Parental influences on offspring phenotype occurring through pathways other than via inherited DNA sequences are known as parental effects. Parental effects profoundly influence offspring behaviour, including behaviour laterality and personality, two traits that are widespread and of fundamental importance in the animal kingdom with clear fitness consequences. However, the impact of parental effects on the interaction between behavioural laterality and personality within the same species has not been previously explored. If such a link exists, it would deepen our understanding of personality traits, extending them to brain laterality and its underlying neurobiology. In addition, if both traits are causally linked, it may constrain evolution as changing one of these traits would affect the other.  The aim of this study was to examine whether offspring personality traits and behavioural laterality are related and can simultaneously be influenced by parental effects, suggesting a common underlying mechanism. Here we exposed parents of an egg laying species, the stickleback (Gasterosteus aculeatus), to predator cue or not and examined the impact of this exposure on the two behavioural traits of their offspring. Shortly after laying, clutches were split: half were reared without predator cues for 12 weeks before behavioural testing, and the other half were used for cortisol analysis. We found that both parents and offspring from predator-exposed parents were bolder, with the offspring more likely to show lateralized behaviour, and were smaller than offspring from parents that were not exposed to predation. The egg cortisol levels were too low to be detected by LC/MS-MS. To our knowledge, these results are the first to indicate that parental effects under varying predation conditions can influence laterality, personality, and growth of offspring within the same individuals, though further evaluation and experiments are needed to determine the role of maternal cortisol.

We observed the response of the adult fish  to a predator model under controlled lab conditions. We tested 26 fish from the predation treatment and 24 from the control. These fish were moved from their home-pond a day before the test and placed into one of four aquaria (80x35x40 cm) to allow them to acclimatize.

 A focal fish was selected from the housing aquarium and gently placed in a novel experimental tank (40x30x40 cm). The experimental tank was furnished with a tube as shelter (5x10 cm), gravel on the floor and a tethered perch model of which the movement was remotely controlled by hand (Fig.2). To further simulate the presence of predators in the environment, we added Perch odour (5ml) to the test aquarium. The experimental tank was positioned in an illuminated wooden box to avoid external disturbance. The duration of the experiment was 10 mins and the behaviour of the subjects was recorded using an overhead camera (Raspberry Pi NoIR Camera Board V2 – 8MP, Raspberry Pi Foundation, UK). We replaced the water in the experimental tank with freshwater and new predator odour each time after 4 fish were tested.

Behaviour was scored using the software BORIS 7.10.2 (Friard & Gamba, 2016). We recorded the time spent in the safe zone (a third of the arena nearest to the shelter) and the predator zone (the third of the tank nearest the predator model).

Offspring boldness

Offspring were tested at 4 months old with a predator inspection test twice, three weeks apart to assess consistency in boldness following (Ramesh et al., 2023). The sample size tested was 100 fish from the control group and 94 from the treatment group. Two tanks placed side-by-side, one (40x20x30 cm) housed a live perch (Perca fluviatilis) and the other (25x20x20 cm) the focal fish and a shelter . The tanks were positioned in an illuminated wooden box to avoid external disturbance. The duration of the experiment was 10 mins. We scored the time spent inside the shelter  and outside the shelter as proxies for shyness and boldness respectively.

Offspring Shoaling (anti-predator behaviour)

To measure shoaling behaviour, 5 individuals from the same home tank were placed in a novel arena (33x18x33 cm) which was surrounded with black plastic. The test lasted 10 minutes. Every 10s a snapshot of the video was taken and the xy coordinates of each fish’s head was identified. The area of the polygon generated by coordinates was calculated using ImageJ 13.1 (Schneider et al., 2012) as a measure of shoal cohesion. We averaged the area generated by the shoal at each snapshot over the total observation time (Bibost & Brown, 2013; Bisazza & Dadda, 2005).

Offspring Laterality

We employed a rotational mirror test to score laterality in a social context (sensu Sovrano et al., 1999). Fish were placed in a circular glass tank (diameter 19 cm, water depth 4 cm) where the wall was covered with mirror foil to create a continuous mirror for the subject to swim alongside their reflection . Subjects were gently placed in the arena, and we recorded which eye the individual used to observe their mirror image every 5 seconds for 10 mins. When the fish was perpendicular to the mirror it was scored as non-lateralized. If the fish was not looking at the mirror as indicated with B in figure 5, it was not included in the lateralized formula. When the fish was looking at the mirror with one eye (<90° to the mirror) we noted if it was using the right or left eye and if the fish was > 90° it was excluded from the data set (Sovrano et al., 1999).We then calculated the total number of times each fish watched the mirror with their left, right or both eyes. The sample size tested was 100 fish from the control group and 94 from the treatment group.