Persistent organic pollutants in green sea turtles (Chelonia mydas) inhabiting two urbanized Southern California habitats.

Within Southern California, east Pacific green sea turtles (Chelonia mydas) forage year-round, taking advantage of diverse food resources, including seagrass, marine algae, and invertebrates. Assessing persistent organic pollutants (POP) in green turtle aggregations in the Seal Beach National Wildlife Refuge (SBNWR, n = 17) and San Diego Bay (SDB, n = 25) can help quantify contamination risks for these populations. Blood plasma was analyzed for polychlorinated biphenyls (PCBs), organochlorinated pesticides (OCPs), and polybrominated diphenyl ethers (PBDEs). PCBs and body size explained much of the separation of turtles by foraging aggregation in a principal component analysis. Turtles from SDB had significantly (p < 0.001) higher total PCBs than SBNWR turtles. Most PCBs detected in turtles were non-dioxin-like PCB congeners (153, 138, 99) that are associated with neurotoxicity. Recaptured turtles' POP levels changed significantly over time indicating significant variation in POP levels through time and space, even among adjacent foraging locations.

Fecal stanols show simultaneous flooding and seasonal precipitation change correlate with Cahokia's population decline.

A number of competing hypotheses, including hydroclimatic variations, environmental degradation and disturbance, and sociopolitical disintegration, have emerged to explain the dissolution of Cahokia, the largest prehistoric population center in the United States. Because it is likely that Cahokia's decline was precipitated by multiple factors, some environmental and some societal, a robust understanding of this phenomenon will require multiple lines of evidence along with a refined chronology. Here, we use fecal stanol data from Horseshoe Lake, Illinois, as a population proxy for Cahokia and the broader Horseshoe Lake watershed. We directly compare the fecal stanol data with oxygen stable-isotope and paleoenvironmental data from the same sediment cores to evaluate the role of flooding, drought, and environmental degradation in Cahokia's demographic decline and sociopolitical reorganization. We find that Mississippi River flooding and warm season droughts detrimental to agriculture occurred circa (ca.) 1150 CE and possibly generated significant stress for Cahokia's inhabitants. Our findings implicate climate change during the Medieval Climatic Anomaly to Little Ice Age transition as an important component of population and sociopolitical transformations at Cahokia, and demonstrate how climate transitions can simultaneously influence multiple environmental processes to produce significant challenges to society.

Effects of environmental exposure to diazepam on the reproductive behavior of fathead minnow, Pimephales promelas.

Pharmaceutical drugs are continuously discharged into the aquatic environment primarily through wastewater discharge; therefore, their possible effects on wildlife is a reason of concern. Diazepam is a widely prescribed benzodiazepine drug used to treat insomnia and anxiety disorders, and it has been found in wastewater effluents worldwide. The present study tested the effects of diazepam on fecundity and the reproductive behavior of the fathead minnow, Pimephales promelas, a fish that exhibits male parental care. Sexually mature fathead minnows were housed at a ratio of one male and two females per tank and exposed to nominal (measured) concentrations of 0, 0.1 (0.14 ± 0.06), 1.0 (1.04 ± 0.15), 10 (13.4 ± 1.5) µg L(-1) for 21 days. Fish receiving the low diazepam treatment had significantly larger clutches than fish receiving the highest concentration but neither were different from controls. Diazepam exposure was not associated with a significant change in fertilization rate, hatchability or time to hatch, but a trend toward a higher number of eggs/day was observed in fish exposed to the low diazepam concentration relative to those exposed to the medium concentration. There were no significant differences in any of the behaviors analyzed when responses were averaged over time. The results showed that exposure to diazepam at concentrations as high as 13 µg L(-1) did not significantly impact the reproductive behavior of fathead minnow.

Differential responses of brain, gonad and muscle steroid levels to changes in social status and sex in a sequential and bidirectional hermaphroditic fish.

Sex steroids can both modulate and be modulated by behavior, and their actions are mediated by complex interactions among multiple hormone sources and targets. While gonadal steroids delivered via circulation can affect behavior, changes in local brain steroid synthesis also can modulate behavior. The relative steroid load across different tissues and the association of these levels with rates of behavior have not been well studied. The bluebanded goby (Lythrypnus dalli) is a sex changing fish in which social status determines sexual phenotype. We examined changes in steroid levels in brain, gonad and body muscle at either 24 hours or 6 days after social induction of protogynous sex change, and from individuals in stable social groups not undergoing sex change. For each tissue, we measured levels of estradiol (E(2)), testosterone (T) and 11-ketotestosterone (KT). Females had more T than males in the gonads, and more E(2) in all tissues but there was no sex difference in KT. For both sexes, E(2) was higher in the gonad than in other tissues while androgens were higher in the brain. During sex change, brain T levels dropped while brain KT increased, and brain E(2) levels did not change. We found a positive relationship between androgens and aggression in the most dominant females but only when the male was removed from the social group. The results demonstrate that steroid levels are responsive to changes in the social environment, and that their concentrations vary in different tissues. Also, we suggest that rapid changes in brain androgen levels might be important in inducing behavioral and/or morphological changes associated with protogynous sex change.

