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Inhibition of glycine transporter-1 in the dorsal vagal complex improves metabolic homeostasis in diabetes and obesity

View Article: PubMed Central - PubMed

ABSTRACT

Impaired glucose homeostasis and energy balance are integral to the pathophysiology of diabetes and obesity. Here we show that administration of a glycine transporter 1 (GlyT1) inhibitor, or molecular GlyT1 knockdown, in the dorsal vagal complex (DVC) suppresses glucose production, increases glucose tolerance and reduces food intake and body weight gain in healthy, obese and diabetic rats. These findings provide proof of concept that GlyT1 inhibition in the brain improves glucose and energy homeostasis. Considering the clinical safety and efficacy of GlyT1 inhibitors in raising glycine levels in clinical trials for schizophrenia, we propose that GlyT1 inhibitors have the potential to be repurposed as a treatment of both obesity and diabetes.

No MeSH data available.


Molecular inhibition of DVC GlyT1 regulates glucose homeostasis in healthy rats.(a) Representative western blots and protein levels of plasma membrane GlyT1 (55, 70 and 90 kDa isoforms) normalized to insulin receptor (IR) in DVC wedges of rats 13-day post DVC lentiviral (LV) injection of GlyT1 shRNA (black bars, n=14) or a mismatch sequence (MM; white bars, n=11) as a control. *P<0.01, **P<0.001 determined by t-test. (b–d) Representative western blots and protein levels of plasma membrane GlyT1 (70 and/or 90 kDa isoforms) normalized to IR in sp5, Sp5C, Sp5I (L), sp5, Sp5C, Sp5I (R) and py wedges of rats 13-day post DVC LV injection of GlyT1 shRNA (black bars, n=5) or MM (white bars, n=5). (e) Glucose infusion rates and (f) glucose production during clamps in rats injected with LV-MM (n=7), LV-GlyT1 shRNA (n=7)or LV-GlyT1 shRNA with DVC MK801 infusion (n=6). (e: *P<0.006; f: *P<0.003 versus LV-MM control and LV-GlyT1 shRNA+MK801 determined by ANOVA and Dunnett's post hoc test). Data are shown as the mean+s.e.m.
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f2: Molecular inhibition of DVC GlyT1 regulates glucose homeostasis in healthy rats.(a) Representative western blots and protein levels of plasma membrane GlyT1 (55, 70 and 90 kDa isoforms) normalized to insulin receptor (IR) in DVC wedges of rats 13-day post DVC lentiviral (LV) injection of GlyT1 shRNA (black bars, n=14) or a mismatch sequence (MM; white bars, n=11) as a control. *P<0.01, **P<0.001 determined by t-test. (b–d) Representative western blots and protein levels of plasma membrane GlyT1 (70 and/or 90 kDa isoforms) normalized to IR in sp5, Sp5C, Sp5I (L), sp5, Sp5C, Sp5I (R) and py wedges of rats 13-day post DVC LV injection of GlyT1 shRNA (black bars, n=5) or MM (white bars, n=5). (e) Glucose infusion rates and (f) glucose production during clamps in rats injected with LV-MM (n=7), LV-GlyT1 shRNA (n=7)or LV-GlyT1 shRNA with DVC MK801 infusion (n=6). (e: *P<0.006; f: *P<0.003 versus LV-MM control and LV-GlyT1 shRNA+MK801 determined by ANOVA and Dunnett's post hoc test). Data are shown as the mean+s.e.m.

Mentions: We alternatively tested the gluco-regulatory role of hindbrain GlyT1 inhibition via the targeted molecular knockdown of GlyT1 within the DVC. We first confirmed that lentiviral injection of GlyT1 shRNA (LV-GlyT1 shRNA) into the DVC selectively reduces the expression of both the 70- and 90-kDa isoforms of GlyT1 in plasma membrane fractions of only the DVC tissue compared with lentiviral injection of mismatch sequence (LV-MM), but not in the two adjacent left and right lateral regions of the DVC containing the Spinal trigeminal track (sp5), Spinal 5nu caudal part (Sp5C) and Spinal 5nu interpolar (Sp5I), and the region inferior to the DVC containing the pyramidal tract (py) of the same rats (Fig. 2a–d, Supplementary Fig. 3i–v). The dominant band at the molecular weight of 70–75 kDa corresponds to GlyT1a and b isoforms and the weaker band at90–100 kDa corresponds to GlyT1c isoform found in the rat brain as described34. The 90–100 kDa band is not detected in the sp5, Sp5C, Sp5I (right) and py regions (Fig. 2c,d). The immunoblot also reveals a strong band at 55 kDa (Fig. 2a), which is consistent with the occurrence of the partially glycosylated form of GlyT1 in the 55–60 kDa range as indicated353637. However, the 55 kDa GlyT1 band in the DVC of LV-GlyT1 shRNA versus LV-MM injected rats is not significantly different as compared with the effect on 70 kDa and 90 kDa bands (Fig. 2a). Nonetheless, the specific metabolic role of various forms of GlyT1 in the brain warrants future investigation.


