In the past century, insects have become models for studying associative learning and memory formation. Learning paradigms have been developed for several insect species and modalities, but they comprise only two main categories: classical and operant conditioning. While the same cues can be learnt in both paradigms, the process by which learning occurs differs on the extent to which animals need to interact with their environment to form a memory. Classical conditioned individuals obtain information about a cue and a reward/punishment passively while operant conditioned animals need perform an action to be rewarded/avoid punishment. Thus, to truly understand how memories are formed and stored in the brain, it becomes essential to identify to which extent self-action within the environment influences learning.

Considering this, I have investigated memory formation in wood ants, Formica rufa, through classical conditioning and found that they are able to passively learn the association between a visual cue and a sugar reward. I have explored this paradigm to investigate particularities of this type of learning and showed that these memories can be lateralised, with short- and medium-term visual memories being formed after training with a reinforcement of the right antenna and long-term memories with a reinforcement on the left antenna.

Additionally, I have developed a paradigm in which wood ants can walk on an air supported ball in open- or closed-loop with a virtual world. To demonstrate this paradigm is functional, I have investigated the use of self-generated optic flow on the integration of distance, speed and time of walking ants. I found that ants display repeatable walking behaviour that does not require but can be changed by variations of self-induced or external optic flow. This paradigm allows for a fully controlled comparison and analysis of active and passive interactions with a virtual environment on a tethered animal from which neural circuits could be accessible.

Ants are excellent navigators using multimodal information for navigation. To accurately localise the nest at the end of a foraging journey, visual cues, wind direction and also olfactory cues need to be learnt. Learning walks are performed at the start of an ant’s foraging career or when the appearance of the nest surrounding has changed. We investigated here whether the structure of such learning walks in the desert ant Cataglyphis fortis takes into account wind direction in conjunction with the learning of new visual information. Ants learnt to travel back and forth between their nest and a feeder, and we then introduced a black cylinder near their nest to induce learning walks in regular foragers. By doing this across days with different wind directions, we were able to probe how ants balance different sensory modalities. We found that (1) the ants’ outwards headings are influenced by the wind direction with their routes deflected such that they will arrive downwind of their target, (2) a novel object along the route induces learning walks in experienced ants and (3) the structure of learning walks is shaped by the wind direction rather than the position of the visual cue.

Several species of insects have become model systems for studying learning and memory formation. Although many studies focus on freely moving animals, studies implementing classical conditioning paradigms with harnessed insects have been important for investigating the exact cues that individuals learn and the neural mechanisms underlying learning and memory formation. Here we present a protocol for evoking visual associative learning in wood ants through classical conditioning. In this paradigm, ants are harnessed and presented with a visual cue (a blue cardboard), the conditional stimulus (CS), paired with an appetitive sugar reward, the unconditional stimulus (US). Ants perform a Maxilla-Labium Extension Reflex (MaLER), the unconditional response (UR), which can be used as a readout for learning. Training consists of 10 trials, separated by a 5-minute intertrial interval (ITI). Ants are also tested for memory retention 10 minutes or 1 hour after training. This protocol has the potential to allow researchers to analyze, in a precise and controlled manner, the details of visual memory formation and the neural basis of learning and memory formation in wood ants.

Wood ants are a model system for studying visual learning and navigation. They can forage for food and navigate to their nests effectively by forming memories of visual features in their surrounding environment. Previous studies of freely behaving ants have revealed many of the behavioural strategies and environmental features necessary for successful navigation. However, little is known about the exact visual properties of the environment that animals learn or the neural mechanisms that allow them to achieve this. As a first step towards addressing this, we developed a classical conditioning paradigm for visual learning in harnessed wood ants that allows us to control precisely the learned visual cues. In this paradigm, ants are fixed and presented with a visual cue paired with an appetitive sugar reward. Using this paradigm, we found that visual cues learnt by wood ants through Pavlovian conditioning are retained for at least one hour. Furthermore, we found that memory retention is dependent upon the ants’ performance during training. Our study provides the first evidence that wood ants can form visual associative memories when restrained. This classical conditioning paradigm has the potential to permit detailed analysis of the dynamics of memory formation and retention, and the neural basis of learning in wood ants.

Ant foragers make use of multiple navigational cues to navigate through the world and the combination of innate navigational strategies and the learning of environmental information is the secret of their navigational success. We present here detailed information about the paths of Cataglyphis fortis desert ants navigating by an innate strategy, namely path integration. Firstly, we observe that the ants’ walking speed decreases significantly along their homing paths, such that they slow down just before reaching the goal, and maintain a slower speed during subsequent search paths. Interestingly, this drop in walking speed is independent of absolute home-vector length and depends on the proportion of the home vector that was completed. Secondly, we find that ants are influenced more strongly by novel or altered visual cues the further along their homing path they are. These results suggest that path integration modulates speed along the homing path in a way that might help ants search for, utilise or learn environmental information at important locations. Ants walk more slowly and sinuously when encountering novel or altered visual cues and occasionally stop and scan the world, this might indicate the re-learning of visual information.