Brain responses to basic tastes

The brain integrates a huge amount of information coming from the environment and the inside of the organism, through different communication pathways. Exposure to food and their post-ingestive consequences generates both exteroceptive (e.g. smell, taste, vision) and interoceptive (i.e. gut perceptions) signals that the brain must integrate and associate to elaborate a multisensory mental representation of food items. This mental representation includes physical and hedonic dimensions depending on the individual experience shaping individual expectations. One of our first objectives was consequently to map the brain responses to basic tastants, such as sweet or bitter compounds, which are known to elicit strong innate appetitive or repulsive reactions (Steiner et al., Reference Steiner, Glaser, Hawilo and Berridge2001). The role of gut nutrient sensing in modulating appetite and conditioning food preferences (notably via sugar gut infusions) has also been extensively described (Sclafani and Ackroff, Reference Sclafani and Ackroff2012). For sweet/sugar perception, our first question was to elucidate whether the detection site might condition the brain responses elicited. Glucose portal infusion (Figure 1e) and duodenal infusion (Figure 1d) both reduced feed intake and ingestion speed in a similar way, but induced different systemic signals and different brain responses (Table 1; Boubaker et al., Reference Boubaker, Val-Laillet, Guerin and Malbert2012), which demonstrates that gut nutrient and/or taste perception can shape brain responses related to hedonic and homeostatic regulation of food intake in the pig. The next step was to compare exteroceptive (i.e. gustatory) and interoceptive (i.e. duodenal) perceptions of sweet taste/nutrient, and more precisely to compare the impact of combined v. dissociated oral and intestinal sucrose detection, on the brain reward processes. Oral and/or duodenal sucrose sensing (Figure 1c and d) induced differential cerebral blood flow changes in brain regions, known to be involved in memory, reward processes and hedonic evaluation of sensory stimuli (Table 1; Clouard et al., Reference Clouard, Meunier-Salaun, Meurice, Malbert and Val-Laillet2014). Such information is valuable to disentangle the respective roles of oral and visceral perception of sweet/sugar in modulating food intake and motivation, and for translational research using the pig model – for example, to assess the impact of additives or artificial sweeteners, design new food products or improve their nutritional quality with a benefits–risks approach in terms of palatability, health, gut–brain axis and behaviour.

More recently, using fMRI, we aimed to compare oral sweet and bitter gustatory stimulation (Table 1; Coquery et al., Reference Coquery, Meurice, Janvier, Bobillier, Quellec, Saint Jalmes, Fu, Roura and Val-Laillet2018). The advantage of fMRI for comparing brain responses to different food stimuli is that multiple stimulations can be performed within the same imaging session, contrary to nuclear imaging (positron emission tomography (PET) and single-photon emission computed tomography (SPECT). The fMRI modality has already been implemented and used in the pig, notably to assess brain activity after deep-brain stimulation (Knight et al., Reference Knight, Min, Hwang, Marsh, Paek, Kim, Felmlee, Abulseoud, Bennet, Frye and Lee2013; Gibson et al., Reference Gibson, Ross, Han, Van Gompel, Min and Lee2016), but to our knowledge, the work performed by Coquery et al. (Reference Coquery, Meurice, Janvier, Bobillier, Quellec, Saint Jalmes, Fu, Roura and Val-Laillet2018) is the first to describe BOLD brain responses to visual (Figure 1a) and food stimuli in the pig. We showed that quinine and sucrose stimulations (Figure 1c) induced different brain activation patterns in gustatory, hedonic and cognitive areas (Table 1; Coquery et al., Reference Coquery, Meurice, Janvier, Bobillier, Quellec, Saint Jalmes, Fu, Roura and Val-Laillet2018). This might contribute to the understanding of the effects of bitter compounds on gut physiology and behaviour, for which significant effects were observed on gastric emptying, intestinal motility, gastrointestinal hormones and food intake (Fu et al., Reference Fu, Manchadi, De Jager, Val-Laillet, Guerin and Roura2016; Roura et al., Reference Roura, Koopmans, Lalles, Le Huerou-Luron, de Jager, Schuurman and Val-Laillet2016).

Brain responses to preferred or aversive food flavours

Previous review papers explained why and how pig models could help disentangle the complex processes underlying food perception and preferences, and how such studies might benefit biomedical research (Clouard et al., Reference Clouard, Meunier-Salaun and Val-Laillet2012c; Roura et al., Reference Roura, Koopmans, Lalles, Le Huerou-Luron, de Jager, Schuurman and Val-Laillet2016). From 2011, we published the first studies using functional imaging to describe brain responses to conditioned food preferences and aversions in pigs, using 99mTc-HMPAO SPECT (Gaultier et al., Reference Gaultier, Meunier-Salaun, Malbert and Val-Laillet2011; Boubaker et al., Reference Boubaker, Val-Laillet, Guerin and Malbert2012; Clouard et al., Reference Clouard, Meunier-Salaun, Meurice, Malbert and Val-Laillet2014) and 18F-FDG PET (Clouard et al., Reference Clouard, Jouhanneau, Meunier-Salaun, Malbert and Val-Laillet2012a) to map cerebral blood flow and brain glucose metabolism, respectively. We validated the use of brain imaging in anaesthetized pigs and confirmed that the brain is still able to integrate and react to food sensory stimuli (e.g. flavours) according to individual positive or negative experience and associated internal states (Table 1; Gaultier et al., Reference Gaultier, Meunier-Salaun, Malbert and Val-Laillet2011 and Reference Clouard, Jouhanneau, Meunier-Salaun, Malbert and Val-Laillet2012c). Different brain responses were identified in a neural network including frontostriatal and limbic areas, which are known to be involved in the regulation of food intake and pleasure in the human and rodents (Figure 1b and c). Associative conditioning between post-ingestive consequences and the meal’s flavours shaped further hedonic valuation and food choices, which highlighted the gut–brain mechanisms influencing eating behaviour in pigs. This model might be useful to investigate the onset of food aversion consequent to invasive medical treatments, such as radiotherapy or chemotherapy, or exacerbated food preferences mediated by hedonic reinforcers such as added sugar (Clouard et al., Reference Clouard, Meunier-Salaun and Val-Laillet2012c).