Inhibition of glycine transporter-1 in the dorsal vagal complex improves metabolic homeostasis in diabetes and obesity
Molecular inhibition of DVC GlyT1 regulates glucose homeostasis in healthy rats.(a) Representative western blots and protein levels of plasma membrane GlyT1 (55, 70 and 90 kDa isoforms) normalized to insulin receptor (IR) in DVC wedges of rats 13-day post DVC lentiviral (LV) injection of GlyT1 shRNA (black bars, n=14) or a mismatch sequence (MM; white bars, n=11) as a control. *P<0.01, **P<0.001 determined by t-test. (b–d) Representative western blots and protein levels of plasma membrane GlyT1 (70 and/or 90 kDa isoforms) normalized to IR in sp5, Sp5C, Sp5I (L), sp5, Sp5C, Sp5I (R) and py wedges of rats 13-day post DVC LV injection of GlyT1 shRNA (black bars, n=5) or MM (white bars, n=5). (e) Glucose infusion rates and (f) glucose production during clamps in rats injected with LV-MM (n=7), LV-GlyT1 shRNA (n=7)or LV-GlyT1 shRNA with DVC MK801 infusion (n=6). (e: *P<0.006; f: *P<0.003 versus LV-MM control and LV-GlyT1 shRNA+MK801 determined by ANOVA and Dunnett's post hoc test). Data are shown as the mean+s.e.m.
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f2: Molecular inhibition of DVC GlyT1 regulates glucose homeostasis in healthy rats.(a) Representative western blots and protein levels of plasma membrane GlyT1 (55, 70 and 90 kDa isoforms) normalized to insulin receptor (IR) in DVC wedges of rats 13-day post DVC lentiviral (LV) injection of GlyT1 shRNA (black bars, n=14) or a mismatch sequence (MM; white bars, n=11) as a control. *P<0.01, **P<0.001 determined by t-test. (b–d) Representative western blots and protein levels of plasma membrane GlyT1 (70 and/or 90 kDa isoforms) normalized to IR in sp5, Sp5C, Sp5I (L), sp5, Sp5C, Sp5I (R) and py wedges of rats 13-day post DVC LV injection of GlyT1 shRNA (black bars, n=5) or MM (white bars, n=5). (e) Glucose infusion rates and (f) glucose production during clamps in rats injected with LV-MM (n=7), LV-GlyT1 shRNA (n=7)or LV-GlyT1 shRNA with DVC MK801 infusion (n=6). (e: *P<0.006; f: *P<0.003 versus LV-MM control and LV-GlyT1 shRNA+MK801 determined by ANOVA and Dunnett's post hoc test). Data are shown as the mean+s.e.m.
Mentions: We alternatively tested the gluco-regulatory role of hindbrain GlyT1 inhibition via the targeted molecular knockdown of GlyT1 within the DVC. We first confirmed that lentiviral injection of GlyT1 shRNA (LV-GlyT1 shRNA) into the DVC selectively reduces the expression of both the 70- and 90-kDa isoforms of GlyT1 in plasma membrane fractions of only the DVC tissue compared with lentiviral injection of mismatch sequence (LV-MM), but not in the two adjacent left and right lateral regions of the DVC containing the Spinal trigeminal track (sp5), Spinal 5nu caudal part (Sp5C) and Spinal 5nu interpolar (Sp5I), and the region inferior to the DVC containing the pyramidal tract (py) of the same rats (Fig. 2a–d, Supplementary Fig. 3i–v). The dominant band at the molecular weight of 70–75 kDa corresponds to GlyT1a and b isoforms and the weaker band at90–100 kDa corresponds to GlyT1c isoform found in the rat brain as described34. The 90–100 kDa band is not detected in the sp5, Sp5C, Sp5I (right) and py regions (Fig. 2c,d). The immunoblot also reveals a strong band at 55 kDa (Fig. 2a), which is consistent with the occurrence of the partially glycosylated form of GlyT1 in the 55–60 kDa range as indicated353637. However, the 55 kDa GlyT1 band in the DVC of LV-GlyT1 shRNA versus LV-MM injected rats is not significantly different as compared with the effect on 70 kDa and 90 kDa bands (Fig. 2a). Nonetheless, the specific metabolic role of various forms of GlyT1 in the brain warrants future investigation.

View Article: PubMed Central - PubMed

ABSTRACT

Impaired glucose homeostasis and energy balance are integral to the pathophysiology of diabetes and obesity. Here we show that administration of a glycine transporter 1 (GlyT1) inhibitor, or molecular GlyT1 knockdown, in the dorsal vagal complex (DVC) suppresses glucose production, increases glucose tolerance and reduces food intake and body weight gain in healthy, obese and diabetic rats. These findings provide proof of concept that GlyT1 inhibition in the brain improves glucose and energy homeostasis. Considering the clinical safety and efficacy of GlyT1 inhibitors in raising glycine levels in clinical trials for schizophrenia, we propose that GlyT1 inhibitors have the potential to be repurposed as a treatment of both obesity and diabetes.

No MeSH data available.