The effect of Roundup on embryonic development, early foxr1 and hsp70 gene expression and hatching of common carp (Cyprinus carpio L.).

The effects of herbicide Roundup (based on glyphosate) on the embryonic development, survival and hatching of common carp (Cyprinus carpio L.) larvae and alteration in foxr1 and hsp70 gene expression were determined. The eggs (obtained from 6 females) were fertilised and incubated in water containing 0; 1 or 10 µl L-1 of Roundup formulation. During early embryonic development (24 and 48 h post-fertilisation - hpf), Roundup caused a statistically important decrease in the embryonic survival rate of common carp. Moreover, retardation of the hatching rate was observed in the group treated with the higher concentration of Roundup at 81 to 99 hpf. At the end of the experiment (99 hpf), an important increase in number of deformed larvae was observed in both groups treated with Roundup in comparison to the control group (52.06; 16.02 and 5.08%, respectively). Significant differences in transcript of the gene foxr1 were found in Roundup-intoxicated groups in comparison to the controls. In the case of hsp70 transcripts, no important changes in exposed groups were observed. These results showed that even small, environmentally relevant amount of Roundup present in the aquatic environment is able to affect the early life stages of common carp and change the transcripts of foxr1, which may have an adverse effect on the later proper development of the reproductive system.

The effects of ghrelin on the in vitro spontaneous and sGnRH-A stimulated luteinizing hormone (LH) release from the pituitary cells of common carp (Cyprinus carpio L.).

In fish, like in mammals, ghrelin affects gonadotropin release acting at the level of the hypothalamus as well as directly on the pituitary gland. In the present study, enzymatically dispersed pituitary cells obtained from sexually mature male and female carp (Cyprinus carpio L.) were incubated in the presence of human ghrelin at the concentration of 10(-7) or 10(-6) M, salmon GnRH analogue (Des-Gly(10), D-Arg(6), Trp(7), Leu(8), Pro(9))-LHRH (sGnRH-A) at the concentration of 10(-8) M or the combination of ghrelin (both concentrations) and sGnRH-A. ELISA method was used for carp LH levels determination in the media collected after 10 or 24 h of incubation. Ghrelin at the concentration of 10(-6) M caused the increase of the spontaneous LH secretion from female pituitary cells only. The combination of ghrelin (both concentrations) with sGnRH-A resulted in the significant elevation of LH levels in the incubations of both male and female pituitary cells in comparison with control incubations as well as with sGnRH-A alone treated cells. The results obtained in this study show that ghrelin functions as LH-stimulating hormone in common carp and that it acts directly on gonadotrophic cells, potentiating also the action of GnRH.

Influence of long-term exposure to dietary cadmium on growth, maturation and reproduction of goldfish (subspecies: Prussian carp Carassius auratus gibelio B.).

The influence of long-term exposure of goldfish to dietary cadmium (Cd) on its accumulation in tissues, growth, ovarian development, luteinizing hormone (LH) secretion and a response to hormonal stimulation of spawning were evaluated. The study was conducted on four groups of females for the period of 3 years, from the age of 10 weeks to second spawning. Four doses of Cd were applied in the feed: 0 (control group), 0.1, 1 and 10 mg Cd g(-1) of feed (wet weight). The highest dose of Cd (10 mg g(-1)) inhibited growth and caused several behavioural effects. In contrast, lower dose of Cd (1 mg g(-1)) stimulated fish growth. The doses of Cd from 0.1 to 1 mg Cd g(-1) did not influence ovarian development. The gonado-somatic index (GSI) and histological analysis of ovaries showed no differences in ovarian development between the control group and the groups receiving these doses of Cd. However, in the group receiving the highest Cd dose, GSI decreased. This was associated with persistent, long-lasting elevation of plasma LH levels. Ovulation did not occur in this group. Injections of salmon GnRH-analogue (sGnRHa) alone or with domperidone (a dopamine receptor antagonist) in sexually mature fish caused an increase of LH levels in all groups, although in the group fed with the highest Cd dose the effect was weaker than in the other groups. After the first spawning season, a negative effect of lower Cd doses (0.1 and 1mg Cd g(-1)) on ovarian recrudescence (rebuilding of ovaries) and on the response to the consecutive hormonal stimulation of spawning was observed (lower number of ovulating females). There was a significantly higher content of Cd in the livers of fish than in their muscles. The results of hormonal stimulation of spawning and histological analysis of ovaries suggest that in goldfish cadmium acts mainly at the level of ovary rather than on the pituitary gland. We suppose that in the natural environment cadmium present in the feed can play an important role in the accumulation of this element in fish tissues and can influence vital physiological processes.