Impact of the metabolic or emotional status on brain responses to food signals

Brain responses to food stimuli can be modulated by previous associative conditioning but also by the individual’s metabolic or psychological state. In rodents, it has been shown that taste/olfactory perception capacity decreased with visceral fat increase (Fernandez-Garcia et al., Reference Fernandez-Garcia, Alcaide, Santiago-Fernandez, Roca-Rodriguez, Aguera, Banos, Botella, de la Torre, Fernandez-Real, Fruhbeck, Gomez-Ambrosi, Jimenez-Murcia, Menchon, Casanueva, Fernandez-Aranda, Tinahones and Garrido-Sanchez2017), and Thanos et al. (Reference Thanos, Michaelides, Gispert, Pascau, Soto-Montenegro, Desco, Wang, Wang and Volkow2008) documented significant changes in brain glucose metabolism in obese compared to lean rats exposed to appetitive food scent. Even though this question was not investigated in the pig model, we explored another relationship between the internal state and brain responses to odours. Emotional and physiological responses to stress are known to impact olfactory perception (Hoenen et al., Reference Hoenen, Wolf and Pause2017). We recently aimed to describe the consequences of a psychosocial stress in pigs on the gut–brain axis, behaviour and brain responses to olfactory food stimuli. We exposed stressed and control animals to low/high concentrations of novel food odours (Figure 1b) and showed activations in the prepyriform, entorhinal, orbitofrontal, and anterior cingulate cortices, dorsal striatum and hippocampus. We also showed that high-concentration olfactory stimuli induced a higher brain activity (Table 1; Coquery et al., in press). More interestingly, pigs with a psychosocial stress had lower brain responses in the right hippocampus compared to unstressed control pigs, as well as lower magnetic resonance spectroscopy-estimated neuronal density in the hippocampus, which suggested an impact of the internal state on cognitive and limbic processes at the brain level (Table 1; Menneson et al., Reference Menneson, Ménicot, Ferret-Bernard, Guérin, Romé, Le Normand, Randuineau, Gambarota, Noirot, Etienne, Coquery and Val-Laillet2019).

Brain anomalies in diet-induced obese minipigs

The use of preclinical models is very important in deciphering the determinants of obesity, eating disorders and metabolic diseases, but also to identify novel predictive biomarkers and innovative therapeutic strategies focused on the gut–brain axis (Val-Laillet et al., Reference Val-Laillet, Aarts, Weber, Ferrari, Quaresima, Stoeckel, Alonso-Alonso, Audette, Malbert and Stice2015; Obrenovich et al., Reference Obrenovich, Fluckiger, Sykes and Donskey2016). The limited size of minipigs enables longitudinal studies at the adult age using the same imaging machines as in humans. The first studies using the minipig as a model of obesity date back to the 1980s (e.g. Phillips et al., Reference Phillips, Westmoreland, Panepinto and Case1982), but most of the publications appeared after 2000 (Johansen et al., Reference Johansen, Hansen, Richelsen and Malmlof2001; Larsen et al., Reference Larsen, Juhl, Porksen, Gotfredsen, Carr, Ribel, Wilken and Rolin2005; Christoffersen et al., Reference Christoffersen, Grand, Golozoubova, Svendsen and Raun2007; Lee et al., Reference Lee, Alloosh, Saxena, Van Alstine, Watkins, Klaunig, Sturek and Chalasani2009; Madsen et al., Reference Madsen, Birck, Fredholm and Cirera2010; Val-Laillet et al., Reference Val-Laillet, Guerin and Malbert2010c). Brain activity was described for the first time in obese minipigs in the Göttingen breed (Val-Laillet et al., Reference Val-Laillet, Layec, Guerin, Meurice and Malbert2011a), and then in the Yucatan (Ochoa et al., Reference Ochoa, Malbert, Meurice and Val-Laillet2016a). Brain functional characteristics or anomalies associated with obesity and weight gain have been well described in the humans, encompassing initial high sensibility to highly palatable food stimuli in high-risk individuals (i.e. reward surfeit), but also decreased brain responses in the prefrontal cortex and striatum in obese patients (i.e. reward deficit), as well as a blunted dopaminergic system (Burger and Stice, Reference Burger and Stice2011; Blum et al., Reference Blum, Thanos, Oscar-Berman, Febo, Baron, Badgaiyan, Gardner, Demetrovics, Fahlke, Haberstick, Dushaj and Gold2015).

