Long-Term Exposure of the Red Cherry Shrimp Neocaridina davidi to Diclofenac: Impact on Survival, Growth, and Reproductive Potential.

The current study was aimed at studying the long-term effects of diclofenac on the freshwater shrimp Neocaridina davidi, concerning survival, somatic growth, and reproduction. In this study, both ovigerous females and males of this species were exposed for 63 d to 0 (control), 0.1, or 1 mg/L of diclofenac. At the highest concentration, significant mortality was detected, and the somatic growth of females was significantly decreased. The percentage of females with a second spawn, observable from day 45, significantly increased at 1 mg/L, while the time between spawns was significantly reduced at both concentrations assayed. However, the gonadal analysis made at the end of the assay in the surviving females showed a significantly lower proportion of advanced oocytes in females exposed to 1 mg/L, as compared to control. Concerning hatching, the percentage of ovigerous females that could have successful hatching was reduced at 1 mg/L of diclofenac, especially for the first spawn. For the second spawn, the low number of juveniles hatched from females exposed to 1 mg/L also showed a significantly higher incidence of morphological abnormalities, such as hydropsy and underdeveloped appendages. Taken together, these results showed that even when diclofenac was able to produce earlier spawns, the reproductive output of each spawn was reduced.

Effects of atrazine on vitellogenesis, steroid levels and lipid peroxidation, in female red swamp crayfish Procambarus clarkii.

Atrazine, a widely use herbicide, has been classified as a potential endocrine disruptor, especially for freshwater species. In this study, we tested the hypothesis that atrazine can affect reproduction in crayfish through dysregulation of vitellogenin expression and hormone synthesis. Adult female crayfish (Procambarus clarkii) were exposed during one month to atrazine at concentrations of either 1 or 5 mg/L. At the end of the exposure, ovaries, hepatopancreas, and hemolymph samples were harvested for analysis of vitellogenin expression and steroid hormone levels. Ovarian tissue was also sampled for both biochemical and histological analyses. Our results show that atrazine-exposed crayfish had a lower expression of vitellogenin in the ovary and hepatopancreas, as well as smaller oocytes, and reduced vitellogenin content in the ovary. Despite these effects, circulating levels of estradiol increased in females exposed to 5 mg/L of atrazine, showing that the inhibiting effect of atrazine on vitellogenin production was not related to a lower secretion of sexual steroids. Instead, some early stimulating effects of estradiol on vitellogenesis could have occurred, particularly in the hepatopancreas. On the other hand, atrazine caused a higher metabolic effort, in terms of lactate production, presumably triggered to provide the energy needed to face the unspecific stress produced by the herbicide. Lipid peroxidation was not affected by atrazine, but glutathione levels were significantly increased.

Effects of glyphosate on egg incubation, larvae hatching, and ovarian rematuration in the estuarine crab Neohelice granulata.

Ovigerous females of the estuarine crab (Neohelice granulate) were exposed to both pure glyphosate (2.5 mg/L and 5 mg/L) and a glyphosate formulation (Roundup Ultramax, containing glyphosate at 2.5 mg/L acid equivalent). At the end of the egg incubation period, a significant reduction in the number of hatched larvae was seen as a result of Roundup exposure. Additionally, several larvae abnormalities were seen in both pure glyphosate (2.5 mg/L) and Roundup treatments, such as hydropsy and hypopigmented eyes, and atrophied eyes were observed in the Roundup treatment. To evaluate the effect of the herbicide on ovarian rematuration, females remained exposed for 32 d. Pure glyphosate at 2.5 mg/L stimulated ovarian maturation over control levels, mainly in terms of a higher gonadosomatic index and a higher percentage of vitellogenic oocytes. A plausible hypothesis to be tested in further experiments is that exposure to glyphosate disrupts the hormonal system controlling reproduction.

In vitro effects of both dopaminergic and enkephalinergic antagonists on the ovarian growth of Cherax quadricarinatus (Decapoda, Parastacidae), at different periods of the reproductive cycle.

