Courtship and spawning behaviour of medaka in a semi-outdoor environment initiating at midnight.

Reproductive timing is a critical ecological trait that directly influences fitness. Medaka (Oryzias latipes), a small freshwater fish, is widely used as a model organism in various scientific fields. However, ecological studies conducted under (semi-) natural conditions remain limited. Although, spawning has been reported to occur within 1 h before and after sunrise, direct observations remain scarce. We investigated the timing of spawning initiation and associated courtship in medaka through 24-h observations using infrared cameras under semi-natural conditions. During the experiments, sunrise occurred at approximately 4:45. Observations of the 31 pairs revealed that spawning occurred between 1:05-9:48, with a peak at 2:00-4:00. Unlike previous reports but like the most recent fieldwork, only 26% of the total spawning events (8/31) were observed within 1 h before and after sunrise. Male courtship behaviours, including following females and quick circle displays, increased from midnight, peaking between 2:00-5:00. This study provides new insights into the natural reproductive timing of medaka, suggesting a possible adaptation to nocturnal spawning, likely as a strategy to reduce predation on both parents and eggs. It also underscores the importance of investigating the ecology of model organisms under (semi-)natural conditions to gain a more comprehensive understanding of biological phenomena observed in laboratory settings.