Standardization of techniques and conditions for in vitro ovulation induction in Astyanax altiparanae

Abstract 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.