Galanin enhances systemic glucose metabolism through enteric Nitric Oxide Synthase-expressed neurons

Abstract

Objective: Decreasing duodenal contraction is now considered as a major focus for the treatment of type 2 diabetes. Therefore, identifying bioactive molecules able to target the enteric nervous system, which controls the motility of intestinal smooth muscle cells, represents a new therapeutic avenue. For this reason, we chose to study the impact of oral galanin on this system in diabetic mice.

Methods: Enteric neurotransmission, duodenal contraction, glucose absorption, modification of gut-brain axis, and glucose metabolism (glucose tolerance, insulinemia, glucose entry in tissue, hepatic glucose metabolism) were assessed.

Results: We show that galanin, a neuropeptide expressed in the small intestine, decreases duodenal contraction by stimulating nitric oxide release from enteric neurons. This is associated with modification of hypothalamic nitric oxide release that favors glucose uptake in metabolic tissues such as skeletal muscle, liver, and adipose tissue. Oral chronic gavage with galanin in diabetic mice increases insulin sensitivity, which is associated with an improvement of several metabolic parameters such as glucose tolerance, fasting blood glucose, and insulin.

Conclusion: Here, we demonstrate that oral galanin administration improves glucose homeostasis via the enteric nervous system and could be considered a therapeutic potential for the treatment of T2D.

Keywords: Diabetes; Enteric nervous system; Galanin.

Graphical abstract

Figure 1

Galanin stimulates duodenal NO release, decreases duodenal contractions to decrease glucose absorption. (A)Ex vivo measurement of duodenal nitric oxide (NO) release amplitude during 5 min in response to Krebs–Ringer (Control), galanin 100 nM and galanin 100 nM plus galantide 100 nM n = 6–12 per group. a, p < 0.01 vs Control; b, p < 0.05 vs galanin + galantide. (B)Ex vivo measurement of duodenal mechanical contraction amplitude during 10 min in response to Krebs–Ringer solution (Control), galanin 100 nM and galanin 100 nM plus galantide 100 nM n = 5 per group. **p < 0.01 vs other groups. (C)In vivo telemetric measurement of duodenal electrical activity during 10 min in response to water (Control), galanin 100 nM and galanin 100 nM plus galantide 100 nM, n = 5 per group. ***p < 0,001 vs other groups. (D)Ex vivo glucose absorption in duodenal everted sacs in response to Krebs–Ringer (Control) or galanin 100 nM n = 8 per group. *p < 0.05 vs Control.

Figure 2

Intestinal galanin modulates the gut-brain axis to control glucose utilization in tissue. (A)In vivo effect of intragastric perfusion of water (Control), galanin 100 nM and galanin 100 nM + galantide 100 nM on nitric oxide (NO) hypothalamic release amplitude. n = 4–7 per group. *p < 0.05 vs other groups. (B)In vivo measurement of glucose entry in muscle, liver and subcutaneous adipose tissue in response to oral gavage of radiolabeled glucose in combination with water (Control) or galanin 100 nM n = 5 per group. *p < 0.05 vs Control.

Figure 3

Oral galanin treatment decreases the duodenal hyper-contractility of diabetic mice and improves diabetic state. (A)Ex vivo measurement of duodenal mechanical contraction amplitude in response to Krebs–Ringer (Control) or galanin 100 nM in high-fat diet (HFD) mice. n = 5 per group. *p < 0.05 vs HFD Control. (B)In vivo measurement of duodenal mechanical contraction amplitude in response to an oral administration of water (HFD Control) or galanin 100 nM during one week. n = 5–8 per group. *p < 0.05 vs HFD Control. (C) Effects of an oral administration of water (HFD Control) or galanin 100 nM during one week on fasted glycemia in HFD mice. n = 12–14 per group. **p < 0.01 vs HFD Control. (D) Oral glucose tolerance test (OGTT) in 6 h fasted HFD mice, after an oral administration of water (HFD Control) or galanin 100 nM during one week. n = 12–14 per group. The adjacent graph represents the average area under the curve (AUC) ***p < 0.001 vs HFD Control. (E) OGTT-associated plasma insulin and glucagon 30 min before and 15 min after oral load of glucose after an oral administration of water (HFD Control) or galanin 100 nM during one week. n = 5 per group. *p < 0.05, **p < 0.01 vs HFD Control. (F) OGTT-associated insulin resistance index (HOMA-IR) in 6 h fasted HFD mice after an oral administration of water (HFD Control) or galanin 100 nM during one week. n = 5 per group. ***p < 0.001 vs HFD Control. (G) Relative expression of endothelial Nitric Oxide Synthase (eNOS) mRNA in hypothalamus of HFD mice after an oral administration of water (HFD Control) or galanin 100 nM during one week. n = 4–5 per group.

Figure 4

Oral galanin improves insulin sensitivity via an AMPK signaling. (A) Hepatic glycogen content of fed mice after an oral administration of water (HFD Control) or galanin 100 nM during one week. n = 4–10 per group. *p < 0.05 vs HFD Control. (B) Relative expression of glucose-6-phosphatase (G6pase) mRNA in liver of HFD mice after an oral administration of water (HFD Control) or galanin 100 nM during one week. n = 4–5 per group. (C) Relative expression of Phosphoenolpyruvate carboxykinase (Pepck) mRNA in liver of HFD mice after an oral administration of water (HFD Control) or galanin 100 nM during one week. n = 4–5 per group. (D)Vastus lateralis muscle glycogen content of fed HFD mice after an oral administration of water (HFD Control) or galanin 100 nM during one week. n = 4–10 per group. *p < 0.05 vs HFD Control. (E) Relative expression of glucose transporter type 4 (Glut-4) mRNA in vastus lateralis muscle of HFD mice after an oral administration of water (HFD Control) or galanin 100 nM during one week. n = 5 per group. (F)Vastus lateralis muscle AMPK and Akt protein expression of HFD mice after an oral administration of water (HFD Control) or galanin 100 nM during one week and the relative quantification. n = 4–5 per group. *p < 0.05 vs Control.