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It is generally argued that distress vocalizations, a common modality for alerting conspecifics across a wide range of terrestrial vertebrates, share acoustic features that allow heterospecific communication. Yet studies suggest that the acoustic traits used to decode distress may vary between species, leading to decoding errors. Here we found through playback experiments that Nile crocodiles are attracted to infant hominid cries (bonobo, chimpanzee and human), and that the intensity of crocodile response depends critically on a set of specific acoustic features (mainly deterministic chaos, harmonicity and spectral prominences). Our results suggest that crocodiles are sensitive to the degree of distress encoded in the vocalizations of phylogenetically very distant vertebrates. A comparison of these results with those obtained with human subjects confronted with the same stimuli further indicates that crocodiles and humans use different acoustic criteria to assess the distress encoded in infant cries. Interestingly, the acoustic features driving crocodile reaction are likely to be more reliable markers of distress than those used by humans. These results highlight that the acoustic features encoding information in vertebrate sound signals are not necessarily identical across species.

Temporal contingency is a key factor in associative learning but remains weakly investigated early in life. Few data suggest simultaneous presentation is required for young to associate different stimuli, whereas adults can learn them sequentially. Here, we investigated the ability of newborn rabbits to perform sensory preconditioning and second-order conditioning using trace intervals between odor presentations. Strikingly, pups are able to associate odor stimuli with 10- and 30-sec intervals in sensory preconditioning and second-order conditioning, respectively. The effectiveness of higher-order trace conditioning in newborn rabbits reveals that very young animals can display complex learning despite their relative immaturity., (© 2022 Coureaud et al.; Published by Cold Spring Harbor Laboratory Press.)

Animals, including humans, detect odours and use this information to behave efficiently in the environment. Frequently, odours consist of complex mixtures of odorants rather than single odorants, and mixtures are often perceived as configural wholes, i.e. as odour objects (e.g. food, partners). The biological rules governing this 'configural perception' (as opposed to the elemental perception of mixtures through their components) remain weakly understood. Here, we first review examples of configural mixture processing in diverse species involving species-specific biological signals. Then, we present the original hypothesis that at least certain mixtures can be processed configurally across species. Indeed, experiments conducted in human adults, newborn rabbits and, more recently, in rodents and honeybees show that these species process some mixtures in a remarkably similar fashion. Strikingly, a mixture AB (A, ethyl isobutyrate; B, ethyl maltol) induces configural processing in humans, who perceive a mixture odour quality (pineapple) distinct from the component qualities (A, strawberry; B, caramel). The same mixture is weakly configurally processed in rabbit neonates, which perceive a particular odour for the mixture in addition to the component odours. Mice and honeybees also perceive the AB mixture configurally, as they respond differently to the mixture compared with its components. Based on these results and others, including neurophysiological approaches, we propose that certain mixtures are convergently perceived across various species of vertebrates/invertebrates, possibly as a result of a similar anatomical organization of their olfactory systems and the common necessity to simplify the environment's chemical complexity in order to display adaptive behaviours., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2022. Published by The Company of Biologists Ltd.)

How animals perceive and learn complex stimuli, such as mixtures of odorants, is a difficult problem, for which the definition of general rules across the animal kingdom remains elusive. Recent experiments conducted in human and rodent adults as well as newborn rabbits suggested that these species process particular odor mixtures in a similar, configural manner. Thus, the binary mixture of ethyl isobutyrate (EI) and ethyl maltol (EM) induces configural processing in humans, who perceive a mixture odor quality (pineapple) that is distinct from the quality of each component (strawberry and caramel). Similarly, rabbit neonates treat the mixture differently, at least in part, from its components. In the present study, we asked whether the properties of the EI.EM mixture extend to an influential invertebrate model, the honey bee Apis mellifera. We used appetitive conditioning of the proboscis extension response to evaluate how bees perceive the EI.EM mixture. In a first experiment, we measured perceptual similarity between this mixture and its components in a generalization protocol. In a second experiment, we measured the ability of bees to differentiate between the mixture and both of its components in a negative patterning protocol. In each experimental series, the performance of bees with this mixture was compared with that obtained with four other mixtures, chosen from previous work in humans, newborn rabbits and bees. Our results suggest that when having to differentiate mixture and components, bees treat the EI.EM in a robust configural manner, similarly to mammals, suggesting the existence of common perceptual rules across the animal kindgdom., Competing Interests: Competing interestsThe authors declare no competing or financial interests., (© 2020. Published by The Company of Biologists Ltd.)