Effects of propranolol on heart rate and development in Japanese medaka (Oryzias latipes) and zebrafish (Danio rerio).

Propranolol is a ß-adrenergic receptor antagonist (ß-blocker) that is frequently used to treat hypertension and other cardiovascular conditions in humans. Detected in surface waters due to discharge of domestic wastewater, propranolol has demonstrated significant species differences in toxicity between fish. The aim of this study was to investigate the effects of propranolol on heart rate and development in embryos of two species of fish; Japanese medaka (JM) Oryzias latipes and zebrafish (ZF) Danio rerio. Parents and fertilized embryos of each species were exposed to nominal (measured) concentrations of 0.1 (0.09), 1 (1.1) and 10 (8.3) µg/L of propranolol. Heart rate was monitored during subsequent exposure in embryos at incremental developmental periods (44, 54, 64 h post-fertilization (hpf) for ZF and 68, 116, 164 hpf for JM). Heart development and morphology was examined using whole mount immunostaining with distance measurements between the sinus venosus (SV) and bulbus arteriosis (BV). Morphological measurements were made at 44 hpf for ZF and 164 hpf for JM. In ZF, a significant reduction in heart rate was observed at 0.08 µg/L propranolol, along with an increase in the SV-BA distance at 44 hpf. Significant reductions in heart rate were also observed in ZF at 54 and 64 hpf at all concentrations of propranolol. For JM, heart rates generally decreased at all developmental timepoints (68, 116 and 164 hpf) after propranolol treatment, with concentration dependent decreases observed at 164 hpf and a lowest observed effect concentration (LOEC) of 0.09 µg/L propranolol at each timepoint. However, significant alterations in cardiac morphology were not observed in JM at 164 hpf. In contrast, heart rates and morphology in ZF were affected with a non-monotonic concentration response in morphology and a LOEC of 0.09 µg/L propranolol for morphological alterations at 44 hpf and for heart rate at each timepoint. These data indicated unique developmental stages of susceptibility between species and that combined parental and embryo exposures may lead to greater impairment of cardiac development and function in offspring than separate exposures of adults and embryos.

Effects of exposure to the ß-blocker propranolol on the reproductive behavior and gene expression of the fathead minnow, Pimephales promelas.

Human pharmaceutical drugs have been found in surface waters worldwide, and represent an increasing concern since little is known about their possible effects on wildlife. Propranolol is a common beta-adrenergic receptor antagonist (ß-blocker) typically prescribed to people suffering from heart disease and hypertension. Propranolol has been detected in United States wastewater effluents at concentrations ranging from 0.026 to 1.90 µg/l. In mammals, there is evidence that ß-blockers can cause sexual dysfunction, and alter serotonergic pathways which may impact reproductive behavior but little is known about the effects on fish behavior. The present study tested the effects of propranolol on fecundity, on brain gene expression and on reproductive behavior of the fathead minnow, Pimephales promelas, a fish that exhibits male parental care. Sexually mature fathead minnows were housed at a ratio of one male and two females per tank and exposed to nominal concentrations of 0, 0.1, 1, 10 µg/l for 21 days. Measured concentrations (±SD) of propranolol were 0.003±0.004, 0.05±0.02, 0.88±0.34 and 4.11±1.19 µg/l. There were no statistically significant differences in fecundity, fertilization rate, hatchability and time to hatch. Propranolol exposure was not associated with a change in nest rubbing behavior, time spent in the nest or approaching the females. There was a significant difference in the number of visits to the nest with males receiving low and medium propranolol treatments. The microarray analysis showed that there were 335 genes up-regulated and 400 genes down-regulated in the brain after exposure to the highest dose of propranolol. Among those genes, myoglobin and calsequestrin transcripts (fold change=10.84 and 5.49, respectively) were highly up-regulated. Ontological analyses indicated changes in genes involved in calcium ion transport, transcription, proteolysis and apoptosis/anti-apoptosis. Pathway analysis indicated that the reduced expression of caspases may lead to impaired neurite outgrowth, neurotransmitter secretion and brain function in developing organisms. The results showed that exposure to propranolol at concentrations as high as 4.11 µg/l did not significantly impact reproductive behavior or spawning abilities of fathead minnow but did alter the regulation of genes within the brain of fish.

Immunohistochemical localization of serotonin in the brain during natural sex change in the hermaphroditic goby Lythrypnus dalli.

