OBJECTIVE: Growth differentiation factor 15 (GDF15), a stress related cytokine, was recently identified as a novel satiety signal acting via the GFRAL receptor located in the hindbrain. Bitter compounds are known to induce satiety via the release of glucagon-like peptide 1 (GLP-1) through activation of bitter taste receptors (TAS2Rs, 25 subtypes) on enteroendocrine cells in the gut. This study aimed to investigate whether and how bitter compounds induce a stress response in intestinal epithelial cells to affect GDF15 expression in patients with obesity, thereby facilitating satiety signaling from the gut.
METHODS: The acute effect of oral intake of the bitter-containing medication Plaquenil (hydroxychloroquine sulfate) on plasma GDF15 levels was evaluated in a placebo-controlled, double-blind, randomized, two-visit crossover study in healthy volunteers. Primary crypts isolated from the jejunal mucosa from patients with obesity were stimulated with vehicle or bitter compounds and the effect on GDF15 expression was evaluated using RT-qPCR or ELISA. Immunofluorescence colocalization studies were performed between GDF15, epithelial cell type markers and TAS2Rs. The role of TAS2Rs was tested by 1) pretreatment with a TAS2R antagonist, GIV3727; 2) determining TAS2R4/43 polymorphisms that affect taste sensitivity to TAS2R4/43 agonists.
RESULTS: Acute intake of hydroxychloroquine sulfate increased GDF15 plasma levels, which correlated with reduced hunger scores and plasma ghrelin levels in healthy volunteers. This effect was mimicked in primary jejunal cultures from patients with obesity. GDF15 was expressed in enteroendocrine and goblet cells with higher expression levels in patients with obesity. Various bitter-tasting compounds (medicinal, plant extracts, bacterial) either increased or decreased GDF15 expression, with some also affecting GLP-1. The effect was mediated by specific intestinal TAS2R subtypes and the unfolded protein response pathway. The bitter-induced effect on GDF15/GLP-1 expression was influenced by the existence of TAS2R4 amino acid polymorphisms and TAS2R43 deletion polymorphisms that may predict patient\'s therapeutic responsiveness. However, the effect of the bitter-tasting antibiotic azithromycin on GDF15 release was mediated via the motilin receptor, possibly explaining some of its aversive side effects.
CONCLUSIONS: Bitter chemosensory and pharmacological receptors regulate the release of GDF15 from human gut epithelial cells and represent potential targets for modulating metabolic disorders or cachexia.

Motilin, produced in endocrine cells in the mucosa of the upper intestine, is an important regulator of gastrointestinal (GI) motility and mediates the phase III of interdigestive migrating motor complex (MMC) in the stomach of humans, dogs and house musk shrews through the specific motilin receptor (MLN-R). Motilin-induced MMC contributes to the maintenance of normal GI functions and transmits a hunger signal from the stomach to the brain. Motilin has been identified in various mammals, but the physiological roles of motilin in regulating GI motility in these mammals are well not understood due to inconsistencies between studies conducted on different species using a range of experimental conditions. Motilin orthologs have been identified in non-mammalian vertebrates, and the sequence of avian motilin is relatively close to that of mammals, but reptile, amphibian and fish motilins show distinctive different sequences. The MLN-R has also been identified in mammals and non-mammalian vertebrates, and can be divided into two main groups: mammal/bird/reptile/amphibian clade and fish clade. Almost 50 years have passed since discovery of motilin, here we reviewed the structure, distribution, receptor and the GI motility regulatory function of motilin in vertebrates from fish to mammals.

