Reproductive stage- and sex-dependant effects of neurohypophyseal nonapeptides on gonadotropin subunit mRNA expression in the catfish Heteropneustes fossilis: An in vitro study.

In the present study, in vitro effects of synthetic vasotocin (VT), isotocin (4Ser, 8Ile- oxytocin; ITb) and the recently cloned IT gene paralog product (8Val-Isotocin, ITa) were studied on the expression of pituitary gonadotropin (GtH) subunit mRNA levels. In male pituitaries of early (preparatory phase) and late (prespawning phase) recrudescing catfish, Heteropneustes fossilis, VT (10 nM, 100 nM and 1000 nM) stimulated fshß expression dose-dependently. But in females, the dose-dependent effect was found only in the preparatory phase. In males, VT stimulated lhß expression only at higher doses. In females, VT produced a significant dose-dependent increase of the lhß expression only in the prespawning phase. VT stimulated the expression of gpα, dose-dependently in the preparatory phase in males and in the prespawning phase in females. The incubation of the pituitaries with ITb did not alter the fshß expression in either sex in both preparatory and prespawning phases. In males, ITb stimulated the expression of lhß and gpα only at the highest concentration (1000 nM) in both phases. In females, ITb stimulated both lhß and gpα expression only at 1000 nM in the preparatory phase and dose-dependently in the prespawning phase. The incubation of the pituitaries with ITa produced effects similar to ITb on the expression of fshß, lhß, and gpα. The results show that the basic peptide VT modulates both fshß and lhß expressions, which are influenced by the sex and reproductive stage. The neutral peptide ITA/ITb exerts an insignificant effect on the fshß expression regardless of sex or season. Both VT and ITa/ITb elicit a significant effect on the lhß expression in late recrudescent phase especially in females.

Effects of altered photoperiod and temperature on expression levels of gonadotrophin subunit mRNAs in the female stinging catfish Heteropneustes fossilis.

Differential effects of photoperiod and temperature on the temporal modulation of gonadotrophin subunit genes (glycoprotein α, gpα), follicle-stimulating hormone ß (fshß) and luteinizing hormone ß (lhß) expression were investigated in the stinging catfish Heteropneustes fossilis. Female H. fossilis were exposed to varying photoperiod and temperature conditions for 14 and 28 days in the early preparatory phase of the annual reproductive cycle. Gonadotrophin subunit gene expression, gonado-somatic index (IG ), ovarian histology and plasma steroid hormone levels were evaluated. The exposure of H. fossilis to long photoperiod (LP) of 16 h light or high temperature (HT) at 28 ± 2° C (mean ± s.e.), alone or in combination, resulted in significant increases in gpα, fshß and lhß messenger (m)RNA levels, IG , plasma oestradiol-17ß (E2 ), testosterone (T) and progesterone (P4 ) levels. The ovaries were filled with advanced yolky oocytes. On the other hand, the short photoperiod (SP) of 8 h light exposure decreased the transcript levels with higher inhibition in the normal temperature (NT) group at 18 ± 2° C (mean ± s.e.) than the HT group at 28 ± 2° C. Furthermore, the inhibition reached the highest level in total darkness (TD) of 24 h light deprivation under NT conditions at 18 ± 2° C. Consequently, the SP and TD treatments inhibited the IG , plasma E2 and T levels and ovarian development. The exposure to high temperature at 28 ± 2° C also modified the short photoperiod effect by elevating plasma E2 level. The plasma T level changed only mildly while the plasma P4 level showed the greatest fluctuations; the level reached the nadir in the SP + HT group but increased in the SP + NT group on day 28. A two-way ANOVA of the data showed differential effects of photoperiod and temperature; photoperiod produced a highly significant effect on fshß expression while temperature had a highly significant effect both on lhß and gpα levels. Thus, the differential expression of the gpα by the environmental variables ensures temporal synchronization of ovarian development and spawning.

Ovaprim, a commercial spawning inducer, stimulates gonadotropin subunit gene transcriptional activity: A study correlated with plasma steroid profile, ovulation and fertilization in the catfish Heteropneustes fossilis.

The commercial fish spawning inducer Ovaprim (OVP) containing a salmon gonadotropin-releasing hormone analogue and domperidone (a dopamine receptor-2 antagonist) has been widely used as an effective spawning inducer in artificial breeding of fishes. It induces a preovulatory LH surge resulting in final oocyte maturation (FOM) and ovulation through a mechanism involving a steroidogenic shift to secrete a maturation-inducing steroid (MIS). In the present study, a 0.5µL/g body weight dose of OVP each injected at 0h and 24h intraperitoneally into gravid female catfish, Heteropneustes fossilis resulted in periovulatory changes in gonadotropin (GtH) subunit gene expression and steroid hormone levels. The OVP injections induced ovulation time-dependently from 6h onwards with 100% ovulation recorded from 24h to 48h. The fertilization rate was high from 6h to 18h and declined from 24h onwards. The OVP treatment up regulated the expression of GtH subunit genes differentially. The expression of glycoprotein-α (GPα) and luteinizing hormone (LHß) peaked at 6h and 12h, and declined at 18h and 24h after the first injection. The second OVP injection at 24h elicited only a transient increase in the GPα expression at 6h and a sustained increase in the LHß expression from 6h to 18h after the second injection, but both transcripts decreased subsequently. The follicle-stimulating hormone (FSHß) expression responded to the OVP treatment from 12h onwards and maintained a constant level from 18h to 36h after the first injection; the second dose had little effect. Plasma steroids were differentially altered: the levels of estradiol-17ß decreased while that of the MIS 17,20ß-dihydroxy-4-pregnen-3-one; 17,20ß-DP increased, causing the steroidogenic shift preceding FOM and ovulation. The present results indicate that LHß expression coincides with the ovulation response and the late induction and maintenance of the FSH expression may be related to post-ovulatory events in the ovary.

Catfish gonadotrophins: cellular origin, structural properties and physiology.

Gonadotrophins (GTHs) play a central role in the regulation of gametogenesis and spawning. The structural duality of the GTHs [luteinising hormone (LH) and follicle-stimulating hormone (FSH)] is established in fishes with the exception of ancestral vertebrates. Most studies indicate that, in teleosts, the GTHs are secreted in separate cells. Phylogenetic analysis shows that the common α-subunit of the GTHs (and also of thyroid-stimulating hormone) and LHß are highly conserved in fishes, as in tetrapods. However, FSHß shows considerable divergence in teleosts. There may be 12 or 13 cysteine residues, with an additional one near the N-terminus. There may be one or two N-linked glycolsyation sites. In catfishes, there are 13 cysteine residues and one N-linked glycosylation site. In an extreme situation, a potential glycosylation site is lacking in some fishes. Both FSH and LH receptors are characterised in teleosts. The FSH receptor is promiscuous and can be cross-activated by LH. By contrast, the LH receptor is highly selective, being activated by its natural ligand or by heterologous ligands (e.g. human chorionic gonadotrophin). Consequently, teleosts show different patterns of LH and FSH secretion. In catfishes, in the absence of native FSH protein, LH controls all aspects of reproduction, from early gametogenesis to spawning.