In olfaction, to preserve the sensitivity of the response, the bioavailability of odor molecules is under the control of odorant-metabolizing enzymes (OMEs) expressed in the olfactory neuroepithelium. Although this enzymatic regulation has been shown to be involved in olfactory receptor activation and perceptual responses, it remains widely underestimated in vertebrates. In particular, the possible activity of OMEs in the nasal mucus, i.e. the aqueous layer that lined the nasal epithelium and forms the interface for airborne odorants to reach the olfactory sensory neurons, is poorly known. Here, we used the well-described model of the mammary pheromone (MP) and behavioral response in rabbit neonates to challenge the function of nasal mucus metabolism in an unprecedented way. First, we showed, in the olfactory epithelium, a rapid glutathione transferase activity toward the MP by ex vivo real-time mass spectrometry (PTR-MS) which supported an activity in the closest vicinity of both the odorants and olfactory receptors. Indeed and second, both the presence and activity of glutathione transferases were evidenced in the nasal mucus of neonates using proteomic and HPLC analysis respectively. Finally, we strikingly demonstrated that the deregulation of the MP metabolism by in vivo mucus washing modulates the newborn rabbit behavioral responsiveness to the MP. This is a step forward in the demonstration of the critical function of OMEs especially in the mucus, which is at the nasal front line of interaction with odorants and potentially subjected to physiopathological changes.

The European rabbit (Oryctolagus cuniculus) is a widely used model in fundamental, medical and veterinary neurosciences. Besides investigations in adults, rabbit pups are relevant to study perinatal neurodevelopment and early behaviour. To date, the rabbit is also the only species in which a pheromone - the mammary pheromone (MP) - emitted by lactating females and active on neonatal adaptation has been described. The MP is crucial since it contributes directly to nipple localisation and oral seizing in neonates, i.e. to their sucking success. It may also be one of the non-photic cues arising from the mother, which stimulates synchronisation of the circadian system during pre-visual developmental stages. Finally, the MP promotes neonatal odour associative and appetitive conditioning in a remarkably rapid and efficient way. For these different reasons, the rabbit offers a currently unique opportunity to determine pheromonal-induced brain processing supporting adaptation early in life. Therefore, it is of interest to create a reference work of the newborn rabbit pup brain, which may constitute a tool for future multi-disciplinary and multi-approach research in this model, and allow comparisons related to the neuroethological basis of social and feeding behaviour among newborns of various species. Here, in line with existing experimental studies, and based on original observations, we propose a functional anatomical description of brain sections in 4-day-old rabbits with a particular focus on seven brain regions which appear important for neonatal perception of sensory signals emitted by the mother, circadian adaptation to the short and single daily nursing of the mother in the nest, and expression of specific motor actions involved in nipple localisation and milk intake. These brain regions involve olfactory circuits, limbic-related areas important in reward, motivation, learning and memory formation, homeostatic areas engaged in food anticipation, and regions implicated in circadian rhythm and arousal, as well as in motricity., (© 2018 Anatomical Society.)

In the nasal olfactory epithelium, olfactory metabolic enzymes ensure odorant clearance from the olfactory receptor environment. This biotransformation of odorants into deactivated polar metabolites is critical to maintaining peripheral sensitivity and perception. Olfactory stimuli consist of complex mixtures of odorants, so binding interactions likely occur at the enzyme level and may impact odor processing. Here, we used the well-described model of mammary pheromone-induced sucking-related behavior in rabbit neonates. It allowed to demonstrate how the presence of different aldehydic odorants efficiently affects the olfactory metabolism of this pheromone (an aldehyde too: 2-methylbut-2-enal). Indeed, according to in vitro and ex vivo measures, this metabolic interaction enhances the pheromone availability in the epithelium. Furthermore, in vivo presentation of the mammary pheromone at subthreshold concentrations efficiently triggers behavioral responsiveness in neonates when the pheromone is in mixture with a metabolic challenger odorant. These findings reveal that the periphery of the olfactory system is the place of metabolic interaction between odorants that may lead, in the context of odor mixture processing, to pertinent signal detection and corresponding behavioral effect.