Motilin\'s role in the regulation of vascular tone and hemodynamic besides gastrointestinal motility is concerned. This study aimed to investigate the expression of motilin receptors in gastrointestinal arteries and motilin-induced relaxation.
The expression of motilin receptors in the left gastric artery (LGA), superior mesenteric artery (SMA), and inferior mesenteric artery (IMA) of adult dogs (1.5-5 years old) were analyzed by immunochemistry, RT-PCR, and western blotting. Motilin\'s effects on the gastrointestinal arteries were evaluated in a multi-wire myograph system.
Immunohistochemical staining showed that motilin receptor was expressed on the membranes of endothelial cells with the fluorescence intensity LGA > SMA > IMA (P < 0.01). The motilin receptor\'s mRNA and protein expression levels shared the same distribution patterns as it in fluorescence intensity (P < 0.01). In isolated LGA preparations precontracted with U46619 (a thromboxaneA2 analog), motilin induced a concentration-dependent relaxation, and the EC50 was 8.8 × 10-8 ± 0.9 × 10-8 M. Motilin-induced relaxation on the three arteries also shared the same pattern as it in fluorescence intensity (P < 0.01) and inhibited by denuded-endothelium and GM-109 (a motilin receptor antagonist) but not by atropine (a muscarinic receptor antagonist).
Motilin receptors are expressed differentially on the membranes of endothelial cells in dog gastrointestinal arteries with a significantly high expression in the LGA. Motilin-induced relaxation is endothelium- and motilin receptor-dependent. The motilin receptor expressed on the endothelial cell membrane of the LGA is the molecular basis for motilin regulating gastric blood flow under physiological conditions in dogs.

1. A previous whole-genome association analysis has identified the motilin receptor gene (MLNR), which regulates gastrointestinal motility and gastric emptying, as a candidate gene related to chicken growth.2. MLNR mRNA was expressed in all tissues tested, and the expression level in digestive tissues was greater than in other tissues. Expression levels in the pancreas, duodenum and glandular stomach at day old and one, two and three weeks of age indicated a possible correlation with the digestive system. This suggested that the MLNR gene plays a central role in gastrointestinal tract function and affects the growth and development of chickens. Moreover, there was a significant difference in expression in the glandular stomach tissue between Ross 308 and Gushi chickens at six weeks of age.3. Re-sequencing revealed an 86-bp insertion/deletion polymorphism in the downstream region of the MLNR gene. The mutation locus was genotyped in 2,261 individuals from nine different chicken breeds. MLNR expression levels in the glandular stomach of chickens with DD genotypes were greater than those in chickens with the ID and II genotypes. The DD genotype was the most dominant genotype in commercial broiler\'s (Ross 308 and Arbor Acres broilers), and the D allele frequency in these breeds exceeded 91%. The deletion mutation tended towards fixation in commercial broilers.4. Association with growth and carcass traits analysed in a Gushi-Anka F2 intercrossed population, showed that the DD genotype was significantly associated with the greatest growth and carcass trait values, whereas values associated with the II genotype were the lowest in the F2 reciprocal cross chickens.5. The results suggest that the mutation is strongly associated with growth related traits and it is likely to be useful for marker-assisted selection of chickens.

Motilin (MLN), a 22-amino-acid peptide hormone, is generally present in the mucosa of the upper gastrointestinal (GI) tract, mainly the duodenum of mammals, and it regulates GI motility, especially that related to interdigestive migrating contraction. However, MLN and its receptor are absent in mice and rats, and MLN does not cause any mechanical responses in the rat and mouse GI tracts. The guinea-pig is also a rodent, but expression of the MLN gene in the guinea-pig has been reported. In the present study, two guinea-pig MLNs, FIPIFTYSELRRTQEREQNKGL found in the Ensemble Genome Database (gpMLN-1) and FVPIFTYSELRRTQEREQNKRL reported by Xu et al. (2001) (gpMLN-2), were synthesized, and their biological activities were evaluated in the rabbit duodenum and guinea-pig GI tract in vitro. Both gpMLNs showed contractile activity in longitudinal muscle strips of the rabbit duodenum. The EC50 values of gpMLN-1 and gpMLN-2 were slightly higher than that of human MLN (hMLN), but the maximum contractions were as same as that of hMLN. Treatment with GM109 and hMLN-induced receptor desensitization decreased the contractile activity of both gpMLNs, indicating that the two gpMLN candidates are able to activate the MLN receptor (MLN-R) of the rabbit duodenum. In guinea-pig GI preparations, hMLN and gpMLNs did not show any mechanical responses in circular muscle strips from the gastric antrum or in longitudinal strips of the duodenum, ileum and colon although acetylcholine and 1,1-dimethyl-4-phenylpiperazinium (DMPP) caused definite mechanical responses. The DMPP-induced neural responses in the gastric circular muscle and ileal longitudinal muscles were not modified by gpMLN-1. Even in the gastric and ileal strips with intact mucosa, no mechanical responses were seen with either of the gpMLNs. Furthermore, RT-PCR using various primer sets failed to amplify the gpMLN-2 mRNA. In conclusion, gpMLNs including one that was already reported and the other that was newly found in a database were effective to the rabbit MLN-R, whereas they did not cause any contractions or modification of neural responses in the guinea-pig GI tract, indicating that the MLN system is vestigial and not functional in regulation of GI motility in the guinea-pig as well as in other rodents such as rats and mice.

