Cholecystokinin

Depending on the amount and composition of the diet, CCK is secreted by the I cells in the duodenal and jejunal mucosa and also by neurons in the enteric nervous system. CCK contributes to satiation and satiety effects. The major effects of CCK are stimulation of gall bladder contraction, delay of gastric emptying and stimulation of pancreatic secretion⁽ Reference Maljaars, Symersky and Kee ^{63 )}. In pigs, CCK release is stimulated by a mixed meal from fasting to postprandial (30–120min) concentrations going from about 10pmol/l to about 30pmol/l, respectively, with a faster response when diets are enriched with starch than with fat⁽ Reference Corring and Chayvialle ^{71 )}. Clutter et al. ⁽ Reference Clutter, Jiang and McCann ^{72 )} measured plasma CCK-8 in pigs, at fasting about 2pmol/l increasing to 10pmol/l at mixed meal feeding whereas Ripken et al. ⁽ Reference Ripken, van der Wielen and van der Meulen ^{73 )} reported concentrations of 0·5pmol/l and 2pmol/l for fasting and postprandial, respectively. All experiments showed a 2- to 5-fold increase in plasma CCK-8 concentrations after mixed meal feeding. Another study in pigs⁽ Reference Jakob, Mosenthin and Zabielski ^{74 )}, using intraduodenal fat infusion, showed plasma CCK concentrations of approximately 5pmol/l both prior and following fat infusion. CCK release was higher to long-chain FA than to medium-chain TAG. In humans, CCK release is mostly stimulated by the presence of dietary protein, whereas carbohydrate provides a weaker stimulus⁽ Reference Liddle, Goldfine and Rosen ^{75 )}. To our knowledge, the effect of dietary protein on plasma CCK secretion has not been investigated to date in pigs. Plasma CCK concentrations are similar in pigs and humans, on average for humans at fasting 2pmol/l and at feeding 10pmol/l⁽ Reference Feinle, Grundy and Otto ^{76 –} Reference Mossner, Grumann and Zeeh ^{79 )}. In both species the magnitude of the CCK response is dependent on the composition of the meal. The effects of single macronutrients or combinations thereof on CCK release have not been investigated systematically in humans or pigs. The pig is a valid model to investigate these relationships.

Anika et al. ⁽ Reference Anika, Houpt and Houpt ^{80 )} administered CCK intravenously at a rate of 0·2nmol/kg per min for total doses of 0·4, 0·8, 1·7 and 3·4nmol/kg to pigs. The lowest dose of CCK reduced food intake by 13 % and the highest dose by 94 %. Houpt⁽ Reference Houpt ^{81 )} showed that intravenous (iv) CCK-8 (the synthetic and bioactive part of the CCK molecule) infusion to pigs of 14, 28 and 56 pmol/kg per min decreased meal size in a dose-dependent manner. A dose of 28pmol/kg per min reduced food intake by 30–40 %. Similar results were found by Baldwin et al. ⁽ Reference Baldwin, Cooper and Parrott ^{82 )} who studied hungry pigs working for food, thirsty pigs responding for water, and non-deprived pigs working for SUC. In this study CCK produced significant dose-related decreases in response rates for all three conditions.

Nutrients exert much of their effects through the CCK-1 receptor⁽ Reference Holzer, Turkelson and Solomon ^{83 )}. There are two regions where CCK acts to produce satiety: the nucleus tractus solitarius in the hindbrain and the medial-basal hypothalamus⁽ Reference Reidelberger, Hernandez and Fritzsch ^{84 )}. CCK acts both at CCK-1 receptors beyond the BBB and by a CCK-1 receptor-mediated mechanism involving abdominal vagal nerves to inhibit food intake⁽ Reference Reidelberger, Hernandez and Fritzsch ^{84 )}. CCK-1 antagonists, such as devazepide (DVZ), are commonly used in pig appetite and satiety studies⁽ Reference Ebenezer, Vellucci and Parrott ^{85 )}. DVZ easily passes the BBB following iv administration inducing a dose-related increase in food intake at doses ranging from 17·5 to 140µg/kg with maximum increases occurring at about 70µg/kg⁽ Reference Reidelberger and O’Rourke ^{86 –} Reference Ebenezer, de la Riva and Baldwin ^{88 )}. Arterial injection of DVZ (0·1mg/kg) in pigs⁽ Reference Gregory, McFadyen and Rayner ^{89 )} abolished the inhibition of food intake to duodenal infusion of emulsified fat and monoacylglycerols. However, it did not alter the inhibition of intake in response to oleic acid, to glycerol or to GLU, suggesting that monoacylglycerol-induced CCK secretion is mainly responsible for the satiety resulting from duodenal fat infusion in the pig. Baldwin & Sukhchai⁽ Reference Baldwin and Sukhchai ^{90 )} reported that intracerebroventricular (icv) injection of DVZ (100µg) before the administration of 1µg CCK abolished the inhibition of GLU intake produced by CCK in pigs. However, DVZ itself had no effect on GLU intake. In a study by Farmer et al. ⁽ Reference Farmer, Roy and Rushen ^{91 )}, pigs were fasted for 24h, injected intravenously with DVZ at 70mg/kg and subsequently subjected to a feed motivation test (operant conditioning). The number of pushes, duration of eating and amount of feed eaten during the feed motivation test were all increased by fasting, and were further increased by DVZ injection, indicating that CCK induces satiation in pigs. An alternative CCK receptor antagonist used in pigs is 2-NAP (2-naphthalenesulfanyl-l-aspartyl-2-(phenethyl) amide), which does not cross the BBB. Baldwin et al. ⁽ Reference Baldwin, de la Riva and Gerskowitch ^{92 )} revealed that iv administration (20 or 40mg/kg) of NAP injected before iv administration of CCK-8 (1µg/kg) abolished the inhibitory effects of CCK-8 on food intake in hungry pigs. Also, NAP abolished the inhibitory effects of CCK-8 on food intake in hungry pigs after icv injection of both compounds. Overall, these results indicate that endogenous CCK is involved in the regulation of satiety in pigs, in a way which is similar to what is known in humans⁽ Reference Wolkowitz, Gertz and Weingartner ^{93 )}, albeit the pig offers the possibility to conduct invasive mechanistic studies on CCK action.