Animals routinely receive information through different sensory channels, and inputs from a modality may modulate the perception and behavioural reaction to others. In spite of their potential adaptive value, the behavioural correlates of this cross-sensory modulation have been poorly investigated. Due to their predator life, crocodilians deal with decisional conflicts emerging from concurrent stimuli. By testing young Crocodylus niloticus with sounds in the absence or presence of chemical stimuli, we show that (i) the prandial (feeding) state modulates the responsiveness of the animal to a congruent, i.e. food-related olfactory stimulus, (ii) the prandial state alters the responsiveness to an incongruent (independent of food) sound, (iii) fasted, but not sated, crocodiles display selective attention to socially relevant sounds over noise in presence of food odour. Cross-sensory modulation thus appears functional in young Nile crocodiles. It may contribute to decision making in the wild, when juveniles use it to interact acoustically when foraging., Competing Interests: We declare we have no competing interests.

In the olfactory epithelium (OE), odorant metabolizing enzymes have the dual function of volatile component detoxification and active clearance of odorants from the perireceptor environment to respectively maintain the integrity of the tissues and the sensitivity of the detection. Although emphasized by recent studies, this enzymatic mechanism is poorly documented in mammals. Thus, olfactory metabolism has been characterized mainly in vitro and for a limited number of odorants. The automated ex vivo headspace gas-chromatography method that was developed here was validated to account for odorant olfactory metabolism. This method easily permits the measurement of the fate of an odorant in the OE environment, taking into account the odorant gaseous state and the cellular structure of the tissue, under experimental conditions close to physiological conditions and with a high reproducibility. We confirmed here our previous results showing that a high olfactory metabolizing activity of the mammary pheromone may be necessary to maintain a high level of sensitivity toward this molecule, which is critical for newborn rabbit survival. More generally, the method that is presented here may permit the screening of odorants metabolism alone or in mixture or studying the impact of aging, pathology, polymorphism or inhibitors on odorant metabolism., (© The Author 2015. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Perception of odors, i.e. usually of mixtures of odorants, is elemental (the odorants' odor qualities are perceived in the mixture) or configural (the odor quality of the mixture differs from the one of each odorant). In human adults, the Red Cordial (RC) mixture is a configurally-processed, 6-odorant mixture. It evokes a red cordial odor quality while none of the elements carries that odor. Interestingly, in newborn rabbits, the same RC mixture is weak configurally perceived: the newborns behaviorally respond to all the elements after conditioning to the whole mixture, but not to the mixture after conditioning to a single element. Thus, they perceive in the RC mixture both the odor quality of the RC configuration and the quality of each element. Here, we aimed to determine whether this perception is modulated by quantitative (number of elements) and/or qualitative bits of information (nature of elements) previously learned by the animals. Newborns were conditioned to RC sub-mixtures of different complexity and composition before behavioral testing to RC. Pups generalized their sucking-related response to RC after learning at least 4 odorants. In contrast, after conditioning to sub-mixtures of another 6-odorant mixture, the elementally perceived MV mixture, pups responded to MV after learning one or two odorants. The different generalization to RC and MV mixtures after learning some of their elements is discussed according to three hypotheses: i) the configural perception of RC sub-mixtures, ii) the ratio of familiar/unfamiliar individual information elementally and configurally perceived, iii) the perception of RC becoming purely elemental. The results allow the first hypothesis to be dismissed, while further experiments are required to distinguish between the remaining two.

Extraction of relevant information from highly complex environments is a prerequisite to survival. Within odour mixtures, such information is contained in the odours of specific elements or in the mixture configuration perceived as a whole unique odour. For instance, an AB mixture of the element A (ethyl isobutyrate) and the element B (ethyl maltol) generates a configural AB percept in humans and apparently in another species, the rabbit. Here, we examined whether the memory of such a configuration is distinct from the memory of the individual odorants. Taking advantage of the newborn rabbit's ability to learn odour mixtures, we combined behavioural and pharmacological tools to specifically eliminate elemental memory of A and B after conditioning to the AB mixture and evaluate consequences on configural memory of AB. The amnesic treatment suppressed responsiveness to A and B but not to AB. Two other experiments confirmed the specific perception and particular memory of the AB mixture. These data demonstrate the existence of configurations in certain odour mixtures and their representation as unique objects: after learning, animals form a configural memory of these mixtures, which coexists with, but is relatively dissociated from, memory of their elements. This capability emerges very early in life., (© 2014 The Author(s) Published by the Royal Society. All rights reserved.)