Even though evidence suggests that chronic exposure to energy-dense palatable foods provokes a switch in the hedonic and motivational brain processes (Burger and Stice, Reference Burger and Stice2011), and that successful BW loss can reverse some obesity-related brain anomalies (Le et al., Reference Le, Pannacciulli, Chen, Salbe, Del Parigi, Hill, Wing, Reiman and Krakoff2007), there was no clear demonstration showing that these brain anomalies are induced by (or concomitant to) weight gain in obese people and not pre-existent to it. Our brain imaging study in the minipig (Table 1; Val-Laillet et al., Reference Val-Laillet, Layec, Guerin, Meurice and Malbert2011a), in association with physiological data (Val-Laillet et al., Reference Val-Laillet, Guerin and Malbert2010b and Reference Val-Laillet, Blat, Louveau and Malbert2010c), was the first to demonstrate that diet-induced obesity could induce brain anomalies in comparison to lean subjects, including decreased basal metabolism in the prefrontal cortex and reward circuit (Figure 1; diet-induced obesity). These results suggest that some characteristics of the reward-deficit phenotype observed in obese humans were probably an acquired feature of obesity, induced by regular excess food intake.

Impact of specific nutrients on brain anomalies in obese minipigs

Following the 2011 demonstration in obese minipigs, two questions arose: Are weight gain and excess calorie intake only liable for these brain anomalies described in obese subjects? And does food quality, independently from the quantity consumed, have a role in shaping brain activity in obese subjects? In two additional independent studies in obese minipigs, we decided to compare different isocaloric obesogenic diets differing in terms of lipid or carbohydrate quality, respectively. The first study compared a 5-week exposure to three obesogenic diets only differing in the nature of lipids used for their composition (i.e. fish oil/n-3, sunflower oil/n-6, or lard/saturated fatty acids). n-3 fatty acids inhibit the production of proinflammatory cytokines (Parker et al., Reference Parker, Gibson, Brotchie, Heruc, Rees and Hadzi-Pavlovic2006), and n-3 supplementation (docosahexaenoic acid (DHA)) can increase the activity of the prefrontal cortex (McNamara et al., Reference McNamara, Able, Jandacek, Rider, Tso, Eliassen, Alfieri, Weber, Jarvis, DelBello, Strakowski and Adler2010), which is one of the brain areas found to be deactivated in obese humans and minipigs (Le et al., Reference Le, Pannacciulli, Chen, Del Parigi, Salbe, Reiman and Krakoff2006; Volkow et al., Reference Volkow, Wang, Telang, Fowler, Goldstein, Alia-Klein, Logan, Wong, Thanos, Ma and Pradhan2009; Val-Laillet et al., Reference Val-Laillet, Layec, Guerin, Meurice and Malbert2011a). We hypothesized that n-3 might prevent the onset of central inflammation and protect brain areas involved in food intake control. We demonstrated significant changes in terms of blood–brain barrier (BBB) permeability and basal brain glucose metabolism according to the diets (Table 1; Val-Laillet et al., Reference Val-Laillet, Meurice, Lalles and Malbert2011b and Reference Val-Laillet, Meurice, Lalles and Malbert2013). Contrary to what was expected, we showed that a diet (overly) enriched with n-3 protected neither the BBB nor the basal metabolism of brain areas that are dysfunctioning in the context of obesity (Figure 1; diet-induced obesity). Consistent results were obtained on intestinal permeability (Lalles et al., Reference Lalles, Perrier, Val-Laillet and Malbert2011 and Reference Lalles, Perrier, Val-Laillet and Malbert2013), suggesting global effects of the diets on the gut–brain axis. Excessive dietary n-3 might increase the fluidity of intestinal and BBB cell membranes, thereby increasing their permeability and facilitating the invasion of proinflammatory compounds towards blood and the brain. Even though this hypothesis still needs validation, these results highlight the need for precise recommendations in terms of dietary n-3, because excessive supplementation might be as deleterious as a deficiency for the gut and brain functions. The European Food Safety Authority (EFSA) stated that dietary recommendations for eicosapentaenoic acid and DHA for European adults are between 250 and 500 mg/day, and that supplemental intakes at doses up to 5 g/day do not raise safety concerns for adults in terms of cardiovascular disease risk, glucose homeostasis or immune function (European Food Safety Authority, 2012). n-3 supplementation in our minipig study was 1.9 g/day in average, which corresponded to 3.2 g of n-3 per day in a normal-weight (63 kg) adult woman. This was above the recommended dose and below the tolerable upper intake level of n-3 according to the EFSA, but no other study in our knowledge ever investigated brain metabolism outcomes at these doses.