Glucagon-like peptide-1

GLP-1 is mostly produced by the L cells in the distal small intestine and colon and contributes to satiety. It performs a variety of functions in the body such as inhibiting acid secretion and gastric emptying, while increasing insulin secretion from the pancreas in a GLU-dependent manner⁽ Reference Holst ^{94 )}. The secretion of GLP-1 is mediated by indirect, duodenal activated neurohumoral mechanisms, as well as by direct contact of nutrients with the distal small intestine⁽ Reference Baldwin, Cooper and Parrott ^{82 )}. Souza da Silva et al. ⁽ Reference Souza da Silva, Haenen and Koopmans ^{95 )} showed that fasting plasma GLP-1 concentrations of 15pmol/l rose to 35pmol/l 1h after the beginning of a complete mash feed. Hooda et al. ⁽ Reference Hooda, Matte and Vasanthan ^{96 )} showed that fasting plasma GLP-1 concentrations in pigs were approximately 8pmol/l and rose postprandial to 20pmol/l, 1–2h after initiation of the mixed-nutrient meal. The rise of plasma GLP-1 in response to intraduodenal infusion of GLU, fat and GLU–fat in pigs was modest (+5pmol/l) for GLU and intermediate (+10pmol/l) for fat, while the effect of combining GLU and fat was additive (+15pmol/l)⁽ Reference Knapper, Morgan and Fletcher ^{97 )}. To our knowledge, the effect of dietary protein on plasma GLP-1 secretion has not been investigated to date in pigs. In general, the GLP-1 responses to fasting/feeding in pigs are comparable with those in humans, with fasting concentrations of about 13pmol/l rising to 30–40pmol/l postprandial⁽ Reference Blom, Lluch and Stafleu ^{78 ,} Reference Lavin, Wittert and Andrews ^{98 ,} Reference Verdich, Toubro and Buemann ^{99 )}.

Although GLP-1 can cross the BBB, several studies suggest that peripheral GLP acts to reduce food intake primarily via activation of the vagal afferent nerve⁽ Reference Asmar ^{100 )}. Little information is available on the effect of GLP-1 on food intake in pigs. Ribel et al. ⁽ Reference Ribel, Larsen and Rolin ^{101 )} showed that administration of a synthetic GLP-1 analogue reduced gastric emptying and therefore may induce satiation in pigs. In general, the pig is recognised as being a good model for studying human conditions⁽ Reference Litten-Brown, Corson and Clarke ^{102 )}, and in the case of GLP-1 analogues, it has found to be predictive of clinical findings in both reduction of hyperglycaemia (through improved insulin secretion and decreased glucagon secretion) and body-weight loss (through reduced gastric emptying and increased satiety)⁽ Reference Knudsen ^{103 )}. The icv administration of GLP-1 inhibited feeding in fasted rats and was counteracted by the icv injection of exendin 9–39, a GLP-1 receptor antagonist⁽ Reference Turton, O’Shea and Gunn ^{104 )}. No data are available in pigs.

