Glutamate promotes nucleotide synthesis in the gut and improves availability of soybean meal feed in rainbow trout

Abstract

Glutamate (Glu) plays various roles directly or through conversions to other amino acids in intracellular metabolisms such as energy source for enterocytes and precursor for nucleic acids. In this study, we examined the effect of single and chronic oral administration of Glu on cell proliferation in intestine and growth in rainbow trout fed soybean meal (SBM) based diet. In the single dose study, 30, 120 and 360 min after oral administration of 50 and 500 mg/kg Glu, the blood and intestine tissues were collected for amino acid concentration and gene expression analysis. Cell-proliferation was detected 24 h after administration using bromo-deoxy uridine (BrdU) in intestine. In the chronic experiment, fish were fed SBM-based diet added 1 and 2 % of Glu for 8 weeks. Final body weight, plasma amino acid concentrations, gene expression and cell-proliferation in the intestine were analyzed. The expressions of some nucleic acid-synthesis related genes were significantly increased 30 min after administration of 50 mg/kg of Glu. After 8 weeks of feeding, the fish fed SBM-based diet showed significantly lower body weight and microvillus thickness in proximal intestine. Supplementation of 2 % of Glu in the SBM-based feed improved both of them. Though it was not significant difference, Glu tended to increase cell-proliferation in the proximal intestine dose-dependently in both single and chronic administration. Our experiment indicates that Glu has positive effect on rainbow trout fed SBM-based feed by reforming proximal intestine through altering cell-proliferation.

Keywords: Amino acids; Feed ingredient; Fishmeal alternatives.

Fig. 1

Alterations of gene expressions in proximal intestine by oral Glu intake. Relative expression of genes related to nucleic acid synthesis in proximal intestine. a cps2: carbamoyl-phosphate synthetase, b ppat: phosphoribosyl pyrophosphate amidotransferase, c pfas: phosphoribosylformylglycinamidine synthase. The tissues were collected before (initial), 30, 120, and 360 min after administration of 50 or 500 mg/kg Glu. The expression of the genes were estimated by quantitative RT-PCR and normalized by arbp. All values are expressed as the mean ± SEM (n = 6). *p < 0.05 as compared to initial group

Fig. 2

Alterations of gene expressions in distal intestine by oral Glu intake. Relative expression of genes related to nucleic acid synthesis in distal intestine. a cps2: carbamoyl-phosphate synthetase, b ppat: phosphoribosyl pyrophosphate amidotransferase, c pfas: phosphoribosylformylglycinamidine synthase. The tissues were collected before (initial), 30, 120, and 360 min after administration of 50, 500 mg/kg Glu p.o. The expression of the genes were estimated by quantitative RT-PCR and normalized by arbp. All values are expressed as the mean ± SEM (n = 6)

Fig. 3

Effects of Glu intake on cell proliferation in the intestine. The number of BrdU positive cells per microvillus in a proximal intestine, b distal intestine. The tissues were collected 24 h after 0, 50, 500 mg/kg Glu (p.o.) and BrdU (i.p.) administration. Data are shown as the average of five villi from each fish. *p < 0.05 as compared to 0 mg/kg group

Fig. 4

Weekly feed intake. The experimental diets were fed by hand to apparent satiation twice a day, 6 days a week for 8 weeks. Data are shown as week total intake per fish

Fig. 5

Alterations of gene expressions in proximal intestine by 8 weeks of Glu supplementation. Relative expression of genes related to nucleic acid synthesis in proximal intestine. A cps2: carbamoyl-phosphate synthetase, A ppat: phosphoribosyl pyrophosphate amidotransferase, C pfas: phosphoribosylformylglycinamidine synthase. The fish were challenged to 8 weeks of experiment. The tissues were collected after the 48 h-fasting. The expression of the genes were estimated by quantitative RT-PCR and normalized by arbp. All values are expressed as the mean ± SEM (n = 8). Different letters indicate significant difference between groups (p < 0.05)

Fig. 6

Alterations of gene expressions in distal intestine by 8 weeks of Glu supplementation in feed. Relative expression of genes related to nucleic acid synthesis in distal intestine. A cps2: carbamoyl-phosphate synthetase, B ppat: phosphoribosyl pyrophosphate amidotransferase, C pfas: phosphoribosylformylglycinamidine synthase. The fishes were challenged to 8 weeks of experiment. The tissues were collected after the 48 h-fasting. The expression of the genes were estimated by quantitative RT-PCR and normalized by arbp. All values are expressed as the mean ± SEM (n = 8). Different letters indicate significant difference between groups (p < 0.05)

Fig. 7

Morphological alteration in distal intestine by SBM feed. Transverse section of distal intestine mucosal fold a FM diet group, b SBM diet group. The fish were challenged to 8 weeks of experiment and fasted for the last 48 h of experiment. The central panel is an extended image of the area surrounded by a broken line in a. dv digestive vacuole, lp lamina propria, pv pinocytotic vacuoles, sm submucosa. Scale bar 100 μm

Fig. 8

Effects of Glu intake on microvilli thickness in the proximal intestine. The thickness of microvillus of proximal intestine. The fish were fasted for the last 48 h of the experiment. All values are expressed as the mean ± SEM (n = 8). Different letters indicate significant difference between groups (p < 0.05)

Fig. 9

Effects of Glu intake on cell proliferation in the intestine. The number of BrdU positive cells per microvillus of A anterior intestine, B posterior intestine. The fish were challenged to 8 weeks of experiment. The fish were fasted for the last 48 h of the experiment. BrdU was administered i.p. 24 h before the sacrifice. All values are expressed as the mean ± SEM (n = 8). Different letters indicate significant difference between groups (p < 0.05)