The neurotransmitter serotonin (5-HT) may play a central role in the inhibition of socially regulated sex change in fish because of its known modulation of both aggressive and reproductive behavior. This is the first study to use immunohistochemical techniques to examine the morphometry of serotonergic neurons at different times during sex change. Using a model species wherein sex change is socially regulated via agonistic social interactions (the bluebanded goby, Lythrypnus dalli), we sampled brains of males and females with different social status, and of females at different times during sex change. Consistent with previous studies on other teleosts, immunoreactive neurons were found in the posterior periventricular nucleus (NPPv), the nucleus of the lateral recess (NRL), the nucleus of the posterior recess (NRP) and in the raphe nucleus. We measured the total area of NPPv, NRL, NRP, and the number and mean cell area of serotonergic neurons in the raphe nucleus. There was no significant difference in any of the brain regions between males, females or sex changing fish, but there was a slight increase in the number of dorsal raphe neurons in the brain of sex changers 2h after male removal. The results show that in L. dalli the serotonergic system does not present any morphological sex and status differences, nor any dramatic modifications during sex change. These data, together with previous results, do not support the hypothesis that serotonin inhibits socially regulated sex change.

Serotonin, social status and sex change in the bluebanded goby Lythrypnus dalli.

In a variety of vertebrates, highly aggressive individuals tend to have high social status and low serotonergic function. In the sex changing fish Lythrypnus dalli, serotonin (5-HT) may be involved as a mediator between the social environment and the reproductive system because social status is a critical cue in regulating sex change. Subordination inhibits sex change in L. dalli, and it is associated with higher serotonergic activity in other species. We tested the hypothesis that high serotonergic activity has an inhibitory effect on sex change. In a social situation permissive to sex change, we administered to the dominant female implants containing the serotonin precursor 5-hydroxytryptophan (5-HTP). In a social situation not conducive to sex change, we administered either the serotonin synthesis inhibitor p-chlorophenylalanine (PCPA) or the 5-HT(1A) receptor antagonist p-MPPI. After three weeks we used HPLC to measure brain levels of 5-HT and its metabolite 5-hydroxyindoleacetic acid (5-HIAA). We also performed PCPA, p-MPPI and fluoxetine injections in size-matched pairs of females to assess its effect on dominance status. Males and newly sex changed fish showed a trend for higher levels of 5-HIAA and 5-HT/5-HIAA ratio than females. The different implants treatments did not affect the probability of sex change. Interestingly, this species does not seem to fit the pattern seen in other vertebrates where dominant individuals have lower serotonergic activity than subordinates.

Variability of GnRH secretion in two goby species with socially controlled alternative male mating tactics.

Male reproductive phenotypic plasticity related to environmental-social conditions is common among teleost fish. In several species, males adopt different mating tactics depending on their size, monopolizing mates when larger, while parasitizing dominant male spawns when smaller. Males performing alternative mating tactics are often characterized by a strong dimorphism in both primary and secondary reproductive traits. According to studies on sex-changing species and on species where only one male morph is reproductively active, male alternative phenotypes are expected to vary also in gonadotropin-releasing hormone (GnRH) neurons in forebrain preoptic area (POA). Here, we compared the intra- and inter-sexual variations in number and size of GnRH neurons, along with gonads and male accessory structure investment, in two goby species, the grass goby, Zosterisessor ophiocephalus, and the black goby, Gobius niger, characterized by male alternative mating phenotypes. In both species, older and larger males defend nests, court and perform parental care, while younger and smaller ones try to sneak territorial male spawning. We found that grass goby and black goby have different patterns of GnRH expression. Grass goby presents a clear intra-sexual dimorphism in GnRH expression, related to the occurrence of alternative mating tactics, while in the black goby, only inter-sexual differences are observed. The inter- and intra-specific variability in the GnRH neurons in these two goby species is discussed in light of the differences in migratory behavior, nest type, and mating system.

Changes along the male reproductive axis in response to social context in a gonochoristic gobiid, Zosterisessor ophiocephalus (Teleostei, Gobiidae), with alternative mating tactics.

Sexual selection has given rise, in several taxa, to intrasexual variation in male phenotype. While evolutionary studies have provided explanations of the adaptive function of this dramatic male phenotypic diversity, the proximate control of its expression has still to be completely understood. Several observations, primarily from sex-changing species, indicated a major role of social interactions in reproductive axis regulation and consequently in the expression of alternative male phenotypes. Here we documented changes along the male reproductive axis in response to social context in a gonochoristic species, the grass goby Zosterisessor ophiocephalus, where fully functional alternative male mating tactics appear to be expressed as an ontogenetic gradient. In the grass goby, larger and older males dig a nest and perform parental care, while smaller males sneak fertilization during territorial male spawning. Territorial males are characterized by a higher number of gonadotropin-releasing hormone (GnRH) neurons in forebrain preoptic area, smaller testes, larger seminal vesicles, and viscous ejaculates that last longer and contain fewer sperm than those of sneakers. To experimentally investigate the role of social factors in inducing changes along the male reproductive axis, sneakers were tested in two different situations: nesting alone or with ripe females. Sneakers that mated and performed parental care showed dramatic changes in brain, reproductive apparatus morphology, and ejaculate traits. GnRH-immunoreactive cells in forebrain preoptic area increased in number, reaching values typical of wild-caught parental males. Testes size decreased while seminal vesicle size increased and ejaculates showed lower sperm densities. These results were discussed within the framework of the social transduction hypothesis, which predicts that social experience should mediate, through a cascade of internal processes, shifts between morphs throughout life.