Peptide YY

PYY is an anorexigenic hormone. It is a member of the pancreatic polypeptide (PP)-fold family of peptides. It is secreted by the L cells of the distal small-intestinal mucosa. The PYY-containing endocrine cells are found in highest numbers in the lower small intestine and colon. The gastrointestinal functions of PYY include inhibition of gastric acid and pepsin secretion, inhibition of pancreatic exocrine secretion, delay of gastric emptying and inhibition of jejunal and colonic motility⁽ Reference Sheikh, Holst and Orskov ^{105 )}. The secretion of PYY is stimulated by the presence of nutrients in the intestine. In humans, fats provide the strongest stimulus followed by carbohydrates and protein⁽ Reference Degen, Oesch and Casanova ^{106 )}. Little is known on the effects of intestinal nutrient loads on PYY secretion in pigs. Plasma PYY concentrations were higher (500pmol/l) in ad libitum-fed pigs compared with fasted pigs (200pmol/l)⁽ Reference Ito, Thidarmyint and Murata ^{107 )}. On the other hand, Souza da Silva et al. ⁽ Reference Souza da Silva, Haenen and Koopmans ^{95 )} showed that fasting plasma PYY concentrations were approximately 700–800pmol/l, showing no response to a mixed-nutrient meal. Plasma PYY concentrations are lower in humans, ranging from 30pmol/l at fasting up to 116pmol/l after an intestinal nutrient load⁽ Reference Seimon, Feltrin and Meyer ^{77 ,} Reference Degen, Oesch and Casanova ^{106 )}. In the fast–refed condition, both single bolus injection (30mg/kg body weight) and iv infusion (0·25mg/kg per min) of PYY3–36 (the synthetic and bioactive part of the PYY molecule) suppressed feed intake in pigs⁽ Reference Ito, Thidarmyint and Murata ^{107 )}, suggesting that circulating PYY3–36 influences satiety and contributes to the termination of a meal such as in humans⁽ Reference Perry and Wang ^{68 )}.

Ghrelin

Ghrelin is an orexigenic intestinal hormone. In pigs, it is mostly produced in the oxyntic and cardiac gland and less commonly in the pyloric glands of the stomach⁽ Reference Govoni, De Iasio and Cocco ^{108 )}, with a similar pattern to that in humans⁽ Reference Kojima and Kangawa ^{109 )}. Ghrelin functions as a neuropeptide signal that reduces satiety and increases hunger. Govoni et al. ⁽ Reference Govoni, De Iasio and Cocco ^{108 )} and Zhang et al. ⁽ Reference Zhang, Yin and Li ^{110 )} showed that fasting increases plasma ghrelin concentrations from 10–25 to 50pmol/l in prepubertal gilts and weanling pigs, respectively. Ghrelin in pigs responded to changes in energy balance and the concentrations increased during fasting from approximately 25 to 75pmol/l⁽ Reference Govoni, De Iasio and Cocco ^{108 ,} Reference Inoue, Watanuki and Myint ^{111 )}. Barretero-Hernandez et al. ⁽ Reference Barretero-Hernandez, Galyean and Vizcarra ^{65 )} reported postprandial plasma ghrelin concentrations of 6pmol/l, increasing to 10pmol/l at fasting in pigs. Scrimgeour et al. ⁽ Reference Scrimgeour, Gresham and Giles ^{112 )} suggested that the dynamics in plasma ghrelin concentrations in the pig appear to be strongly influenced by prolonged fasting and change from 15pmol/l (feeding) to 100pmol/l (fasting). These ghrelin responses to fasting/feeding are comparable with those observed in humans⁽ Reference Blom, Lluch and Stafleu ^{78 ,} Reference Cummings, Purnell and Frayo ^{113 )}. However, the concentrations in humans are higher, being approximately 150pmol/l postprandial and 250pmol/l at fasting.

Salfen et al. ⁽ Reference Salfen, Carroll and Keisler ^{114 )} showed that there was an increase in body weight of weanling pigs given exogenous ghrelin chronically, suggesting that feeding behaviour was influenced by the treatment. Ghrelin stimulates food intake in humans too. However, in vagotomised patients, ghrelin does not increase food intake, suggesting that an intact vagus nerve is required for exogenous ghrelin to increase appetite in humans⁽ Reference Le Roux, Neary and Halsey ^{115 )}. No pig studies have been conducted addressing the contribution of the vagus nerve to the orexogenic action of ghrelin.

Dominant phyla

The largest microbiota of the body is located in the GIT and the set of gene products provides a diverse range of biochemical and metabolic activities to complement host physiology. Hundreds of species are present in the GIT lumen, only belonging to a few microbial phyla. Firmicutes and Bacteroidetes are the two dominant bacterial phyla in the human and mouse gut, with the Proteobacteria, Actinobacteria, Fusobacteria and Verrucomicrobia phyla as subdominant phyla⁽ Reference Ley, Hamady and Lozupone ^{171 ,} Reference Eckburg, Bik and Bernstein ^{182 –} Reference Ley, Turnbaugh and Klein ^{184 )}. Similarly, the GIT microbiota in pigs, as well as in wild suidae, mainly consists of the Firmicutes and Bacteroidetes phyla⁽ Reference Leser, Amenuvor and Jensen ^{185 ,} Reference Guo, Xia and Tang ^{186 )}(Table 3). Recently two different enterotype-like clusters, primarily distinguished by unclassified Ruminococcus and Prevotella, have been identified in pig faeces⁽ Reference Mach, Berri and Estelle ^{175 )}. Their phylogenetic composition was highly similar to two of the enteroptypes described in humans⁽ Reference Arumugam, Raes and Pelletier ^{173 )}. Interestingly, in pigs, as in humans, enterotype-like clustering distribution can vary within an individual over time⁽ Reference Knights, Ward and McKinlay ^{187 )}.

