Transgenic Reexpression of GLUT1 or GLUT2 in Pancreatic β Cells Rescues GLUT2-null Mice from Early Death and Restores Normal Glucose-stimulated Insulin Secretion

Abstract

GLUT2-null mice are hyperglycemic, hypoinsulinemic, hyperglucagonemic, and glycosuric and die within the first 3 weeks of life. Their endocrine pancreas shows a loss of first phase glucose-stimulated insulin secretion (GSIS) and inverse ␣ to ␤ cell ratio. Here we show that reexpression by transgenesis of either GLUT1 or GLUT2 in the pancreatic ␤ cells of these mice allowed mouse survival and breeding. The rescued mice had normal-fed glycemia but fasted hypoglycemia, glycosuria, and an elevated glucagon to insulin ratio. Glucose tolerance was, however, normal. In vivo, insulin secretion assessed following hyperglycemic clamps was normal. In vitro, islet perifusion studies revealed that first phase of insulin secretion was restored as well by GLUT1 or GLUT2, and this was accompanied by normalization of the glucose utilization rate. The ratio of pancreatic insulin to glucagon and volume densities of ␣ to ␤ cells were, however, not corrected. These data demonstrate that 1) reexpression of GLUT1 or GLUT2 in ␤ cells is sufficient to rescue GLUT2-null mice from lethality, 2) GLUT1 as well as GLUT2 can restore normal GSIS, 3) restoration of GSIS does not correct the abnormal composition of the endocrine pancreas. Thus, normal GSIS does not depend on transporter affinity but on the rate of uptake at stimulatory glucose concentrations.Genetic inactivation of the glucose transporter GLUT2 by homologous recombination in embryonic stem cells allowed us to generate mice deficient in the activity of this transporter (1). These GLUT2-null mice were unable to survive past the third week of life and displayed a form of diabetes mellitus characterized by hyperglycemia, relative hypoinsulinemia, and elevated glucagon levels. Their glucose tolerance was severely impaired, and as assessed in islet perifusion experiments, their pancreatic ␤ cells had completely lost the first phase while maintaining a second phase of insulin secretion. The cellular composition of the islets of Langerhans was also modified with an inversion of the ␣ to ␤ cell ratio due to an absolute increase in ␣ cell number and absolute decrease in ␤ cell numbers. As this change in cellular composition appeared only after birth, which is the time at which glucose sensitivity by pancreatic ␤ cells is being established, we postulated that the altered cellular composition of the islets was due to a defect in the secretion by ␤ cells of autocrine proliferation factors or of paracrine factors normally restricting ␣ cell proliferation.Glucose-stimulated insulin secretion (GSIS) 1 is initiated by the uptake of glucose by the glucose transporter GLUT2. Glucose is then phosphorylated by glucokinase and further metabolized through the glycolytic pathway. Activation of the mitochondrial metabolism then leads to generation of coupling factors, ATP (2, 3) and glutamate (4), which trigger the distal steps in insulin granule exocytosis. The high K m (6 mM) hexokinase IV, referred to as glucokinase, is the rate-limiting step in glucose metabolism by ␤ cells, and it therefore has a h...

Oscillatory control of insulin secretion

“…It was therefore unexpected that human β-cells preferentially express low K m GLUT1 (De Vos et al, 1995), whose capacity is close to saturation already at threshold concentrations of glucose. However, consistent with the latter observation, transgenic re-expression of GLUT1 or GLUT2 into β-cells of GLUT2-null mice were equally efficient in restoring normal glucose sensing (Thorens et al, 2000). The explanation is probably a very high glucose transport capacity that is not rate limiting.…”

Expression of the gene encoding the high-K m glucose transporter 2 by the early postimplantation mouse embryo is essential for neural tube defects associated with diabetic embryopathy

“…RNA was isolated and reverse-transcribed, and serial dilutions of the reverse transcription products were amplified by PCR as described previously [35]. The cDNA products were amplified using primers complementary to the Glut2 amplicon [43] and the PCR conditions described below, or to constitutively expressed 36B4 (also known as Arbp), which was used as a normalisation control, as described [35,44]. Autoradiographs of the electrophoresed PCR products were scanned, and unsaturated bands were quantified using Adobe Photoshop (version 9.0.1, Adobe Systems, Inc., San Jose, CA).…”

“…Genotype analysis of Glut2 allele Glut2 +/− pups were identified by PCR analysis of tail DNA using the previously reported primer sequences [42,43] …”

Glial hypothalamic inhibition of GLUT2 expression alters satiety, impacting eating behavior

“…Due to its kinetic properties and tissue localization, GLUT2 is involved in the glucosensing mechanism as it has a uniquely low affinity for glucose (Km ∼17 mM) and can also use mannose, galactose, and fructose as low affinity substrates for transport (Thorens, Guillam, Beermann, Burcelin, & Jaquet, 2000). GLUT2 is the major glucose transporter present in hepatocytes, enterocytes, kidney epithelial cells, and cells of the hepatoportal vein (Thorens, 2015).…”

“…GLUT2 is also the major glucose transporter in pancreatic ß‐cells, where its genetic inactivation impairs glucose uptake and suppresses glucose‐stimulated insulin secretion. GLUT2 −/− mice die at around the weaning period, and transgenic expression of another glucose transporter, GLUT1, in β‐cells (RIPGlut1;GLUT2 −/− ) restores normal glucose‐stimulated insulin biosynthesis (Bady et al, 2006; Guillam et al, 1997; Thorens et al, 2000). …”