Motilin (MOT), a 22-amino-acid peptide hormone produced in the duodenal mucosa, stimulates gastrointestinal motility in mammals and birds, and it is a mediator of interdigestive motor complexes. Recently, expression of MOT-like peptide (MOTLP) and its receptor mRNAs was identified in zebrafish. The aim of the present study was to determine whether the zebrafish MOTLP (zfMOTLP, HIAFFSPKEMRELREKE) affects zebrafish gastrointestinal motility, with comparison to the effect of human MOT, in which five amino acids are identical to zfMOTLP at positions 5, 9, 15, 16, and 17. zfMOTLP caused small contractions of the rabbit duodenum and chicken ileum but, the sensitivity was about 3000-times lower than that of human MOT. zfMOTLP-induced contraction in the rabbit duodenum was decreased by pretreatment of the MOT receptor antagonist GM109, indicating that zfMOTLP could bind to the MOT receptor. zfMOTLP (3-100nM) increased the intracellular Ca2+ concentration in zfMOT receptor-expressing HEK293 cells, but human MOT did not cause responses even at 100nM. In in vitro study using isolated zebrafish gastrointestinal strips, zfMOTLP caused only small contractions even at high doses (1-10μM). zfMOT receptor mRNA is detected in the gastrointestinal tract and brain to almost the same extent, and the expression level (40-70 copies/100ng total RNA) is much lower than that in the chicken gastrointestinal tract. These results suggest that the MOTLP/MOT receptor system is present in zebrafish, but its physiological role for regulation of gastrointestinal motility might be not significant due to the weak contractile activity and low expression level of the receptor.

OBJECTIVE: To study the neural mechanism by which electroacupuncture (EA) at RN12 (Zhongwan) and BL21 (Weishu) regulates gastric motility.
METHODS: One hundred and forty-four adult Sprague Dawley rats were studied in four separate experiments. Intragastric pressure was measured using custom-made rubber balloons, and extracellular neuron firing activity, which is sensitive to gastric distention in the dorsal vagal complex (DVC), was recorded by an electrophysiological technique. The expression levels of c-fos, motilin (MTL) and gastrin (GAS) in the paraventricular hypothalamic nucleus (PVN) were assayed by immunohistochemistry, and the expression levels of motilin receptor (MTL-R) and gastrin receptor (GAS-R) in both the PVN and the gastric antrum were assayed by western blotting.
RESULTS: EA at RN12 + BL21 (gastric Shu and Mu points), BL21 (gastric Back-Shu point), RN12 (gastric Front-Mu point), resulted in increased neuron-activating frequency in the DVC (2.08 ± 0.050, 1.17 ± 0.023, 1.55 ± 0.079 vs 0.75 ± 0.046, P < 0.001) compared with a model group. The expression of c-fos (36.24 ± 1.67, 29.41 ± 2.55, 31.79 ± 3.00 vs 5.73 ± 2.18, P < 0.001), MTL (22.48 ± 2.66, 20.76 ± 2.41, 19.17 ± 1.71 vs 11.68 ± 2.52, P < 0.001), GAS (24.99 ± 2.95, 21.69 ± 3.24, 23.03 ± 3.09 vs 12.53 ± 2.15, P < 0.001), MTL-R (1.39 ± 0.05, 1.22 ± 0.05, 1.17 ± 0.12 vs 0.84 ± 0.06, P < 0.001), and GAS-R (1.07 ± 0.07, 0.91 ± 0.06, 0.78 ± 0.05 vs 0.45 ± 0.04, P < 0.001) increased in the PVN after EA compared with the model group. The expression of MTL-R (1.46 ± 0.14, 1.26 ± 0.11, 0.99 ± 0.07 vs 0.65 ± 0.03, P < 0.001), and GAS-R (1.63 ± 0.11, 1.26 ± 0.16, 1.13 ± 0.02 vs 0.80 ± 0.11, P < 0.001) increased in the gastric antrum after EA compared with the model group. Damaging the PVN resulted in reduced intragastric pressure (13.67 ± 3.72 vs 4.27 ± 1.48, P < 0.001). These data demonstrate that the signals induced by EA stimulation of acupoints RN12 and BL21 are detectable in the DVC and the PVN, and increase the levels of gastrointestinal hormones and their receptors in the PVN and gastric antrum to regulate gastric motility.
CONCLUSIONS: EA at RN12 and BL21 regulates gastric motility, which may be achieved through the PVN-DVC-vagus-gastric neural pathway.