Table 3 Mean values of the amount of total SCFA throughout life in pig and human faeces

* SCFA content in meconium; values adapted from Midtvedt & Midtvedt⁽ Reference Midtvedt and Midtvedt ^{414 )}.

† Commercial pigs (Large White × Landrace × Pietrain breed) weaned 28d of age, data expressed per kg of faecal DM (Montagne et al. ⁽ Reference Montagne, Le Floc’h and Arturo-Schaan ^{415 )}).

‡ 5d post-weaning.

§ Post-pubescent (6·4 and 7·5 months-old Large White × Landrace × Pietrain breed) and gestating sows (1·5–2 years-old Large White × Landrace breed), data expressed per kg of faecal fresh matter (I Le Huerou-Luron, unpublished results).

Postnatal and early life microbial colonisation

During the first few hours after birth (postnatal), contact with environmental and colonising bacteria is essential for healthy intestinal and immune maturation. The major role of microbiota in the development of the neonatal GIT was confirmed in conditions where colonisation was modified early in life through exposure to micro-organisms of maternal and environmental origins, through nutrients consumed and through antibiotic treatments. During infancy, the composition of the intestinal microbiota is unstable and more variable than in older children and adults. Diet-induced adaptation of the microbiota may vary from the proximal to the distal parts of the intestine. The composition that is typically measured from faecal samples does not reflect the large bacterial diversity along the intestinal tract. Animal models, specifically cannulated pigs, are useful for obtaining a better understanding of the interactions between microbiota present in different niches of the intestine and physiology relevant to humans⁽ Reference Saraoui, Parayre and Guernec ^{188 )}. Studies with germ-free piglets clearly show that bacteria are essential for the growth and development of the digestive tract⁽ Reference Chowdhury, King and Willing ^{189 )}. The comparison of gene expression profiles in enterocytes of germ-free compared with conventional piglets has brought insight on the impact of microbiota on the GIT function⁽ Reference Chowdhury, King and Willing ^{189 )}. Bacterial colonisation induces the maturation and function of several components of the mucosal immune system and defence in order to prevent inflammatory responses that would compromise the barrier function⁽ Reference Chowdhury, King and Willing ^{189 –} Reference Hooper ^{191 )}. In humans, neonates compared with older individuals have a decreased innate defence, a low production of IgA and a defective interaction between dendritic cells, T lymphocytes and regulatory T cells⁽ Reference El Aidy, Dinan and Cryan ^{192 )}. Similarly, the mucosal immune system is essentially absent in the neonatal piglet, even though the systemic immune tissue is well developed⁽ Reference Bailey, Haverson and Inman ^{193 )}, and piglets begin to synthetise secretory IgA from the second week of age⁽ Reference Le Huërou-Luron and Ferret-Bernard ^{194 )}. The balance between T lymphocytes helper 1 and helper 2 responses in human and pig neonates is skewed toward the helper 2 profile, resulting in a high susceptibility to intracellular pathogen infection⁽ Reference Le Bourgot, Ferret-Bernard and Apper-Bossard ^{195 ,} Reference Adkins, Leclerc and Marshall-Clarke ^{196 )}. The impaired protection of the neonate against infections may be partly attributed to a deficient secretion of interferon⁽ Reference Le Bourgot, Ferret-Bernard and Apper-Bossard ^{195 ,} Reference Wilson, Westall and Johnston ^{197 )}.

It only takes a few hours for bacteria to appear in the faeces of mammalian neonates⁽ Reference Thompson, Wang and Holmes ^{198 ,} Reference Dore and Corthier ^{199 )}. Facultative anaerobic bacteria, such as Proteobacteria, are the first colonisers. These bacteria reduce oxygen concentration in the GIT and allow strict anaerobes, such as members from the genus Bacteroides and the phyla Actinobacteria and Firmicutes, to colonise the intestine. During the first year of life in humans and the first 6 months in pigs, the intestinal microbiota composition fluctuates widely between individuals and over time, before resembling the adult status (Table 4). The potential use of piglets as a model for human studies is reinforced by the early colonisers, Bacteroides and Escherichia/Shigella being similar in humans. However, the substantial presence of Lactobacillus and Streptococcus in pigs is unparalleled in humans.

Table 4 Comprehensive summary of the existing literature on the relationship between nutrition and brain composition/development in pig models

FA, fatty acids; ARA, arachidonic acid; DPA, docosapentaenoic acid; LA, linoleic acid; ALA, α-linolenic acid; MCT, medium-chain TAG; l-DOPA, l-3,4-dihydroxyphenylalanine; HC, high cholesterol; LC, low cholesterol; ppm, parts per million.

