AMPK-mTOR pathway is involved in glucose-modulated amino acid sensing and utilization in the mammary glands of lactating goats

Abstract

Background: The local supply of energy-yielding nutrients such as glucose seems to affect the synthesis of milk components in the mammary gland (MG). Thus, our study was conducted to investigate the effects of locally available MG glucose supply (LMGS) on amino acid (AA) sensing and utilization in the MG of lactating dairy goats. Six dosages of glucose (0, 20, 40, 60, 80, and 100 g/d) were infused into the MG through the external pudendal artery to investigate the dose-dependent changes in mammary AA uptake and utilization (Exp.1) and the changes in mRNA and protein expression of the AMPK-mTOR pathway (Expt.2).

Results: In Exp.1, total milk AA concentration was highest when goats were infused with 60 g/d glucose, but lower when goats were infused with 0 and 100 g/d glucose. Increasing LMGS quadratically changed the percentages of α_S2-casein and α-lactalbumin in milk protein, which increased with infusions from 0 to 60 g/d glucose and then decreased with infusions between 60 and 100 g/d glucose. The LMGS changed the AA availability and intramammary gland AA utilization, as reflected by the mammary AA flux indexes. In Exp.2, the mRNA expression of LALBA in the MG increased quadratically with increasing LMGS, with the highest expression at dose of 60 g/d glucose. A high glucose dosage (100 g/d) activated the general control nonderepressible 2 kinase, an intracellular sensor of AA status, resulting in a reduced total milk AA concentration.

Conclusions: Our new findings suggest that the lactating MG in dairy goats may be affected by LMGS through regulation of the AA sensory pathway, AA utilization and protein synthesis, all being driven by the AMPK-mTOR pathway.

Keywords: AMPK signaling; Amino acid profile; Glucose supply; Mammary gland; Milk protein component.

Fig. 1

Relative mRNA expression of genes encoding the main milk proteins in the mammary gland of lactating goats infused with low (LDG, 0 g/d), middle (MDG, 60 g/d) or high (HDG, 100 g/d) glucose dosages through the external pudendal artery in Exp.2. Error bars represent pooled SEM. ^a,b,cValues without a common letter for a gene differ (P ≤ 0.05). A BLG = β-lactoglobulin; B CSN1S2 = α_S2-casein; C CSN2 = β-casein; D CSN3 = κ-casein; E LALBA = α-lactalbumin

Fig. 2

Relative mRNA expression of genes involved in the AMPK-mTOR pathway in lactating mammary glands of dairy goats locally treated with low (LDG, 0 g/d), middle (MDG, 60 g/d) or high (HDG, 100 g/d) glucose dosages in Exp.2. Error bars represent the pooled SEM. ^a,b,cValues without a common letter for a gene differ (P ≤ 0.05). A EEF1A1 = eukaryotic translation elongation factor 1 α 1; B EEF2 = eukaryotic translation elongation factor 2; C EIF4EBP1 = eukaryotic translation initiation factor 4E binding protein 1; D MTOR = mechanistic target of rapamycin; E PRKAA1 = protein kinase AMP-activated catalytic subunit alpha 1; F RPS6KB1 = ribosomal protein S6 kinase β-1

Fig. 3

Protein abundance and phosphorylation state of AMPK and mTOR in lactating mammary glands of dairy goats locally treated with low (LDG, 0 g/d), middle (MDG, 60 g/d) or high (HDG, 100 g/d) glucose dosages in Exp.2. Error bars represent the pooled SEM. ^a,b,cValues without a common letter differ (P ≤ 0.05). A AMPK = AMP-activated protein kinase; B p-AMPK = phosphorylated AMP-activated protein kinase; C p-AMPK/AMPK = the ratio of phosphorylated AMP-activated protein kinase to AMP-activated protein kinase; D mTOR = mechanistic target of rapamycin; E p-mTOR = phosphorylated mechanistic target of rapamycin; F p-mTOR/mTOR = the ratio of phosphorylated mechanistic target of rapamycin to phosphorylated mechanistic target of rapamycin

Fig. 4

Relative mRNA expression of genes involved in amino acid response pathways in lactating mammary glands of dairy goats locally treated with low (LDG, 0 g/d), middle (MDG, 60 g/d) or high (HDG, 100 g/d) glucose dosages in Exp.2. Error bars represent pooled SEM. ^a,b,cValues without a common letter were significantly different (P ≤ 0.05). A ATF4 = activating transcription factor 4; B GCN2 = general control nonderepressible 2; C NUPR1 = stress protein P8