The second study relating nutrient quality and brain activity compared a long-term exposure to three obesogenic isocaloric diets, only differing in the nature of their carbohydrates (i.e. starch as control, compared to glucose and fructose) (Ochoa et al., Reference Ochoa, Malbert, Lalles, Bobillier and Val-Laillet2014, Reference Ochoa, Malbert, Meurice and Val-Laillet2016a and Reference Ochoa, Val-Laillet, Lalles, Meurice and Malbert2016b). Fructose, which is overly used in industrial food products, has been incriminated for its role in the onset of metabolic syndrome and its impact on gut–brain metabolic signals in rodents (Dornas et al., Reference Dornas, de Lima, Pedrosa and Silva2015; Ochoa et al., Reference Ochoa, Lalles, Malbert and Val-Laillet2015; Legeza et al., Reference Legeza, Marcolongo, Gamberucci, Varga, Banhegyi, Benedetti and Odermatt2017), but counterarguments have been raised (Rizkalla, Reference Rizkalla2010), and the results obtained in the minipig model feed the controversy. Obesogenic diets altered fat accretion and metabolic profiles independently of dietary carbohydrates, but only pigs chronically fed the fructose diet developed a strong preference for it (Ochoa et al., Reference Ochoa, Malbert, Lalles, Bobillier and Val-Laillet2014 and Reference Ochoa, Val-Laillet, Lalles, Meurice and Malbert2016b). After 7 months of diet, all groups presented a similar weight gain and insulin sensitivity index; and compared to starch, chronic exposure to fructose and glucose induced bilateral activations in several reward-related brain regions, including prefrontal and striatal areas (Figure 1; diet-induced obesity), with no significant differences between fructose and glucose groups (Table 1; Ochoa et al., Reference Ochoa, Malbert, Meurice and Val-Laillet2016a). Consistent with the dynamic vulnerability model (Stice et al., Reference Stice, Figlewicz, Gosnell, Levine and Pratt2013), it is plausible that our minipigs did not develop a blunted reward response yet, since they were food-restricted for the entire experimental diet exposure and became only moderately obese. Brain metabolism differences observed here would rather be in favour of the reward surfeit theory and hypersensitization of the reward circuit, which might precede the reward deficit status, which was previously validated in a massively obese minipig model (Val-Laillet et al., Reference Val-Laillet, Layec, Guerin, Meurice and Malbert2011a).

Dynamic relationship between brain activity, body weight and diet changes

Among the hot questions on the relationship between excessive consumption of palatable foods and the emergence of brain dysfunctions is the onset dynamic of these anomalies and its relationship with BW status. It is still unknown whether regular consumption of highly palatable foods without weight gain, or different strategies to lose weight, can also influence neural networks and related behavioural functions. Gautier Y., et al. (unpublished data) aimed at following the neurocognitive processes in a cohort of adult minipigs, from a normal-weight status when exposed daily to a high-fat-sugar (HFS) diet rationed not to exceed the metabolic needs, to the onset of obesity when ad libitum food intake was permitted, and finally to a third stage when animals were subjected either to a restrictive diet or bariatric surgery to make them lose weight. This is the first study describing longitudinally the impact of diet switch and/or BW variations on metabolic and neurocognitive processes, using two in vivo brain imaging modalities (PET and fMRI) in the same minipigs. Our preliminary and yet unpublished data indicated that brain responses to sucrose were dependent on both the metabolic/nutritional status and BW, and that significant variations of brain activity were detected in the dorsal striatum, prefrontal cortex and insular cortex. Complete results are still pending, but we hope to identify brain imaging markers that might predict weight gain or weight loss after nutritional interventions, in combination with metabolic markers (e.g. hormonal, transcriptomics, metabolomics and metagenomics) to hypothesize the potential underlying microbiota–gut–brain mechanisms influencing eating behaviour and disorders.

Developmental origins of health and diseases

The concept of developmental origins of health and diseases, initially described by David Barker (Barker, Reference Barker2001), postulates that early nutritional environment can modify the progeny’s metabolism, physiology and behaviour, and influence its susceptibility to declare further pathologies. In humans, it is admitted that maternal obesity increases the prevalence of obesity and metabolic syndrome in the progeny (McGuire et al., Reference McGuire, Dyson and Renfrew2010). The effects of high-calorie and high-lipid diets on glucose metabolism, energy balance, cardiovascular function and adiposity of the progeny have been demonstrated in rodents, pigs and non-human primates (Albuquerque et al., Reference Albuquerque, Sardinha, Telles, Watanabe, Nascimento, Tavares do Carmo and Ribeiro2006; Williams et al., Reference Williams, Seki, Vuguin and Charron2014; Wang et al., Reference Wang, Zhou, Liu, Li, Zhao, Deng, Cao, Che, Fang, Xu, Lin, Feng, Li and Wu2016), similarly to the susceptibility to declare cognitive, emotional, psychiatric and behavioural disorders (Bale et al., Reference Bale, Baram, Brown, Goldstein, Insel, McCarthy, Nemeroff, Reyes, Simerly, Susser and Nestler2010). Maternal obesity and/or a plethoric HFD during gestation and lactation also induce brain anomalies, with the possible onset of neuroinflammation, decreased spatial memory, neurogenesis and brain plasticity alterations (e.g. in the hippocampus and reward circuit). The microbiota represents a major actor of the gut–brain communication (Bienenstock et al., Reference Bienenstock, Kunze and Forsythe2015; Carabotti et al., Reference Carabotti, Scirocco, Maselli and Severi2015). In mice, maternal consumption of a HFD during gestation and lactation impacts intestinal immunity and microbiota of the progeny (Myles et al., Reference Myles, Fontecilla, Janelsins, Vithayathil, Segre and Datta2013), and a direct relationship exists between excessive food intake before weaning, microbiota composition and body adiposity accretion (Sefcikova et al., Reference Sefcikova, Bujnakova, Racek, Kmet and Mozes2011).