* Swainsonine is a plant toxin.

† Quercetin is a dietary polyphenol with potential health benefits.

Early disturbances of the microbial colonisation process, such as induced by high-hygiene environments or by antibiotic treatment, have major consequences for the developmental sequence of the GIT microbiota and for host metabolism⁽ Reference Cox and Blaser ^{200 )}. An advantage of the porcine model is the flexibility to compare different early environmental-rearing conditions, using, for example, outdoor and indoor sow-reared piglets or isolator-reared neonates. Mulder et al. ⁽ Reference Mulder, Schmidt and Stokes ^{201 )} showed large differences in composition of ileal-adherent microbiota between outdoor and indoor sow-reared animals, which corresponded to major differences in intestinal immune activation. Excessive hygiene appears to interfere with the normal processes of bacterial stabilisation and alters immune development. Mulder et al. ⁽ Reference Mulder, Schmidt and Lewis ^{202 )} showed that the succession of events that lead to a stable adult microbiota depends on colonisation during the first 2d of life, and also on continuous exposure to highly diverse microbiota during the early development at least up to weaning at 4 weeks of age. The use of antibiotics, in combination with stressors in early life, was shown to affect adult pig microbiota and intestinal gene expression, including genes involved in immune-related processes⁽ Reference Schokker, Zhang and Vastenhouw ^{203 )}. This observation in pigs corroborates human studies indicating that changing the environmental conditions, and in particular microbial exposure, throughout early life affects the development of immune diseases⁽ Reference Pinsk, Lemberg and Grewal ^{204 )}. Whether inducing early change in immune homeostasis by modifying microbiota would lead to different sensitivity of pigs to infectious or inflammatory challenge, such as recently reported with early spray-dried supplementation⁽ Reference Boyer, D’Costa and Edwards ^{205 )}, warrants further investigation.

Many beneficial strategies have been suggested to strengthen the postnatal development of presumably beneficial microbiota and GIT functions, including: changing the composition of maternal food during gestation and lactation; changing the composition of infant formulas; and favouring breast-feeding over formula-feeding during the suckling period⁽ Reference Heinritz, Mosenthin and Weiss ^{180 ,} Reference Le Huërou-Luron, Blat and Boudry ^{206 )}. For example, feeding neonatal piglets with formula supplemented with prebiotics increased the bacterial numbers by 5-fold, the content of folic acid by 53 % and growth of the colon⁽ Reference Aufreiter, Kim and O’Connor ^{207 )}. Similarly, supplementation of the sow diet with prebiotics during gestation and lactation was associated with 50 % greater fermentative activity of the caecal microbiota, accelerated development of the intestinal immunity, and improved intestinal protection by increasing ileal Peyer’s patch production of secretory IgA in the offspring by 46 %⁽ Reference Le Bourgot, Ferret-Bernard and Apper-Bossard ^{195 )}. Faecal secretory IgA also increased by 170 % in healthy infants who receive a prebiotic-supplemented formula⁽ Reference Scholtens, Alliet and Raes ^{208 )}. These results in pigs and humans underline the key role of maternal nutrition during pregnancy in supporting neonatal development of the GIT immune system via modulation of microbiota.

The health benefits of breast-feeding have been recognised for a long time. Breast-feeding is associated with earlier colonisation with bifidobacteria, partly in relation to the presence of oligosaccharides in human maternal milk⁽ Reference Donovan, Wang and Li ^{177 )}. As in humans, breast-fed piglets showed lower intestinal growth and permeability compared with high protein formula-fed ones⁽ Reference Boudry, Morise and Seve ^{166 ,} Reference Donovan, Wang and Li ^{177 )}. One major issue in human studies on the effect of breast- v. formula-feeding on gut function is the great number of confounding factors which are difficult to circumvent, including quantification of food intake in breast-fed infants, variable length of exclusive breast-feeding, and variability of the composition of milk formulas. Animal models are used to help control these confounding factors, in particular, use of an automatic milk feeder that provides neonatal piglets with artificial milk as similar as possible to maternal milk⁽ Reference Morise, Seve and Mace ^{209 )}. These studies using a formula-fed piglet model provide strong support for the idea that short dietary changes before weaning associated with a modification of the early intestinal bacterial colonisation can have a long-term impact on the severity of inflammatory responses without changing the basal physiology of the intestinal barrier and cytokine profile in the intestine⁽ Reference Chatelais, Jamin and Gras-Le Guen ^{157 ,} Reference Boudry, Jamin and Chatelais ^{158 )}. No similar data are available from human studies due to the invasive procedure required for intestine functionality research. However, breast-feeding is clearly associated with lower incidence of necrotising enterocolitis and diarrhoea in both human and pig neonates⁽ Reference Le Huërou-Luron, Blat and Boudry ^{206 ,} Reference Siggers, Siggers and Thymann ^{210 )}.

