Leucine (and lysine) increased plasma levels of the satiety hormone cholecystokinin (CCK), and phenylalanine of the incretin glucagon-like peptide 1 (GLP-1) after oral gavages in pigs

Abstract

Excess dietary amino acids (AA) has been associated with reduced feed intake, increased satiation, and extended satiety in pigs. Recent ex vivo studies suggested that satiety peptide cholecystokinin (CCK) and insulinotropic glucagon-like peptide 1 (GLP-1), mediated the anorexigenic or insulinotropic effects of Lys, Glu, Phe, Ile, and Leu. However, the ex vivo model limitations require validation in vivo. The aim of the present study was to assess the effect of orally administered AA in vivo in pigs. It was hypothesized that oral Lys, Ile, and Leu have an anorexigenic effect via CCK, while Glu and Phe have an insulinotropic effect increasing circulating levels of GLP-1. Eight entire male pigs (Landrace × Large White) of 18.23 ± 1.06 kg of body weight were administered an oral gavage of water (control) or a 3 mmol/kg of Glu, Ile, Leu, Lys, Phe, or glucose (positive control for GLP-1 release) following an overnight fasting during 5 consecutive days using an incomplete latin square design. Blood samples were collected from the jugular vein before (-5 min, baseline value) and after the gavage (5, 15, 30, 60 and 90 min) to assess CCK and GLP-1 plasma levels. Pigs administered the oral gavage of Leu (P < 0.05), or Lys (P < 0.1) had increased levels of plasma CCK from 0 to 90 min post-gavage when compared to the control. A strong association (P < 0.001) was observed between GLP-1 plasma levels with Phe intake. The impact was significant starting 30 min post-gavage and was sustained until the end of the experiment (90 min post-gavage). Glucose administration increased GLP-1 early after the intake at the 5 min mark (P < 0.1). A positive correlation (P < 0.05, r = 0.89) driven by the impact of Phe at the 60 to 90 min post-gavage was identified between CCK and GLP-1 indicating feedback mechanisms between proximal and distal small intestine. In conclusion, oral gavages of Leu and Lys increased anorexigenic hormone CCK plasma levels in pigs. Phe caused a significant long-lasting increase in incretin GLP-1 plasma levels. Blood CCK and GLP-1 concentrations in Phe gavaged pigs were positively correlated indicating a potential feedback mechanism between proximal (CCK) and distal (GLP-1) small intestine. The present results are compatible with the known anorexigenic effects of excess dietary Leu and Lys, and the insulinotropic effect of Phe in pigs. These results demonstrate the relevance of accurate feed formulation practices particularly in post weaning pigs.

Keywords: amino acid; blood; cholecystokinin; glucagon-like peptide 1; oral gavage; pig.

Figure 1.

Study protocol for the collection of blood samples in orally gavaged pigs. At t = −5 min, baseline blood samples were collected. Immediately after the first blood, sampling light anesthesia with isoflurane was applied by mask ventilation. At t = 0, oral gavages with water (control) or 3 mmol/kg solutions of glucose (GLP-1 positive control), Glu, Ile, Leu, Lys, or Phe were administered with a 25 cm plastic extension tube. Following a 2 to 3 min period of observation after the gavage, animals were placed back into their corresponding pen. For the next 90 min after the gavage, blood samples were collected at t = 5, 15, 30, 60, and 90 min. At t = 90, pigs had ad libitum access to feed again.

Figure 2.

Changes in the blood kinetic of CCK following an oral gavage with AA or glucose in pigs. CCK plasma levels and AUC (0 to 90 min) in young pigs after an oral gavage with water (control) or 3 mmol/kg solutions of Lys (A), Phe (B), Leu (C), Ile (D), Glu (E), or glucose (F). The AUC was derived using a Best Linear Unbiased Estimator (BLUE) to estimate missing data. Baseline values are represented as time 0. Data (CCK plasma levels and AUC) are expressed as the mean + SEM (n = 4 or 5). Statistical differences of the AUC between AA treatments and glucose compared to the control are represented as: ^*P < 0.05.

Figure 3.

Changes in the blood kinetic of GLP-1 following an oral gavage with AA or glucose in pigs. GLP-1 plasma levels and AUC (0 to 90 min) in young pigs after an oral gavage with water (control) or 3 mmol/kg solutions of Lys (A), Phe (B), Leu (C), Ile (D), Glu (E), or glucose (F). The AUC was derived using a Best Linear Unbiased Estimator (BLUE) to estimate missing data. Baseline values are represented as time 0. Data (GLP-1 plasma levels and AUC) are expressed as the mean + SEM (n = 4 or 5). Statistical differences of the GLP-1 values between the AA treatment or Glucose and the baseline are represented as: ^#P < 0.05, and ^##P < 0.001.

Figure 4.

Relationship between plasma cholecystokinin (CCK) and glucagon-like peptide (GLP-1) levels following an oral gavage with amino acids (AA) or glucose in pigs. Relationship between CCK and GLP-1 plasma concentrations (expressed as AUC) for all treatments combined at 0 to 90 min (A) and 60 to 90 min (B) as well as for individual treatments [Lys (C), Phe (D), Leu, (E), Ile (F), Glu (G) and glucose (H)] at 60 to 90 min after an oral gavage in young pigs. The correlation analyses between CCK and GLP-1 data were performed by the Pearson’s correlation test. The P < 0.05 was set as the statistical significance.