Early nutritional programming of neurocognitive functions in the minipig

The pig provides a great opportunity to investigate the consequences of nutritional programming for the gastrointestinal tract (Guilloteau et al., Reference Guilloteau, Zabielski, Hammon and Metges2010) and neurodevelopment (Mudd and Dilger, Reference Mudd and Dilger2017). Recent studies focused on the impact of perinatal nutrition on brain functions in pigs, showing, for instance, that perinatal choline deficiency and maternal dietary choline both influence brain development in young pigs (Mudd et al., Reference Mudd, Getty, Sutton and Dilger2016 and Reference Mudd, Getty and Dilger2018). Most of the translational research on perinatal programming of obesity and associated diseases has been performed in rodent models, while the use of large animals such as pigs is scarce (Gonzalez-Bulnes and Chavatte-Palmer, Reference Gonzalez-Bulnes and Chavatte-Palmer2017).

To fill this gap, we performed an experiment in the Yucatan minipig model and constituted two groups of sows fed either a standard or an HFS diet during the last third of gestation and all the lactation period. Maternal daily food rations were calculated to prevent obesity and BW differences between control and HFS groups (contrary to most studies in rodents). All the piglets from both groups were fed a standard diet after weaning. The HFS maternal diet increased plasma levels of triglycerides and free fatty acids in weaned piglets, but more interestingly, it significantly decreased the faecal amount of short-chain fatty acids (SCFA) 100 days after birth, even though all animals were fed a standard diet after weaning (Val-Laillet et al., Reference Val-Laillet, Besson, Guerin, Coquery, Randuineau, Kanzari, Quesnel, Bonhomme, Bolhuis, Kemp, Blat, Le Huerou-Luron and Clouard2017). This indicates a decreased fermentation activity of the intestinal microbiota, already detected in HFS sows. An alternative hypothesis would be an increased intestinal absorption/use of SCFAs, and further investigations in terms of intestinal metabolism or metagenomic sequencing are required to disentangle this question. In the holeboard test (spatial learning task), where the animals have to learn and remember the location of food rewards hidden in a complex environment, Yucatan piglets born from HFS sows had increased reference and working memory scores compared to control (Val-Laillet et al., Reference Val-Laillet, Besson, Guerin, Coquery, Randuineau, Kanzari, Quesnel, Bonhomme, Bolhuis, Kemp, Blat, Le Huerou-Luron and Clouard2017), which corroborated previous data in conventional piglets (Clouard et al., Reference Clouard, Kemp, Val-Laillet, Gerrits, Bartels and Bolhuis2016). This might be explained by increased cognitive abilities and/or an increased motivation to perform/complete the cognitive task since it was reinforced by highly palatable food rewards (chocolate/sugar-coated peanuts). Any change in the motivation to obtain these food rewards can influence the animals’ memory scores. This is where the exploration of brain functions and activity is fundamental to interpret such ambivalent results.

After post-weaning behavioural tests, some piglets were euthanized to explore their brain plasticity and neuroinflammation in the hippocampus and prefrontal cortex. The hippocampus is involved in spatial learning, while the prefrontal cortex has a major role in hedonic valuation of food and goal-oriented behaviour. Interestingly, the piglets born from HFS mothers had a smaller hippocampal granular cell layer and a decreased neurogenesis (Val-Laillet et al., Reference Val-Laillet, Besson, Guerin, Coquery, Randuineau, Kanzari, Quesnel, Bonhomme, Bolhuis, Kemp, Blat, Le Huerou-Luron and Clouard2017). An increased microglial cell density was also observed in the prefrontal cortex, and the activation of microglia was increased in the hippocampus and prefrontal cortex (Val-Laillet D., Kanzari A. et al., unpublished data). These observations rather praise in favour of decreased cognitive abilities and a possible alteration of the prefrontal processes involved in food valuation and motivation, due to neuroinflammation processes.

Long-term impact of early nutritional programming

Some piglets were kept alive until the adult age to perform new behavioural tests, metabolic measures and brain exploration with in vivo imaging techniques. The differences observed in the young age for the holeboard test were not found again, but during the maze test (another spatial cognitive test that is not reinforced by food rewards, contrary to the holeboard test), minipigs born from HFS mothers presented more stress and reduced cognitive abilities (Table 1; Gautier et al., Reference Gautier, Luneau, Coquery, Meurice, Malbert, Guerin, Kemp, Bolhuis, Clouard, Le Huerou-Luron, Blat and Val-Laillet2018). More interesting is the fact that these animals showed a reduced basal glucose metabolism in the prefrontal cortex and nucleus accumbens (Figure 1; maternal HFS diet), which are both parts of the brain reward circuit. We also demonstrated that these animals also presented a decreased dopamine and/or serotonin transporter availability in the striatum and hippocampus. These brain anomalies are usually observed in obese subjects, as previously demonstrated in humans (Wang et al., Reference Wang, Volkow, Logan, Pappas, Wong, Zhu, Netusil and Fowler2001; Le et al., Reference Le, Pannacciulli, Chen, Del Parigi, Salbe, Reiman and Krakoff2006; Volkow et al., Reference Volkow, Wang, Telang, Fowler, Thanos, Logan, Alexoff, Ding, Wong, Ma and Pradhan2008 and Reference Volkow, Wang, Telang, Fowler, Goldstein, Alia-Klein, Logan, Wong, Thanos, Ma and Pradhan2009) and minipigs (Val-Laillet et al., Reference Val-Laillet, Layec, Guerin, Meurice and Malbert2011a; Ochoa et al., Reference Ochoa, Malbert, Meurice and Val-Laillet2016a). It is consequently striking to find them in animals that were fed a standard diet from weaning without abnormal weight gain or metabolic disease until the adult age. This suggests that the HFS diet might program the brain even in the absence of weight problem and possibly modulate adaptation mechanisms to further dietary challenge. Further research is needed in this model to demonstrate that the brain and the digestive system, including gut microbiota, can influence each other with causal relationship to modulate the susceptibility to health and diseases, and notably behavioural and metabolic disorders.