Dominant phyla

Differences in the most abundant genera exist between the human and the pig intestinal microbiota⁽ Reference Heinritz, Mosenthin and Weiss ^{180 )}. Belonging to the Bacteroidetes phylum, the most abundant genus is Bacteroides in humans, averaging 9 to 42 % of total bacteria⁽ Reference Dore and Corthier ^{199 )}, while the most abundant genus is Prevotella in weaned pigs, accounting for more than 20 % of total bacteria⁽ Reference Mach, Berri and Estelle ^{175 ,} Reference Kim, Borewicz and White ^{211 )}. In adult humans, the phylum Actinobacteria may represent up to 15 % of total bacteria. It comprises bifidobacteria, the most predominant group detected in infants (40 % in average in faecal samples of 6-week-old European infants)⁽ Reference Fallani, Young and Scott ^{212 )}. The population of bifidobacteria present in the intestine of pigs is considerably lower, with less than 0·1 % of total sequences in the faecal samples of 22-week-old commercial pigs⁽ Reference Kim, Borewicz and White ^{211 )}. Dietary, environmental and behavioural (such as feeding habits) factors contribute to the species-specificity of the composition of microbiota.

Epigenetic mechanisms

The concept that early developmental dietary insults (poor or inadequate pre- or postnatal nutrition, for example) can have long-term consequences on health later in life has been termed developmental programming, or ‘developmental origins of health and disease’. The GIT microbiota appears to have an important role in the GIT programming as its initial composition creates distinct individuality during the lifespan⁽ Reference Weng and Walker ^{167 ,} Reference Palmer, Bik and DiGiulio ^{213 ,} Reference Jakobsson, Jernberg and Andersson ^{214 )}. A mechanism leading to these long-term effects may be due to epigenetically active fermentation metabolites such as in histone acetylation⁽ Reference Strasak, Bartova and Harnicarova ^{215 ,} Reference Mischke and Plosch ^{216 )}. However, no studies on epigenetic modifications underlying long-term effects on early microbial colonisation and gut function are available in pigs. Despite limited studies, use of the pig as a model for humans to assess the effects of early nutrition on the development of microbiota is gaining acceptance amongst the scientific community.

Brain responses to food signals

Describing brain responses to food signals is important to investigate food pleasure and motivation, or to decipher the brain networks underlying sensory and nutrient perception. An extensive literature is available on this topic in the human, describing mostly via functional MRI the brain responses to various food signals, according to different internal states (for example, hungry v. sated) or conditions (for example, lean v. obese), and many review papers are available on this topic (for example, Rolls⁽ Reference Rolls ^{221 )}; Stice et al. ⁽ Reference Stice, Spoor and Ng ^{222 )}; Carnell et al. ⁽ Reference Carnell, Gibson and Benson ^{223 )}). Studies using large animal models have not yet completely made use of the opportunities provided by in vivo brain imaging. The very first studies using functional imaging to describe food-induced brain responses in pigs used ^99mTc-HMPAO (technetium hexamethylpropyleneamine oxime) SPECT (single photon emission computed tomography) and [¹⁸F]fluorodeoxyglucose positron emission tomography to map cerebral blood flow⁽ Reference Boubaker, Val-Laillet and Guerin ^{224 –} Reference Gaultier, Meunier-Salaun and Malbert ^{226 )} and brain GLU metabolism⁽ Reference Clouard, Jouhanneau and Meunier-Salaun ^{227 )}, respectively. These studies addressed the use of the pig to study food conditioning, looking at specific modulations of the response of the brain reward circuit after exposure to flavours with positive or negative hedonic values⁽ Reference Gaultier, Meunier-Salaun and Malbert ^{226 ,} Reference Clouard, Jouhanneau and Meunier-Salaun ^{227 )}. These studies provide two major outcomes: (1) functional imaging in anaesthetised pigs can be implemented to explore brain responses to different food signals, as in humans; and (2) the brain circuits activated by the perception of food signals are similar to those described in the human (for example, frontostriatal areas, amygdala and insular cortex). Boubaker et al. ⁽ Reference Boubaker, Val-Laillet and Guerin ^{224 )} showed in pigs that duodenal and portal GLU infusions led to different systemic and brain responses in areas regulating food intake and pleasure. Clouard et al. ⁽ Reference Clouard, Meunier-Salaün and Meurice ^{225 )} compared congruent v. dissociated oral and duodenal SUC perception, and found different brain responses in the limbic and reward circuits. Studies in human subjects showed that brain responses to energy-providing sugars and sweeteners are not the same in the reward circuit (for reviews, see Low et al. ⁽ Reference Low, Lacy and Keast ^{228 )} and Ochoa et al. ⁽ Reference Ochoa, Lallès and Malbert ^{229 )}), which resembles the results obtained by Clouard et al. ⁽ Reference Clouard, Meunier-Salaün and Meurice ^{225 )} in pigs. These studies are important to understand how the human brain correlates sugar cravings, as well as the neurobehavioural changes that could emerge due to the chronic consumption of, for example; sugars or non-energy sweeteners.