Interacting with the mother during the daily nursing, newborn rabbits experience her body odor cues. In particular, the mammary pheromone (MP) contained in rabbit milk triggers the typical behavior which helps to localize and seize the nipples. It also promotes the very rapid appetitive learning of simple or complex stimuli (odorants or mixtures) through associative conditioning. We previously showed that 24 h after MP-induced conditioning to odorants A (ethyl isobutyrate) or B (ethyl maltol), newborn rabbits perceive the AB mixture in a weak configural way, i.e., they perceive the odor of the AB configuration in addition to the odors of the elements. Moreover, after conditioning to the mixture, elimination of the memories of A and B does not affect the memory of AB, suggesting independent elemental and configural memories of the mixture. Here, we evaluated whether configural memory persistence differs from elemental one. First, whereas 1 or 3-day-old pups conditioned to A or B maintained their responsiveness to the conditioned odorant for 4 days, those conditioned to AB did not respond to the mixture after the same retention period. Second, the pups conditioned to AB still responded to A and B 4 days after conditioning, which indicates stronger retention of the elements than of the configuration when all information are learned together. Third, we determined whether the memory of the elements competes with the memory of the configuration: after conditioning to AB, when the memories of A and B were erased using pharmacological treatment, the memory of the mixture was extended to day 5. Thus, newborn rabbits have access to both elemental and configural information in certain odor mixtures, and competition between these distinct representations of the mixture influences the persistence of their memories. Such effects certainly occur in the natural context of mother-pup interactions and may contribute to early acquisition of knowledge about the surroundings.

Smelling monomolecular odors hardly ever occurs in everyday life, and the daily functioning of the sense of smell relies primarily on the processing of complex mixtures of volatiles that are present in the environment (e.g., emanating from food or conspecifics). Such processing allows for the instantaneous recognition and categorization of smells and also for the discrimination of odors among others to extract relevant information and to adapt efficiently in different contexts. The neurophysiological mechanisms underpinning this highly efficient analysis of complex mixtures of odorants is beginning to be unraveled and support the idea that olfaction, as vision and audition, relies on odor-objects encoding. This configural processing of odor mixtures, which is empirically subject to important applications in our societies (e.g., the art of perfumers, flavorists, and wine makers), has been scientifically studied only during the last decades. This processing depends on many individual factors, among which are the developmental stage, lifestyle, physiological and mood state, and cognitive skills; this processing also presents striking similarities between species. The present review gathers the recent findings, as observed in animals, healthy subjects, and/or individuals with affective disorders, supporting the perception of complex odor stimuli as odor objects. It also discusses peripheral to central processing, and cognitive and behavioral significance. Finally, this review highlights that the study of odor mixtures is an original window allowing for the investigation of daily olfaction and emphasizes the need for knowledge about the underlying biological processes, which appear to be crucial for our representation and adaptation to the chemical environment.

This study evaluated whether olfactory preconditioning is functional in newborn rabbits and based on joined or independent memory of odorants. First, after exposure to odorants A+B, the conditioning of A led to high responsiveness to odorant B. Second, responsiveness to B persisted after amnesia of A. Third, preconditioning was also functional with two overlapping pairs of odorants (A+B and B+C) and amnesia of one odorant did not affect memory of the others. Thus, incidental pairing of odorants allows reinforcement of one odorant to implicitly reinforce the others, the bond then vanishes, and the memory of each element becomes independent.