Impact of food functional ingredient/supplementation on brain activity

Because many evidences indicate a crucial role for the microbiota in regulating different aspects of behaviour and mental health, as well as comorbidities related to eating and metabolic disorders (van de Wouw et al., Reference van de Wouw, Schellekens, Dinan and Cryan2017), pre- and probiotics open the way to novel biotherapeutic strategies. Pinto-Sanchez et al. (Reference Pinto-Sanchez, Hall, Ghajar, Nardelli, Bolino, Lau, Martin, Cominetti, Welsh, Rieder, Traynor, Gregory, De Palma, Pigrau, Ford, Macri, Berger, Bergonzelli, Surette, Collins, Moayyedi and Bercik2017) recently demonstrated that probiotic supplementation reduced depression scores and increased the quality of life in patients with inflammatory bowel disease. Interestingly, changes in brain activity via fMRI, with reduced reactivity of the limbic system, were also shown. Comparison data are lacking in the pig model, but the use of maternal prebiotics can impact the progeny’s growth, intestinal microbiota, morphology and immunity in pigs (Le Bourgot et al., Reference Le Bourgot, Ferret-Bernard, Le Normand, Savary, Menendez-Aparicio, Blat, Appert-Bossard, Respondek and Le Huerou-Luron2014, Reference Le Bourgot, Ferret-Bernard, Blat, Apper and Le Huërou-Luron2016, and Reference Le Bourgot, Le Normand, Formal, Respondek, Blat, Apper, Ferret-Bernard and Le Huerou-Luron2017). The use of dietary prebiotics early in life in piglets also supported recognition memory and modulated the concentrations of SCFAs in the colon, blood and the brain, as well as hippocampal serotonin (Fleming et al., Reference Fleming, Monaikul, Patsavas, Waworuntu, Berg and Dilger2017). Interestingly, we recently showed that oral supplementation of butyrate might shape brain activity and plasticity (Val-Laillet et al., Reference Val-Laillet, Guerin, Coquery, Nogret, Formal, Rome, Le Normand, Meurice, Randuineau, Guilloteau, Malbert, Parnet, Lalles and Segain2018). Butyrate is an SCFA that is naturally produced in the gut through bacterial fermentation of dietary fibres, playing an essential role in gastrointestinal functions (Guilloteau et al., Reference Guilloteau, Zabielski, Hammon and Metges2010). This microbiota metabolite (i.e. postbiotic) has a trophic role for colonocytes, modulatory properties on gut morphology and functional plasticity, and is part of the histone deacetylase inhibitors having positive effects on long-term memory processes (Guilloteau et al., Reference Guilloteau, Zabielski, Hammon and Metges2010; Val-Laillet et al., Reference Val-Laillet, Guerin, Coquery, Nogret, Formal, Rome, Le Normand, Meurice, Randuineau, Guilloteau, Malbert, Parnet, Lalles and Segain2018). For that matter, some authors suggested that butyrate could prevent memory impairment and improve cognition through the upregulation of neurotrophic factors (Moretti et al., Reference Moretti, Valvassori, Varela, Ferreira, Rochi, Benedet, Scaini, Kapczinski, Streck, Zugno and Quevedo2011; Valvassori et al., Reference Valvassori, Varela, Arent, Dal-Pont, Bobsin, Budni, Reus and Quevedo2014). To tackle this question in the pig model, we compared two groups of juvenile animals that were exposed for 3 weeks to either oral sodium butyrate or control treatment (Table 1; Val-Laillet et al., Reference Val-Laillet, Guerin, Coquery, Nogret, Formal, Rome, Le Normand, Meurice, Randuineau, Guilloteau, Malbert, Parnet, Lalles and Segain2018). We demonstrated that oral butyrate supplementation triggered basal brain glucose metabolism changes in the hippocampus and nucleus accumbens, which are involved in cognitive and hedonic processes (Figure 1g). Interestingly, post-mortem immunohistochemistry data revealed a higher granular cell layer volume, cell proliferation and neurogenesis with butyrate treatment, supporting a positive modulatory role of this postbiotic on hippocampal plasticity and neurocognitive processes (Val-Laillet et al., Reference Val-Laillet, Guerin, Coquery, Nogret, Formal, Rome, Le Normand, Meurice, Randuineau, Guilloteau, Malbert, Parnet, Lalles and Segain2018).