The influence of gabaergic drugs upon LH secretion from the pituitary of the common carp: an in vitro study.

In the present study, the perifusions of whole pituitary glands of spermiating male common carp were performed in the presence of several GABAergic drugs. Muscimol (agonist of GABA(A) receptors) and bicuculline (the antagonist of the same type of GABA receptors) did not modify basal LH release. LH basal secretion was not modified when pituitaries were perifused with baclofen--an agonist of GABAB receptors. On the other hand, baclofen at doses of 10(-8) and 10(-4) M significantly decreased GnRH-A-induced LH release to about 86% and 88% of LH levels in control group, respectively. In our previous study we have shown that GABA decreased basal and GnRH-A-stimulated in vivo and in vitro LH release. In conclusion, it can be suggested that in the mature male carp GABA exerts an inhibitory influence on GnRH-stimulated LH release, probably through the inhibition of the GnRH action on gonadotropes. This inhibition seems to be mediated by the B type of GABA receptors.

Substrate concentration affects the in vitro metabolism of 17-hydroxyprogesterone by ovaries of the carp, Cyprinus carpio.

Carp ovarian tissue was incubated with (3)H-17-hydroxyprogesterone in the presence of 0, 0.1, 1, 10, and 100 µg ml(-1) unlabeled 17-hydroxyprogesterone. The pattern of metabolites formed showed a marked variation with substrate concentration. Formation of glucuronide and sulphate conjugates was important only at low substrate concentration. At high substrate concentration (10 and 100 µg ml(-1)) 17,20α-dihydroxy-4-pregnen-3-one was the major metabolite, but at intermediate concentrations polar 7α-hydroxypregnanetetrols predominated. The results support the hypothesis that at low substrate concentrations conjugating, 5α-reducing and 7α-hydroxylating enzymes, of high activity but low capacity, act as scavengers to deactivate any steroids formed during the relatively low pituitary gonadotrophin secretions which are necessary for oocyte development, but that during the prespawning gonadotrophin surge when high levels of substrate are present these enzymes are saturated and 17,20α-dihydroxy-4-pregnen-3-one (17,20αP) becomes the major ovarian steroid. The possible role of 17,20αP during oocyte final maturation requires further examination.

Effects of N,N-dimethylnitrosamine (DMNA) on in vitro oocyte maturation and embryonic development of fertilized eggs of carp (Cyprinus carpio L.) kept in eutrophied ponds.

Toxic effects of N,N-dimethylnitrosamine (DMNA) at doses of 100 or 500 micrograms l-1 on in vitro carp oocyte maturation (steroidogenesis), embryonic development and hatching of larvae (obtained as a result of artificial spawning of females kept for four seasons in normal and eutrophicated ponds) were investigated. There were no significant effects of DMNA on oocyte maturation and steroidogenesis during 24 h of incubation. The DMNA decreased the hatching of fertilized eggs derived from control females. This decrease reached a level of significance at a dose of 500 micrograms l-1. However, the effect of long-term exposure of female fish to eutrophied water was very much higher. The trend of the in vitro DMNA effect was the same, but it did not reach a statistically significant level. The results suggest that nitrosamines, through their effect on egg hatchability, may reduce fish populations along with increasing aquatic eutrophication.