The in vitro effect of both spiperone (dopaminergic antagonist) and naloxone (enkephalinergic antagonist), was assayed on small pieces of ovary dissected from C. quadricarinatus females, with the eventual addition of some neuroendocrine organs, such as thoracic ganglion or eyestalk tissue. The incorporation of tritiated leucine by the ovary was measured in order to estimate the ovarian growth. During the post-reproductive period, both mentioned antagonists were able to significantly stimulate the ovary in the presence of thoracic ganglion, but did not produce any significant effect in the preparation containing ovary and eyestalk tissue, or only ovary. No significant effects of the assayed antagonists were noted during the pre-reproductive period. These results were in accordance with previous models describing the neuroendocrine control of crustacean reproduction, and represent new findings about the hormonal context in different periods of the reproductive cycle of crayfish. Besides, by means of the experimental combination of the tested antagonists with dopamine or met-enkephalin, a new model dealing with the interaction of these two neurotransmitters on the hormonal secretion of thoracic ganglion has been proposed.

Interference of cadmium and copper with the endocrine control of ovarian growth, in the estuarine crab Chasmagnathus granulata.

The effects of cadmium and copper on the hormonal control of ovarian growth were evaluated on the estuarine crab Chasmagnathus granulata, by means of both in vivo (14 days exposure) and in vitro (24 h) assays. For both kind of assays, heavy metal concentrations of 0 (control), 0.5 mg/L of cadmium or 0.1 mg/L of copper were used. No significant (P > 0.05) change of the gonadosomatic index was observed in the in vivo assays with intact females exposed to heavy metals, while eyestalk-ablated exposed females showed significantly (P < 0.05) lower gonadosomatic index values than their respective controls. This latter result led us to consider the possibility that the interfered with extra-eyestalk hormones. In this sense, no differences were noted between control and heavy metals-exposed groups after co-incubating ovary with thoracic ganglion (the source of the gonad stimulating hormone). However, when ovary was incubated with methyl farnesoate or 17-hydroxyprogesterone, 3H-leucine incorporation was significantly lower in the heavy metals-exposed groups than in the controls, indicating a possible interference of cadmium and copper with the transduction pathway of those hormones. On the other hand, ovaries co-incubated in vitro with eyestalk tissue and exposed to either heavy metal showed significantly higher 3H-leucine incorporation than did the controls, suggesting an inhibitory effect of both heavy metals on the secretion of the gonad inhibiting hormone from the eyestalk tissue. Interference by copper and cadmium with the transduction mechanisms of gonad inhibiting hormone at the ovarian level does not appear to be a viable hypothesis, because the addition of eyestalk extracts to the incubation medium reversed the effect caused by each heavy metal. The results from the in vitro assays were in accordance with those obtained with the intact crabs in vivo.

Inhibition of molting by cadmium in the crab Chasmagnathus granulata (Decapoda Brachyura).

The effect of cadmium on molting of the estuarine crab Chasmagnathus granulata was assayed. Adult males were eyestalk-ablated, in order to synchronically induce molting, and were then chronically exposed to cadmium at 0.25 or 0.50 mg/l. At the highest concentration, a significant mortality was detected at the time of molting, in the few crabs that could reach the E stage. However, most of the crabs exposed from the beginning of the premolt period (D(0) stage) to 0.50 mg/l of cadmium were arrested at the D(1)" stage. This effect was not seen when crabs were exposed to the same cadmium concentration from either D(1)"' or D(3) premolt stages. Crabs arrested by cadmium did not present any difference in the calcium content of carapace, compared to controls, while ecdysteroid levels of those crabs were similar to the ones of control crabs that were in the same premolt stage but could finally molt. These results suggest that cadmium could be preventing the normal peaking of ecdysteroids needed for molting. Since eyestalk-ablated crabs were used, a presumably direct effect of cadmium on Y-organ seems likely, by affecting cytoplasmatic calcium concentration and/or other actions.