Apart from pre-, pro- and postbiotics, other kinds of food functional ingredients can be used to modulate behaviour and brain activity. Stressful environments/events are known to alter well-being and eating behaviour, sometimes leading to appetite loss and anorexia or to food cravings and hyperphagia. This has been described in humans (Guarda et al., Reference Guarda, Schreyer, Boersma, Tamashiro and Moran2015; Geiker et al., Reference Geiker, Astrup, Hjorth, Sjodin, Pijls and Markus2018) as in pigs (Clouard et al., Reference Clouard, Meunier-Salaun and Val-Laillet2012c). In young pigs, we demonstrated positive effects of sensory feed functional ingredients based on natural plant extracts on eating behaviour, with increased feed preference and intake after a feed transition, for example (Clouard et al., Reference Clouard, Meunier-Salaün and Val-Laillet2012b; Clouard and Val-Laillet, Reference Clouard and Val-Laillet2014). Then, we explored brain responses to one of these feed functional ingredients and showed that the perception of the ingredient (Figure 1b and c) in familiarized individuals induced different responses in reward and perception brain areas compared to familiar control feed flavour in naive animals (Table 1; Val-Laillet et al., Reference Val-Laillet, Meurice and Clouard2016). In addition to its potential application in animal production to improve well-being and feed intake, translational applications might be assessed in fragile human populations suffering from appetite loss, decreased motivation to eat and/or denutrition. These food functional ingredients might also be tested in the context of degraded/stressful environments to assess their ability to counterbalance the effects of stressors at the brain level.

Impact of chronic vagus nerve stimulation on brain activity

Chronic vagus nerve stimulation (VNS) is used in clinical practice as a therapy for refractory epilepsy and some psychiatric disorders (Vonck et al., Reference Vonck, De Herdt and Boon2009), but it is also a promising potential strategy to fight against obesity and an original methodological approach to understand how the gut–brain signals transmitted via the vagus nerve can modulate brain activity, metabolism and eating behaviour. The pig model was rapidly in good position among candidate animal models to assess VNS against obesity (for review see Val-Laillet et al., Reference Val-Laillet, Biraben, Randuineau and Malbert2010a). The vagus nerves convey much information emanating from the gut, notably information related to gastric distension, one of the first satiation signals. We showed in young pigs that the activity of several brain areas was found to be significantly associated with an increase in gastric distension pressure (Figure 1f), such as the olfactory bulb, prefrontal cortex and amygdala (Layec et al., Reference Layec, Lapouble, Val-Laillet, Guerin, Chauvin, Heresbach and Malbert2009; Sauleau et al., Reference Sauleau, Lapouble, Val-Laillet and Malbert2009). But after bilateral vagotomy, only a limited number of brain areas still responded to gastric pressure (Table 1; Sauleau et al., Reference Sauleau, Lapouble, Val-Laillet and Malbert2009). A pilot study also demonstrated that chronic gastric distension (via a gastric balloon; Figure 1f) activated the nucleus accumbens (Layec et al., Reference Layec, Lapouble, Val-Laillet, Guerin, Chauvin, Heresbach and Malbert2009). With VNS, the aim is to mimic the nervous gut–brain signals emitted with stomach distension. The first pilot study investigating brain responses to VNS was performed in lean juvenile pigs (Table 1; Biraben et al., Reference Biraben, Guérin, Bobillier, Val-Laillet and Malbert2008). At that time, nuclear functional brain imaging in the pig model was emerging, and our digital pig brain atlas (Saikali et al., Reference Saikali, Meurice, Sauleau, Eliat, Bellaud, Randuineau, Verin and Malbert2010) was not available, which explains why the identification of activated brain structures was tricky and the interpretation very limited. Two years later, we reiterated a similar experiment in obese adult minipigs and demonstrated that chronic VNS during 14 weeks decreased feed intake, prevented the animals from gaining further BW contrary to sham animals, and modified the feed preferences with a decreased consumption of high-sucrose feed and increased consumption of standard feed (Val-Laillet et al., Reference Val-Laillet, Biraben, Randuineau and Malbert2010a).

With time, both VNS and brain imaging procedures were improved in the pig model, and two new studies were published (Malbert et al., Reference Malbert, Picq, Divoux, Henry and Horowitz2017b and Reference Malbert, Bobillier, Picq, Divoux, Guiraud and Henry2017a). A first experiment compared three chronic VNS patterns in growing pigs (S1, S2 and S3; Table 1). Only S2 and S3 activated the dorsal vagal complex and had an impact on the prefrontal cortex, contrary to S1 (Table 1; Malbert et al., Reference Malbert, Bobillier, Picq, Divoux, Guiraud and Henry2017a; Figure 1h). Only S2 and S3 significantly reduced daily ingestion, the duration of eating bouts during the meal, as well as the consumption of the high-lipid diet during the three-choice feed test, where the animals had simultaneous access to control, high-lipid and high-glucose feeds provided by a three-trough robotic feeder (Malbert et al., Reference Malbert, Bobillier, Picq, Divoux, Guiraud and Henry2017a). Taken together, these results demonstrate how specific nervous signals emanating from the gut can modulate the activity of the brain network involved in food intake regulation and goal-oriented behaviour. This also highlights the importance of the prefrontal cortex in modulating food motivation and preferences. Another study in obese minipigs reused the same VNS procedure as that previously successfully tested (Val-Laillet et al., Reference Val-Laillet, Biraben, Randuineau and Malbert2010a) to investigate the VNS impact on metabolism and brain activity (Table 1; Malbert et al., Reference Malbert, Picq, Divoux, Henry and Horowitz2017b). Mean whole-body insulin sensitivity and hepatic glucose uptake rate were significantly lower in non-stimulated obese minipigs compared to lean minipigs, and contrary to stimulated obese minipigs. Stimulated animals also had increased glucose metabolism in the anterior prefrontal cortex and cingulate cortex compared to non-stimulated animals (Table 1; Malbert et al., Reference Malbert, Picq, Divoux, Henry and Horowitz2017b; Figure 1h). Once again, this study demonstrated that chronically mimicking satiation signals emanating from the gut could improve the activity of the prefrontal cortex, which is known to have inhibitory inputs towards the orexigenic brain areas and to be deactivated in obese individuals.

