Multiscale study of social learning and memory in Drosophila melanogaster: from genes to animal culture – MoleCulture

Behavioral Protocol

A female Drosophila can use public information to select a mate from a group. For instance, an "observer" female will preferentially mate with a male dusted with an artificial color (e.g., pink) that she previously saw successfully mating with a "demonstrator" female. This preference holds true over a male dusted with another color (e.g., green) that she saw being rejected by another demonstrator female. Thus, Drosophila females can utilize socially learned information. These findings could have significant evolutionary implications, as socially acquired sexual preferences might lead to reproductive isolation, potentially paving the way for speciation.

This research project explores the mechanisms and evolutionary consequences of mate-copying (MC) in Drosophila.

Neurobiological and Molecular Foundations

From a neurobiological and molecular perspective, the study delves into observational social learning. The rutabaga (rut) protein, involved in cAMP synthesis, has been identified as crucial for this process, acting as a "coincidence detector" between stimuli. The project aims to identify the key brain structures for MC, including the ellipsoid body (EB), the fan-shaped body (FSB), and the mushroom bodies (MBs), all known for their roles in visual learning and memory. The role of dopamine and serotonin as unconditioned stimulus (US) pathways will also be examined using pharmacological and neurogenetic approaches, incorporating ecologically relevant stimuli (actual mating) and free-moving flies.

Social Long-Term Memory (SoLTM)

Regarding Social Long-Term Memory (SoLTM), the project seeks to demonstrate its existence in Drosophila through repeated demonstrations. It's expected to show that this memory depends on protein synthesis (a characteristic of long-term memory) and to identify the involved neural centers by using genetic rescue of the rutabaga mutation.

Evolutionary Consequences of Mate-Copying

For the evolutionary consequences of MC, the objective is to prove that D. melanogaster possesses the cognitive abilities for the cultural transmission of sexual preferences across multiple generations, leading to durable local traditions. The project will explore conformity in Drosophila females by observing whether they conform to the majority of observed mate choices, using a hexagonal setup. Finally, long transmission chains will be conducted to empirically test the stability of this cultural transmission.

Methodological Development

In terms of methodological development, the complexity of repeated demonstrations can be overcome by creating a new methodology based on photos of couples. This will allow for the standardization and acceleration of experiments, testing whether visual cues alone suffice for MC, and refining the study of influential cues.

This project revealed that Drosophila possesses the cognitive abilities necessary to culturally transmit a preference for a trait (Danchin et al., 2018). This finding challenges the view of culture as solely human prerogative and contributes to a broader debate on cultural transmission in vertebrates.

To prove cultural transmission, five criteria were verified in Drosophila:

1. Social learning through observation of conspecifics.

2. Learning from older to younger individuals.

3. Social Long-term memory.

4. The behavior concerns individual characteristics (here, male color).

5. The behavior is conformist, with individuals learning the most frequent preference.

This research highlighted the key role of conformism and revolutionized our perception of cultural transmission capacity in invertebrates, providing a mechanistic definition of culture in a broad sense. This opens promising avenues for the study of culture's evolution.

Decoding the Behavioral Process and Stimuli of MC

The behavioral process of mate-copying (MC) has been finely analyzed. It was established that observer females base their decisions on observing the male's acceptance by the demonstrator female, not on the rejection of a spurned male (Noebel et al., 2022). This distinction is crucial for understanding the positive nature of the learning signal.

Regarding the necessary stimuli, innovative experiments showed that demonstration does not require in vivo interactions. Even highly simplified 2D images of mating pairs are sufficient to trigger MC in naive females (Noebel et al., 2022).

The order of social information was also examined. When faced with contradictory and sequential demonstrations, Drosophila females exhibit a primacy bias, meaning they prefer the male characterized by the trait observed first (Santiago et al., 2024).

A mechanistic model of this social learning has been proposed, where the observation of mating acts as an unconditioned stimulus associated with the male's color as a conditioned stimulus. Observer females thus form a simple associative memory. Research on the rutabaga (rut) mutant, deficient in adenylyl cyclase (Rut+) essential for cAMP synthesis and involved in various associative learnings, showed that Rut+ in Kenyon cells (KC) gamma (gamma-KC) of the mushroom bodies is necessary for MC (Noebel et al., 2023). Furthermore, additional alpha-KC and beta-KC Kenyon cells are recruited (Noebel et al., 2025) to establish long-term memory, demonstrating a specificity of brain structures in different memory phases, like non-social memory.

Building upon a paradigm developed for observational social learning – mate copying (MC) in Drosophila – our new project aims to identify and explore the neurobiological mechanisms of social learning and memory in a brain containing a million times fewer neurons than a human's. Our previous work has demonstrated that Drosophila possesses the cognitive abilities necessary to transmit this social learning at the population level. However, it remains to be determined if this transmission can persist in a natural context.

Leveraging the genetic tools available in Drosophila, we plan to:

1. Identify and functionally characterize the specific neural networks involved in MC with cellular resolution: We will develop a novel in vivo imaging setup that allows us to examine the physiological properties of newly identified neurons in response to the visual stimuli involved in MC. This will provide functional data on MC, from which we can model this observational social learning.

2. Determine the effects of MC and test the involvement of identified MC neurons at the population level: We will conduct a semi-natural experiment to assess the impact of MC and the role of specific neurons in this process.

By doing so, we will clarify the anatomical and functional foundations of this social learning and evaluate its importance in cultural transmission.

Neuroscientists are unanimous in recognizing that experiences influence brain, which in turn influences experiences. For instance, individuals can learn from their own experience. However, individuals can also learn from the observation of the experience of other individuals, a process called “observational learning”, and which constitutes a form of social learning. This process constitutes an alternative to "trial-and-error" strategies, which can be very costly in some contexts such as tasting toxic foods, confronting a predator or even more so in reproductive decisions given the small numbers of reproductive events in a life-time.
From an evolutionary point of view, observational learning, or social learning in general, has the potential to foster the emergence of cultural transmission and evolution2-6 that in return can modify the selection pressures acting on individuals (through the effect of social constraints). While social learning is documented in a vast array of species, beyond Humans, cultural transmission has been documented in only a small number of mammals and birds. However, we recently proposed on the basis of a detailed experimental and theoretical study of social learning in the context of mate-choice, that social learning in drosophila may foster the emergence of real cultural traditions in mating preference that may vary among populations. In view of the importance of mate-choice in terms of sexual selection, we now propose to merge the study of the neurobiology of cognition and that of the evolutionary origin and consequences of that type of social learning to better understand its evolutionary and mechanistic characteristics.
Usually, mechanisms of social learning are studied at the sole level of individual’s behavior so that its neurobiological underpinning and the selective pressure that prevailed during their evolution remain poorly investigated. This is because there is currently no recognized simple and tractable biological model that integrates and processes social stimuli to enable the study of how the brain learns and memorizes social information. In addition, research on social learning mostly focuses either on proximate or ultimate causes, but only rarely integrate these two levels of analysis.
This project is interdisciplinary in its essence. Its goal is to start exploring the neurobiological mechanisms of social learning while accounting for their consequences at the individual, population and evolutionary scale. As a corollary, this project will establish Drosophila melanogaster as a new mechanistic model of social learning.