We previously identified quantitative trait loci (QTL) for body weight and average daily gain in a common region between ADL0198 (chr 1: 171.7 Mb) and ABR0287 (chr 1: 173.4 Mb) on chicken chromosome 1 in an F2 resource population produced by crossing low- and high-growth lines of the Hinai-dori breed. Motilin receptor (MLNR) is a candidate gene affecting growth traits in the region. In this study, we genotyped polymorphisms of the MLNR gene and investigated its association with growth traits in a Hinai-dori F2 intercross population. All the exons of the MLNR gene in the parental population were subjected to PCR amplification, nucleotide sequenced and haplotypes identified. To distinguish resultant diplotype individuals in the F2 population, a mismatch amplification mutation assay was performed. Three haplotypes (Haplotypes 1-3) were accordingly identified. Six genotypes produced by the combination of three haplotypes (Haplotype 1, 2, and 3) were examined in order to identify associations between MLNR haplotypes and growth traits. The data showed that Haplotype 1 was superior to Haplotype 2 and 3 in body weight at 10 and 14 weeks of age, average daily gain between 4 and 10 weeks, 10 and 14 weeks, and 0 and 14 weeks of age in female in F2 females. It was concluded that MLNR is a useful marker of growth traits and could be used to develop strategies for improving growth traits in the Hinai-dori breed.

OBJECTIVE: To investigate the effects of intrathecal morphine and fentanyl combined with low-dose naloxone on the expression of motilin and its receptor in a rat model of postoperative pain.
METHODS: An intrathecal catheter was implanted, and saline, opioids (morphine and fentanyl) and naloxone were intrathecally administered 7 days later. An incisional pain model was established to induce pain behaviors in rats by unilateral plantar incision. Thermal hyperalgesia and mechanical allodynia were measured by using a radiant heat and electronic Von Frey filament, respectively. The expression of motilin in the hippocampus, stomach, duodenum, and plasma was determined by ELISA; and the expression of motilin receptor in the hippocampus was detected by Western blot assay.
RESULTS: Motilin and its receptor were detected in the hippocampus. Acute incisional pain increased the motilin expression in the hippocampus and duodenum, while decreasing its expression in the gastric body and plasma. Postoperative analgesia with morphine+fentanyl upregulated the expression of motilin in the hippocampus; however, motilin was downregulated in peripheral sites. Naloxone at 1 ng/kg restored motilin to baseline levels. Acute pain, morphine+fentanyl, and naloxone all induced the expression of motilin receptor in the hippocampus.
CONCLUSIONS: Acute pain, postoperative analgesia with opioids, and naloxone significantly impacted the expression of hippocampal and peripheral motilin. Variation trends in all sites were not identical. Intrathecal injection of low-dose naloxone upregulated paw withdrawal thermal latency and enhanced the analgesic effects of opioids. The findings presented here provide a new basis for central and peripheral regulations in GI motility, clinical postoperative analgesia, and management of analgesic complications.