We have previously shown an increase on the ovulation rate of pacu, Piaractus mesopotamicus, using hypophysation with prostaglandin F-2 alpha (PGF(2 alpha)) (HPG protocol) rather than hypophysation (H); only, however, a persistent lack of ovulation in some females could not be further explained by PGF(2 alpha) and steroid plasma levels at time of ovulation. Considering this, here we compared the meiotic process and steroids and PGF(2 alpha) levels during spawning induction using both protocols. To that, H and HPG were treated with conventional hypophysation (0.6 mg/kg priming and 5.4 mg/kg resolving dose), and the latter received synthetic PGF(2 alpha) (2 mL/fish) at resolving dose. A negative control (saline solution), a SPG group (receiving only PGF(2 alpha)) and a PPG group (receiving priming and PGF(2 alpha)) were included. An initial PGF(2 alpha) peak between one to four hours post-resolving dose (1-4 hr) occurred in groups receiving PGF(2 alpha) (HPG, SPG and PPG). Only H and HPG showed a 17 alpha-20 beta-dihydroxy-4-pregnen-3-one (DHP) peak 4 hr, concomitant with meiosis resumption, and a final PGF(2 alpha) peak 12 hours. The HPG protocol resulted in higher PGF(2 alpha) plasma levels for longer periods (1-4 and 12 hr) than H (single peak 12 hr), occasioning in the former higher and concomitant levels of both PGF(2 alpha) and DHP at 4 h. Considering that DHP is known to induce expression of prostaglandin receptors in fish, the concomitant plasma peaks of DHP and PGF(2 alpha) may also explain the ovulation rate gain previously published, using HPG protocol, which can be further improved adjusting the priming dose and defining the exact interval of PGF(2 alpha) ovulatory action in this species.

This study aimed to understand how germ cell development occurs in females of Devario aequipinnatus, by morphologically describing oogenesis and the reproductive phases. Sexually mature females of D. aequipinnatus (n = 70) were obtained from commercial fisheries and delivered to the Laboratório de Ictiologia Neotropical, UNESP, Ilha Solteira, SP, Brazil. The ovaries were removed, fragmented and fixed following the usual techniques for light microscopy. The stages of ovarian development in D. aequipinnatus begin with the oogonia, which proliferate into new cells or differentiate into prophasic oocytes that, at the end of this process, form the ovarian follicle and end folliculogenesis. In the previtellogenic stage, the oocytes were characterized mainly by the gradual loss of basophilia and an increase in oocyte diameter. Vitellogenesis was marked mainly by the incorporation of yolk granules. Mature oocytes were defined by their migration from the nucleus to the micropyle. Postovulatory follicles and atresic oocytes were also observed. The reproductive phases were classified as: immature, early and final developing, spawning capable, regressing and regenerating. Therefore, the development of an understanding of cell modifications that occurs up to oogenesis is a basic step that is essential for the description of the reproductive biology of D. aequipinnatus, given the lack of information about the reproductive aspects of this species.

This study aimed to establish in vitro culture conditions for inducing ovulation in lambari (Astyanax altiparanae). All fish received a priming dose of either 0.6 mg/kg or 100 IU hCG/kg (human chorionic gonadotropin) in vivo. In experiment 1, 0.6 mg and 5.4 mg of carp pituitary extract (CPE)/ kg of ovarian fragment were tested as resolving doses. In experiment 2, prostaglandin F2α (PGF2α) analogue at 100 ng/mL was added to 5.4 mg CPE/kg as the resolving dose. Since ovulation did not occur, in experiment 3 and 4, we compared diverse forms of obtaining follicles, comparing manual follicle dissociation and collagenase at 100, 200, and 400 CDU/mL. Additionally, in experiment 4, the resolving dose of CPE and hCG was replaced by 1 µg/mL 17α,20β-dihydroxy-4-pregnen-3-one (DHP). Ovulation was successful only in Experiment 4, using mechanically dissociated follicles with a priming dose of either CPE or hCG and 1 µg/mL DHP as the resolving dose. Key findings include that 5% CO2 is unnecessary, mechanical dissociation of ovarian fragments is optimal, a priming dose of CPE or hCG is required, and DHP at 1 µg/mL is effective. These results establish a standardized protocol for in vitro ovulation induction in A. altiparanae, offering a valuable tool to study ovarian function and spawning failure in tropical species.

Melatonin (MTN) plays a direct role in fish ovulation, and we hypothesized that aligning ovulation with an endogenous MTN peak levels could improve reproductive performance. To that, in experiment 1, we assessed circadian variation of plasma MTN in pacu (Piaractus mesopotamicus) females. In experiment 2, pacu females were divided into two groups: one receiving the hypophysation dose at 7 pm (dark onset) and the other at midnight (five hours after dark). Experiment 1 showed that circulating MTN levels increased at 7 pm and remained stable through the dark phase. In experiment 2, no differences in latency or reproductive performance were observed between groups. However, MTN levels at ovulation increased significantly only in the 7 pm group. Strong positive correlations between MTN levels at ovulation and reproductive parameters, such as fertility, hatching rates, and fecundity, were observed. These results confirm that PGF2α and DHP peaks at ovulation are associated with successful ovulation in pacu, consistent with literature that shows that MTN influences ovulation by acting through the Mtnr1a receptor, triggering arachidonic acid release and prostaglandin synthesis. Our findings indicate that the timing of spawning induction influences MTN levels at ovulation, which are positively correlated with reproductive success. This is the first report linking plasma MTN levels at ovulation to successful reproductive performance in fish induced by hypophysation, highlighting the relevance of spawning timing for pacu reproduction.

This study evaluated the potential of lambari (Astyanax altiparanae) as a pituitary donor for hormone-induced spawning in fish. The effectiveness of the extracted hormone was demonstrated over four consecutive years, successfully inducing spawning in captive lambari as well as in two other rheophilic species, curimbatá (Prochilodus lineatus) and tambaqui (Colossoma macropomum). When administered in the same manner as crude carp pituitary extract, a total dose of 6 mg·kg⁻¹, divided into two applications (10% + 90%), proved to be effective, since spawning was obtained in the three tested species. Under the applied management conditions, donor fish maintained a gonadosomatic index of approximately 15%, with ovaries predominantly composed of mature vitellogenic oocytes. These females exhibited basal levels of estradiol (around 0.9 ng.ml-1) and 17,20β-dihydroxy-4-pregnen-3-one (17,20β-P) (around 0.2 ng.ml-1), along with stable lhβ expression levels among different baths and between males and females. The lambari pituitary extract, when injected into females of the same species, induced a significant increase in 17,20β-P levels from approximately 0.2 to 1.0 ng·ml⁻¹. The use of lambari pituitary extract confirms that donor fish, both males and females, must be reproductively mature and in the breeding season. To our knowledge, most previous studies on extract donors are based on wild donors, making this one of the few that describe their physiological characteristics under aquaculture conditions. Low rearing densities seem to be crucial for obtaining a potent extract, although fish from the grow-out phase may also be suitable, a possibility that still requires evaluation. Our findings highlight the potential of lambari pituitary glands as a valuable alternative for hormone-induced spawning in small and medium-sized fish species, supported by their rapid maturation (4 months), low trophic level, increasing commercial production, and the prospect of using canned lambari to efficiently recover pituitary glands from fish heads.