Young and adult mammals are constantly exposed to chemically complex stimuli. The olfactory system allows for a dual processing of relevant information from the environment either as single odorants in mixtures (elemental perception) or as mixtures of odorants as a whole (configural perception). However, it seems that human adults have certain limits in elemental perception of odor mixtures, as suggested by their inability to identify each odorant in mixtures of more than 4 components. Here, we explored some of these limits by evaluating the perception of three 6-odorant mixtures in human adults and newborn rabbits. Using free-sorting tasks in humans, we investigated the configural or elemental perception of these mixtures, or of 5-component sub-mixtures, or of the 6-odorant mixtures with modified odorants' proportion. In rabbit pups, the perception of the same mixtures was evaluated by measuring the orocephalic sucking response to the mixtures or their components after conditioning to one of these stimuli. The results revealed that one mixture, previously shown to carry the specific odor of red cordial in humans, was indeed configurally processed in humans and in rabbits while the two other 6-component mixtures were not. Moreover, in both species, such configural perception was specific not only to the 6 odorants included in the mixture but also to their respective proportion. Interestingly, rabbit neonates also responded to each odorant after conditioning to the red cordial mixture, which demonstrates their ability to perceive elements in addition to configuration in this complex mixture. Taken together, the results provide new insights related to the processing of relatively complex odor mixtures in mammals and the inter-species conservation of certain perceptual mechanisms; the results also revealed some differences in the expression of these capacities between species putatively linked to developmental and ecological constraints.

Elemental and configural olfactory perception allows interaction with the environment from very early in life. To evaluate how newborn rabbits can extract and respond to information from the highly complex chemical surroundings, and how experience acts on this sensory, cognitive and behavioural capability, we ran a study in four steps including a total of eight experiments. We mainly used a binary AB mixture comprising ethyl isobutyrate (component A) and ethyl maltol (component B), previously shown as a bearer of blending properties; in rabbit pups (as in human adults), the mixture elicits a weak configural perception, i.e. the perception of a configural odour different from the odours of the components. First, a repeated exposure to one component of AB led to a more elemental perception of this mixture; conversely, a repeated exposure to AB facilitated its configural processing. Second, similar impact of experience did not appear with a non-blending AC mixture (ethyl isobutyrate-guaïacol). Third, repeated exposure to AB impacted not only the perception of AB, but also and in the same way the perception of the AC mixture sharing one component, and reciprocally. However, facilitation to perceive one mixture in one mode (configural/elemental) was not generalized to a mixture sharing no components with the experienced mixture [AB versus DE (damascenone and vanillin)]. Thus, experience contributes to the neonatal perception of odour mixtures and adds plasticity to the perceptual system. However, this impact remains dependent on the chemical composition of the mixtures.

The mammary pheromone promotes the acquisition of novel odorants (CS1) in newborn rabbits. Here, experiments pinpoint that CS1 becomes able to support neonatal learning of other odorants (CS2). We therefore evaluated whether these first- and second-order memories remained dependent after reactivation. Amnesia induced after CS2 recall selectively blocked this memory, when recall and amnesia of CS1 left the souvenir of CS2 safe; this finding partially differed from results obtained in adult mammals. Thus, in this model of neonatal appetitive odor learning, second-order memory seems to depend on first-order memory for its formation but not for its maintenance.

The processing of odour mixtures by young organisms is poorly understood. Recently, the perception of an AB mixture, known to engage configural perception in adult humans, was suggested also to be partially configural in newborn rabbits. In particular, pups did not respond to AB after they had learned A or B. However, two alternative hypotheses might be suggested to explain this result: the presence in the mixture of a novel odorant that inhibits the response to the learned stimulus, and the unevenness of the sensory and cognitive processes engaged during the conditioning and the behavioural testing. We conducted four experiments to explore these alternative hypotheses. In experiment 1, the learning of A or B ended in responses to mixtures including a novel odorant (AC or BC). Experiment 2 pointed to the absence of overshadowing. Therefore, a novelty effect cannot explain the non-response to AB after the learning of A or B. In experiment 3, pups having learned A or B in AC or BC did not respond to AB. However, they generalized odour information acquired in AB to AC or BC in experiment 4. Thus, the balancing of the perceptual tasks between the conditioning and retention test does not enhance the response to the AB mixture. To sum up, the present experiments give concrete support to the partially configural perception of specific odour mixtures by newborn rabbits.