Impact of diet on brain activity, neurotransmission and cognition

Minipigs have become a widely accepted model for studying obesity and the metabolic syndrome⁽ Reference Johansen, Hansen and Richelsen ^{230 –} Reference Val-Laillet, Guerin and Malbert ^{234 )}. They can be used to investigate the obesity-induced central modifications in humans, including decreased activity of the prefrontal cortex⁽ Reference Le, Pannacciulli and Chen ^{235 –} Reference Volkow, Wang and Telang ^{237 )} and altered dopaminergic function⁽ Reference Volkow, Wang and Telang ^{237 ,} Reference Wang, Volkow and Logan ^{238 )}. Val-Laillet et al. ⁽ Reference Val-Laillet, Layec and Guerin ^{239 )} demonstrated in the Göttingen minipig that brain alterations similar to those described in obese humans exist in this model and that they are an acquired feature of obesity correlated to weight gain. In Pitman–Moore minipigs, Val-Laillet et al. ⁽ Reference Val-Laillet, Meurice and Lalles ^{240 )} also described the effects of three high-lipid diets differing in their lipid sources and found that the basal GLU metabolism of the anterior prefrontal cortex and nucleus accumbens was highest with a diet enriched with sunflower-seed oil, intermediary with a diet enriched with lard, and lowest with a diet enriched with fish oil. These results demonstrate that specific dietary nutrients can modify brain metabolism independently from body weight, and that specific nutrients in excess might favour the onset of brain metabolism anomalies.

In humans, cognitive test scores were positively related to breast milk DHA and negatively related to linoleic acid, suggesting that high levels of dietary linoleic acid may impair cognition⁽ Reference Lassek and Gaulin ^{241 )}. Individual consumption of dietary FA had an impact on cognitive measures in children⁽ Reference Lassek and Gaulin ^{242 )}, with n-3 FA being positively related to cognitive test scores in male and female children, while n-6 showed the reverse relationship. Higher scores in tests of neurodevelopment were found in infants fed formula with DHA than in infants fed formulas without DHA⁽ Reference Willatts, Forsyth and DiModugno ^{243 )}. DHA supplementation in young boys increased the prefrontal cortex activation during sustained attention⁽ Reference McNamara, Able and Jandacek ^{244 )}. These results are consistent with the hypothesis that dietary DHA is assimilated by the brain and has a positive influence on cognition. Autopsy data show lower DHA in brain of human infants fed formula rather than those who were breast-fed⁽ Reference Farquharson, Cockburn and Patrick ^{245 ,} Reference Makrides, Neumann and Byard ^{246 )}, which resembles the results obtained in pigs (Table 4).

In pigs, dietary FA significantly made an impact on the frontal cortex and striatum concentrations of neurotransmitters (for example, dopamine, serotonin)⁽ Reference de la Presa Owens and Innis ^{247 ,} Reference de la Presa Owens and Innis ^{248 )}. Another study showed fewer arm-entries in a maze in pigs receiving a low-PUFA diet compared with a high-PUFA diet, with the effect being probably dependent on central dopamine metabolism⁽ Reference Ng and Innis ^{249 )}. Epidemiological and clinical studies also suggest a relationship between dietary FA and altered functions of the nervous system, including neurocognitive disorders⁽ Reference Grosso, Galvano and Marventano ^{250 )}. Dietary n-3 FA could also protect against brain disorders on neurotransmission, neuroprotection and neurogenesis⁽ Reference Denis, Potier and Heberden ^{251 )}. However, data are lacking for pigs providing an opportunity to investigate the relationship between diet, brain activity and cognitive functions relevant to humans via in vivo imaging.

Peripheral neuromodulation to regulate eating behaviour in pigs

Vagal nerve stimulation (VNS) is a therapy for refractory epilepsy and psychiatric disorders⁽ Reference Hotujac and Kuzman ^{252 ,} Reference Vonck, De Herdt and Boon ^{253 )}, but it has also received attention as a way to modulate food intake. Animal models, including pigs, were used to investigate this question⁽ Reference Val-Laillet, Biraben and Randuineau ^{254 )}. Diaz-Guemes et al. ⁽ Reference Diaz-Guemes, Sanchez and Luis ^{255 )} found that VNS increased central nervous activity, but no effect was observed on feeding behaviour. In contrast, other authors demonstrated a decreased weight gain, decreased fat gain and plasma insulin-like growth factor I⁽ Reference Sobocki, Krolczyk and Herman ^{256 )}, or decreased food intake and specific activations in several brain areas associated with altered gastric myoelectric activity⁽ Reference Matyja, Thor and Sobocki ^{257 )} in growing pigs⁽ Reference Biraben, Guérin and Bobillier ^{258 )}. Another on-going study⁽ Reference Malbert, Guérin and Bobillier ^{259 )} showed VNS-induced metabolism differences in the brain reward circuit only 7d after VNS onset, meaning that quick central neuroplasticity phenomena can be induced by VNS, possibly modulating homeostatic and cognitive processes. In Göttingen minipigs fed a Western diet, VNS prevented further weight gain, decreased food intake and sweet cravings⁽ Reference Val-Laillet, Biraben and Randuineau ^{254 )}, proving the therapeutic potential of this strategy. Similarly, some studies assessing the impact of VNS on eating behaviours and weight in individuals with other psychiatric and neurological disorders showed significant modulation of food cravings and body weight⁽ Reference McClelland, Bozhilova and Campbell ^{260 ,} Reference Val-Laillet, Aarts and Weber ^{261 )}, but there are significant discrepancies between studies. Overall, functional imaging in pigs has the potential to help validate and optimise therapies before their application to human patients.