Impact of bariatric surgery on brain activity

To conclude this section on interventional strategies, it is interesting to evoke what is still one of the most effective therapeutic compromises against morbid obesity: the Roux-en-Y gastric bypass (RYGB; Kang and Le, Reference Kang and Le2017). A recent paper reviewed the evidences derived from neuroimaging and animal (mainly rodent) studies (Hankir et al., Reference Hankir, Seyfried, Miras and Cowley2018), explaining how a metabolic surgical procedure such as the RYGB, which is drastically reorganizing gut, including peptide hormones and microbiota metabolites, is also reshaping homeostatic and hedonic brain processes. Different animal models of bariatric surgery have been described (Rao et al., Reference Rao, Rao and Kini2010), and the first demonstrations of RYGB in the pig model, to our knowledge, date back to the 1990s (Frantzides et al., Reference Frantzides, Carlson and Schulte1995; Potvin et al., Reference Potvin, Gagner and Pomp1997). Since then, the pig and minipig models have been widely used to describe the functional mechanisms of RYGB. Not being exhaustive, we can refer to studies having investigated gastrointestinal histological morphology (Gentileschi et al., Reference Gentileschi, Gagner, Milone, Kini and Fukuyama2006), long-term weight loss in superobese minipigs (Birck et al., Reference Birck, Vegge, Stockel, Gogenur, Thymann, Hammelev, Sangild, Hansen, Raun, von Voss and Eriksen2013), changes in adipose tissue distribution (Chavez-Talavera et al., Reference Chavez-Talavera, Baud, Spinelli, Daoudi, Kouach, Goossens, Vallez, Caiazzo, Ghunaim, Hubert, Lestavel, Tailleux, Staels and Pattou2017), β-cell function, biliary function and glucose metabolism (Lindqvist et al., Reference Lindqvist, Spegel, Ekelund, Garcia Vaz, Pierzynowski, Gomez, Mulder, Hedenbro, Groop and Wierup2014; Sham et al., Reference Sham, Simianu, Wright, Stewart, Alloosh, Sturek, Cummings and Flum2014; Baud et al., Reference Baud, Daoudi, Hubert, Raverdy, Pigeyre, Hervieux, Devienne, Ghunaim, Bonner, Quenon, Pigny, Klein, Kerr-Conte, Gmyr, Caiazzo and Pattou2016; Chavez-Talavera et al., Reference Chavez-Talavera, Baud, Spinelli, Daoudi, Kouach, Goossens, Vallez, Caiazzo, Ghunaim, Hubert, Lestavel, Tailleux, Staels and Pattou2017; Lindqvist et al., Reference Lindqvist, Ekelund, Garcia-Vaz, Stahlman, Pierzynowski, Gomez, Rehfeld, Groop, Hedenbro, Wierup and Spegel2017a), as well as incretins regulation (Lindqvist et al., Reference Lindqvist, Ekelund, Pierzynowski, Groop, Hedenbro and Wierup2017b), for example. Despite all of this work, we found no study on the consequence of RYGB (or any other bariatric surgery) on brain activity and metabolism in pigs.

Identifying a way to reproduce the gut–brain effects of RYGB without a real surgery per se would free the patients from possible deleterious consequences of invasive bariatric surgery (Lupoli et al., Reference Lupoli, Lembo, Saldalamacchia, Avola, Angrisani and Capaldo2017). That is why preclinical research in a relevant model, such as the pig, is necessary. In a longitudinal study (Gautier Y., Bergeat D., et al., unpublished data), we aimed at phenotyping diet-induced obese Yucatan minipigs subjected either to the RYGB (cf. Baud et al., Reference Baud, Daoudi, Hubert, Raverdy, Pigeyre, Hervieux, Devienne, Ghunaim, Bonner, Quenon, Pigny, Klein, Kerr-Conte, Gmyr, Caiazzo and Pattou2016 for surgery procedure) or a pair-fed restrictive dietary treatment. Peptide and hormone measurements, gastrointestinal histological morphology and immunohistochemistry, transcriptomic, metabolomic and metagenomic analyses were performed to provide a systemic description of the intestinal (including microbiota) and metabolic phenotype. The obese minipigs were also subjected to fMRI and PET brain imaging before and 3 months after the surgery (RYGB or sham) to investigate brain responses to sucrose (Figure 1d and f). Our preliminary results (Table 1; Bergeat D., et al., unpublished data) are in line with human brain imaging studies showing a modulation of the mesolimbic and frontostriatal pathway after RYGB (Ochner et al., Reference Ochner, Kwok, Conceicao, Pantazatos, Puma, Carnell, Teixeira, Hirsch and Geliebter2011; Scholtz et al., Reference Scholtz, Miras, Chhina, Prechtl, Sleeth, Daud, Ismail, Durighel, Ahmed, Olbers, Vincent, Alaghband-Zadeh, Ghatei, Waldman, Frost, Bell, le Roux and Goldstone2014). To date, the minipig is the only preclinical model for which such an analogy has been described, which opens the way to further interventional studies aiming at unravelling the complex gut–brain interactions shaping eating behaviour after bariatric surgery.