Mammary pheromone (MP)-induced odor memory is a new model of appetitive memory functioning early in a mammal, the newborn rabbit. Some properties of this associative memory are analyzed by the use of anisomycin as an amnesic agent. Long-term memory (LTM) was impaired by anisomycin delivered immediately, but not 4 h after either acquisition or reactivation. Thus, the results suggest that this form of neonatal memory requires both consolidation and reconsolidation. By extending these notions to appetitive memory, the results reveal that consolidation and reconsolidation processes are characteristics of associative memories of positive events not only in the adult, but also in the newborn.

In flavor perception, both experience with the components of odor/taste mixtures and the cognitive strategy used to examine the interactions between the components influence the overall mixture perception. However, the effect of these factors on odor mixtures perception has never been studied. The present study aimed at evaluating whether 1) previous exposure to the odorants included in a mixture or 2) the synthetic or analytic strategy engaged during odorants mixture evaluation determines odor representation. Blending mixtures, in which subjects perceived a unique quality distinct from those of components, were chosen in order to induce a priori synthetic perception. In the first part, we checked whether the chosen mixtures presented blending properties for our subjects. In the second part, 3 groups of participants were either exposed to the odorants contributing to blending mixtures with a "pineapple" or a "red cordial" odor or nonexposed. In a following task, half of each group was assigned to a synthetic or an analytical task. The synthetic task consisted of rating how typical (i.e., representative) of the target odor name (pineapple or red cordial) were the mixtures and each of their components. The analytical task consisted of evaluating these stimuli on several scales labeled with the target odor name and odor descriptors of the components. Previous exposure to mixture components was found to decrease mixture typicality but only for the pineapple blending mixture. Likewise, subjects engaged in an analytical task rated both blending mixtures as less typical than did subjects engaged in a synthetic task. This study supports a conclusion that odor mixtures can be perceived either analytically or synthetically according to the cognitive strategy engaged.

Mammalian neonates depend on their mother's food supply and use a defined sequence of actions to find her mammary area. Their behavior is initially uncertain and demanding but rapidly becomes optimal. Efficient learning is thus operating in newborns. For instance, European rabbit (Oryctolagus cuniculus) pups localize the nipples through typical orocephalic movements. These movements are released by the mammary pheromone secreted in milk or by prenatally learned odor cues. During daily nursing, they also learn odors associated with the mother, supposedly with sucking as the main reinforcer. We here investigate the role of the mammary pheromone as an enforcer of early olfactory learning in newborn rabbits. In testing more than 950 pups, we show that the mammary pheromone promotes learning of neutral odorants paired with the pheromone in single and short trials. The pheromone-induced learning is efficient from birth and supports successive acquisition of distinct odorants. This reveals that a mammalian pheromone can function as a "cognitive organizer" that promotes early learning of relevant environmental cues.

Mammals owe part of their evolutionary success to the harmonious exchanges of information, energy and immunity between females and their offspring. This functional reciprocity is vital for the survival and normal development of infants, and for the inclusive fitness of parents. It is best seen in the intense exchanges taking place around the mother's offering of, and the infant's quest for, milk. All mammalian females have evolved behavioural and sensory methods of stimulating and guiding their inexperienced newborns to their mammae, whereas newborns have coevolved means to respond to them efficiently. Among these cues, maternal odours have repeatedly been shown to be involved, but the chemical identity and pheromonal nature of these cues have not been definitively characterized until now. Here we focus on the nature of an odour signal emitted by the female rabbit to which newborn pups respond by attraction and oral grasping, and provide a complete chemical and behavioural description of a pheromone of mammary origin in a mammalian species.

This study was aimed at characterising the impact of immediate postnatal sucking on pup survival and development. The interactive effects of postnatal success with the day 0 weight of pups, the nest-access regimen (controlled or free) or parity of females was investigated. Pups (n = 900) were categorised according to their initial ingestion of colostrum. In primiparous does: (1) pup mortality between d0-d10 was higher for unsuccessful than for successful early suckers; (2) lighter d0-weight reduced survival for unsuccessful but not for successful pups; (3) free nest-access of females annihilated the survival advantage fostered by the initial sucking success. In secondiparous does, these impacts waned. Finally, whatever the does' parity, only d0-weight influenced pup weight-gain between d0-21. Thus, pup survival seemed to depend (at least in primiparae) on their ability to suck right after birth, and to display a pattern of energy saving without being disturbed by the females' nest entries.