17,20beta-dihydroxy-4-pregnen-3-one-20-sulfate (1720betaP-S); and androstenedione (AD). Males were observed during exposure to nanomolar concentrations of each steroid over a 2-h period. We observed chasing, nudging (courtship behaviors) and pushing (an aggressive behavior). Each steroid elicited a different set of behaviors. 1720betaP, which is released by ovulatory females, elicited a low level of chasing and nudging that persisted throughout the experiment. Exposure to 1720betaP-S, which is released primarily by ovulatory females, triggered a large increase in nudging and chasing that lasted for only 5 min. In contrast, AD, which is released by females early in the ovulatory cycle and by mature males, elicited increases in aggressive behavior. 1720betaP and 1720betaP-S both caused increases in GtH-II release while AD did not. These results demonstrate that goldfish can discriminate components found in the female pheromone blend, suggesting that goldfish, and likely other fish species, may employ blends of hormonal products as pheromones.