Central neuromodulation to regulate eating behaviour in pigs

Recent development suggests that the deep-brain stimulation (DBS), a procedure for the treatment of Parkinson’s or depression, might also be used to combat obesity⁽ Reference Halpern, Wolf and Bale ^{262 –} Reference Howland ^{265 )}. The minipig has emerged as an ideal model for basic and preclinical studies on DBS⁽ Reference Sorensen, Nielsen and Rosendal ^{266 )}. Hypothalamic DBS was validated in the Göttingen minipig⁽ Reference Bjarkam, Nielsen and Glud ^{267 ,} Reference Ettrup, Tornoe and Sorensen ^{268 )} and resulted in reduced weight gain⁽ Reference Melega, Lacan and Gorgulho ^{269 )}, as well as in behavioural and physiological changes that could be related to the activation of limbic and autonomic brain networks⁽ Reference Ettrup, Sorensen and Rodell ^{270 )}. These results with pigs are similar to those described in human studies (for reviewes, see McClelland et al. ⁽ Reference McClelland, Bozhilova and Campbell ^{260 )} and Val-Laillet et al. ⁽ Reference Val-Laillet, Aarts and Weber ^{261 )}). Shon et al. ⁽ Reference Shon, Lee and Goerss ^{271 )} showed that DBS of the subthalamic nuclei in pigs can stimulate striatal dopamine release, which is related to food motivation and obesity⁽ Reference Volkow, Wang and Baler ^{272 ,} Reference Narayanaswami, Thompson and Cassis ^{273 )}, whereas Sauleau et al. ⁽ Reference Sauleau, Lapouble and Val-Laillet ^{12 )} managed to modify food motivation and learning. Knight et al. ⁽ Reference Knight, Min and Hwang ^{274 )} demonstrated that DBS of the nucleus accumbens, a putative target to combat obesity⁽ Reference Halpern, Wolf and Bale ^{262 )}, modulated the activity of the prefrontal, cingulate and insular cortices, which are brain regions involved in eating behaviour. A similarly low metabolic activity of the prefrontal cortex was observed in obese humans⁽ Reference Le, Pannacciulli and Chen ^{235 –} Reference Volkow, Wang and Telang ^{237 )} and minipigs⁽ Reference Val-Laillet, Layec and Guerin ^{239 )}, an anomaly that was normalised via DBS of the cortex. The combination of DBS and MRI has been explored in pigs, in terms of image-guided brain navigation⁽ Reference White, Woolley and Bienemann ^{275 )}, network activation⁽ Reference Knight, Min and Hwang ^{274 ,} Reference Min, Hwang and Marsh ^{276 )} and safety⁽ Reference Shrivastava, Abosch and Hanson ^{277 –} Reference Gorny, Presti and Goerss ^{279 )}. These studies show the (mini) pig is a convenient model to study the impact of DBS on eating behaviour and nutritional diseases, and to test medical innovations in preclinical trials before being safely applied to humans.

New imaging approaches in pigs

In addition to the aforementioned neuromodulation studies, there are many innovative methods related to nutrition and brain activity used in humans that could potentially be investigated in pigs. Alstrup & Smith⁽ Reference Alstrup and Smith ^{280 )} reviewed 10 years of positron emission tomography findings on neuromolecular processes in the living porcine brain and listed all the validated brain radio ligands including several molecules of interest for nutrition studies. Other methods can be used for molecular imaging in pigs, such as the wireless instantaneous neurotransmitter concentration system which allows the measure of neurotransmitter release in specific brain areas⁽ Reference Agnesi, Tye and Bledsoe ^{281 ,} Reference Van Gompel, Chang and Goerss ^{282 )}. The non-invasive magnetoencephalography and electrocorticography⁽ Reference Bowyer, Okada and Papuashvili ^{283 )}, as well as the functional near-IR spectroscopy and cortical imaging have been successfully implemented in the pig model⁽ Reference Uga, Saito and Sano ^{284 )} and could be applied in the future to map brain responses to food and nutrient stimulations.