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Positive regulation by GABA(B)R1 subunit of leptin expression through gene transactivation in adipocytes.

Nakamura Y, Hinoi E, Takarada T, Takahata Y, Yamamoto T, Fujita H, Takada S, Hashizume S, Yoneda Y - PLoS ONE (2011)

Bottom Line: However, no prominent expression was seen with mRNA for GABA(B)R2 subunit required for heteromeric orchestration of the functional GABA(B)R by any adipocytic cells and tissues.Knockdown by siRNA of GABA(B)R1 subunit led to significant decreases in leptin promoter activity and leptin mRNA levels in 3T3-L1 cells.Our results indicate that GABA(B)R1 subunit is constitutively expressed by adipocytes to primarily regulate leptin expression at the transcriptional level through a mechanism not relevant to the function as a partner of heterodimeric assembly to the functional GABA(B)R.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Molecular Pharmacology, Division of Pharmaceutical Sciences, Kanazawa University Graduate School of Natural Science and Technology, Kanazawa, Ishikawa, Japan.

ABSTRACT

Background: The view that γ-aminobutyric acid (GABA) plays a functional role in non-neuronal tissues, in addition to an inhibitory neurotransmitter role in the mammalian central nervous system, is prevailing, while little attention has been paid to GABAergic signaling machineries expressed by adipocytes to date. In this study, we attempted to demonstrate the possible functional expression of GABAergic signaling machineries by adipocytes.

Methodology/principal findings: GABA(B) receptor 1 (GABA(B)R1) subunit was constitutively expressed by mouse embryonic fibroblasts differentiated into adipocytes and adipocytic 3T3-L1 cells in culture, as well as mouse white adipose tissue, with no responsiveness to GABA(B)R ligands. However, no prominent expression was seen with mRNA for GABA(B)R2 subunit required for heteromeric orchestration of the functional GABA(B)R by any adipocytic cells and tissues. Leptin mRNA expression was significantly and selectively decreased in adipose tissue and embryonic fibroblasts, along with drastically reduced plasma leptin levels, in GABA(B)R1- mice than in wild-type mice. Knockdown by siRNA of GABA(B)R1 subunit led to significant decreases in leptin promoter activity and leptin mRNA levels in 3T3-L1 cells.

Conclusions/significance: Our results indicate that GABA(B)R1 subunit is constitutively expressed by adipocytes to primarily regulate leptin expression at the transcriptional level through a mechanism not relevant to the function as a partner of heterodimeric assembly to the functional GABA(B)R.

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Phenotypes of GABABR1- mice.Plasma was collected from blood of male mice at 4 weeks old, followed by measurement of (A) leptin and (B) insulin using ELISA kits. Various tissues were dissected from male mice at 4 weeks old, followed by measurement of wet weights of (C) WAT and (D) other tissues for calculation of the percentage over body weight. (E) Mice were fed with powder diets for 1 week, followed by measurement of the accumulated amount of food intake during 1 week. Open-field tests were carried out using male mice at 4 weeks old for (F) the number of crossing and (G) the number of rearing. (H) Body temperature was measured at the rectum in mice. (I) Mice were fasted for 15 to 16 h and then injected ip with 2 g/kg glucose, followed by determination of blood glucose levels 15 to 120 min after the injection. (J) Basal blood glucose levels were measured in male mice at 4 weeks old. Values are the mean ± S.E. from different experiments shown in the figure. *P<0.05; **P<0.01, significantly different from each control value obtained in WT mice. B.W., body weight; Gb1, GABABR1.
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pone-0020167-g004: Phenotypes of GABABR1- mice.Plasma was collected from blood of male mice at 4 weeks old, followed by measurement of (A) leptin and (B) insulin using ELISA kits. Various tissues were dissected from male mice at 4 weeks old, followed by measurement of wet weights of (C) WAT and (D) other tissues for calculation of the percentage over body weight. (E) Mice were fed with powder diets for 1 week, followed by measurement of the accumulated amount of food intake during 1 week. Open-field tests were carried out using male mice at 4 weeks old for (F) the number of crossing and (G) the number of rearing. (H) Body temperature was measured at the rectum in mice. (I) Mice were fasted for 15 to 16 h and then injected ip with 2 g/kg glucose, followed by determination of blood glucose levels 15 to 120 min after the injection. (J) Basal blood glucose levels were measured in male mice at 4 weeks old. Values are the mean ± S.E. from different experiments shown in the figure. *P<0.05; **P<0.01, significantly different from each control value obtained in WT mice. B.W., body weight; Gb1, GABABR1.

Mentions: For further evaluation of the possible function of GABABR1 subunit expressed by adipocytes, GABABR1- mice were analyzed for potential fat phenotypes under the normal chow diet at 4 weeks of age. GABABR1- mice exhibited a drastic decrease in plasma leptin levels (Figure 4A), with plasma insulin levels being unchanged (Figure 4B), compared with those in WT littermates. A significant reduction was seen in the ratio of WAT weight over body weight in GABABR1- mice compared with control littermates (Figure 4C), with no marked changes in the weights of other organs such as BAT, liver, kidney, pancreas, spleen and hypophysis (Figure 4D). By contrast, a significant increase was observed in the ratios of brain and heart weights over body weight in GABABR1- mice compared with those in WT mice.


Positive regulation by GABA(B)R1 subunit of leptin expression through gene transactivation in adipocytes.

Nakamura Y, Hinoi E, Takarada T, Takahata Y, Yamamoto T, Fujita H, Takada S, Hashizume S, Yoneda Y - PLoS ONE (2011)

Phenotypes of GABABR1- mice.Plasma was collected from blood of male mice at 4 weeks old, followed by measurement of (A) leptin and (B) insulin using ELISA kits. Various tissues were dissected from male mice at 4 weeks old, followed by measurement of wet weights of (C) WAT and (D) other tissues for calculation of the percentage over body weight. (E) Mice were fed with powder diets for 1 week, followed by measurement of the accumulated amount of food intake during 1 week. Open-field tests were carried out using male mice at 4 weeks old for (F) the number of crossing and (G) the number of rearing. (H) Body temperature was measured at the rectum in mice. (I) Mice were fasted for 15 to 16 h and then injected ip with 2 g/kg glucose, followed by determination of blood glucose levels 15 to 120 min after the injection. (J) Basal blood glucose levels were measured in male mice at 4 weeks old. Values are the mean ± S.E. from different experiments shown in the figure. *P<0.05; **P<0.01, significantly different from each control value obtained in WT mice. B.W., body weight; Gb1, GABABR1.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3105007&req=5

pone-0020167-g004: Phenotypes of GABABR1- mice.Plasma was collected from blood of male mice at 4 weeks old, followed by measurement of (A) leptin and (B) insulin using ELISA kits. Various tissues were dissected from male mice at 4 weeks old, followed by measurement of wet weights of (C) WAT and (D) other tissues for calculation of the percentage over body weight. (E) Mice were fed with powder diets for 1 week, followed by measurement of the accumulated amount of food intake during 1 week. Open-field tests were carried out using male mice at 4 weeks old for (F) the number of crossing and (G) the number of rearing. (H) Body temperature was measured at the rectum in mice. (I) Mice were fasted for 15 to 16 h and then injected ip with 2 g/kg glucose, followed by determination of blood glucose levels 15 to 120 min after the injection. (J) Basal blood glucose levels were measured in male mice at 4 weeks old. Values are the mean ± S.E. from different experiments shown in the figure. *P<0.05; **P<0.01, significantly different from each control value obtained in WT mice. B.W., body weight; Gb1, GABABR1.
Mentions: For further evaluation of the possible function of GABABR1 subunit expressed by adipocytes, GABABR1- mice were analyzed for potential fat phenotypes under the normal chow diet at 4 weeks of age. GABABR1- mice exhibited a drastic decrease in plasma leptin levels (Figure 4A), with plasma insulin levels being unchanged (Figure 4B), compared with those in WT littermates. A significant reduction was seen in the ratio of WAT weight over body weight in GABABR1- mice compared with control littermates (Figure 4C), with no marked changes in the weights of other organs such as BAT, liver, kidney, pancreas, spleen and hypophysis (Figure 4D). By contrast, a significant increase was observed in the ratios of brain and heart weights over body weight in GABABR1- mice compared with those in WT mice.

Bottom Line: However, no prominent expression was seen with mRNA for GABA(B)R2 subunit required for heteromeric orchestration of the functional GABA(B)R by any adipocytic cells and tissues.Knockdown by siRNA of GABA(B)R1 subunit led to significant decreases in leptin promoter activity and leptin mRNA levels in 3T3-L1 cells.Our results indicate that GABA(B)R1 subunit is constitutively expressed by adipocytes to primarily regulate leptin expression at the transcriptional level through a mechanism not relevant to the function as a partner of heterodimeric assembly to the functional GABA(B)R.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Molecular Pharmacology, Division of Pharmaceutical Sciences, Kanazawa University Graduate School of Natural Science and Technology, Kanazawa, Ishikawa, Japan.

ABSTRACT

Background: The view that γ-aminobutyric acid (GABA) plays a functional role in non-neuronal tissues, in addition to an inhibitory neurotransmitter role in the mammalian central nervous system, is prevailing, while little attention has been paid to GABAergic signaling machineries expressed by adipocytes to date. In this study, we attempted to demonstrate the possible functional expression of GABAergic signaling machineries by adipocytes.

Methodology/principal findings: GABA(B) receptor 1 (GABA(B)R1) subunit was constitutively expressed by mouse embryonic fibroblasts differentiated into adipocytes and adipocytic 3T3-L1 cells in culture, as well as mouse white adipose tissue, with no responsiveness to GABA(B)R ligands. However, no prominent expression was seen with mRNA for GABA(B)R2 subunit required for heteromeric orchestration of the functional GABA(B)R by any adipocytic cells and tissues. Leptin mRNA expression was significantly and selectively decreased in adipose tissue and embryonic fibroblasts, along with drastically reduced plasma leptin levels, in GABA(B)R1- mice than in wild-type mice. Knockdown by siRNA of GABA(B)R1 subunit led to significant decreases in leptin promoter activity and leptin mRNA levels in 3T3-L1 cells.

Conclusions/significance: Our results indicate that GABA(B)R1 subunit is constitutively expressed by adipocytes to primarily regulate leptin expression at the transcriptional level through a mechanism not relevant to the function as a partner of heterodimeric assembly to the functional GABA(B)R.

Show MeSH