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Pannexin 1 is required for full activation of insulin-stimulated glucose uptake in adipocytes.

Adamson SE, Meher AK, Chiu YH, Sandilos JK, Oberholtzer NP, Walker NN, Hargett SR, Seaman SA, Peirce-Cottler SM, Isakson BE, McNamara CA, Keller SR, Harris TE, Bayliss DA, Leitinger N - Mol Metab (2015)

Bottom Line: Finally, we measured Panx1 mRNA in human visceral adipose tissue samples by qRT-PCR and compared expression levels with glucose levels and HOMA-IR measurements in patients.Our data show that adipocytes express functional Pannexin 1 (Panx1) channels that can be activated to release ATP.We show that Panx1 channel activity regulates insulin-stimulated glucose uptake in adipocytes and thus contributes to control of metabolic homeostasis.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology, University of Virginia, Charlottesville, VA 22908, USA ; Cardiovascular Research Center, University of Virginia, Charlottesville, VA 22908, USA.

ABSTRACT

Objective: Defective glucose uptake in adipocytes leads to impaired metabolic homeostasis and insulin resistance, hallmarks of type 2 diabetes. Extracellular ATP-derived nucleotides and nucleosides are important regulators of adipocyte function, but the pathway for controlled ATP release from adipocytes is unknown. Here, we investigated whether Pannexin 1 (Panx1) channels control ATP release from adipocytes and contribute to metabolic homeostasis.

Methods: We assessed Panx1 functionality in cultured 3T3-L1 adipocytes and in adipocytes isolated from murine white adipose tissue by measuring ATP release in response to known activators of Panx1 channels. Glucose uptake in cultured 3T3-L1 adipocytes was measured in the presence of Panx1 pharmacologic inhibitors and in adipocytes isolated from white adipose tissue from wildtype (WT) or adipocyte-specific Panx1 knockout (AdipPanx1 KO) mice generated in our laboratory. We performed in vivo glucose uptake studies in chow fed WT and AdipPanx1 KO mice and assessed insulin resistance in WT and AdipPanx1 KO mice fed a high fat diet for 12 weeks. Panx1 channel function was assessed in response to insulin by performing electrophysiologic recordings in a heterologous expression system. Finally, we measured Panx1 mRNA in human visceral adipose tissue samples by qRT-PCR and compared expression levels with glucose levels and HOMA-IR measurements in patients.

Results: Our data show that adipocytes express functional Pannexin 1 (Panx1) channels that can be activated to release ATP. Pharmacologic inhibition or selective genetic deletion of Panx1 from adipocytes decreased insulin-induced glucose uptake in vitro and in vivo and exacerbated diet-induced insulin resistance in mice. Further, we identify insulin as a novel activator of Panx1 channels. In obese humans Panx1 expression in adipose tissue is increased and correlates with the degree of insulin resistance.

Conclusions: We show that Panx1 channel activity regulates insulin-stimulated glucose uptake in adipocytes and thus contributes to control of metabolic homeostasis.

No MeSH data available.


Related in: MedlinePlus

Weight gain, energy expenditure, serum free fatty acids and insulin are not different between high fat fed WT and AdipPanx1KO mice. (A) Intraperitoneal glucose tolerance test was performed on age-matched, male, WT and AdipPanxKO littermates that had been fed chow or high fat diet (60% fat) for 12 weeks. Mice were injected i.p. with 1 g/kg glucose, and blood glucose was measured in tail vein blood via glucometer (One Touch Ultra). Data are presented as mean ± s.e.m and representative of 3 independent experiments (WT HFD n = 6, AdipPanxKO HFD n = 9, WT chow n = 7, AdipPanxKO chow n = 6). Combined area under the curve (AUC) analysis of glucose tolerance tests reveals that AdipPanxKO mice are significantly more glucose intolerant after high fat feeding compared to WT mice (WT HFD n = 18, AdipPanxKO HFD n = 14, WT chow n = 7, AdipPanxKO chow n = 4); *p = 0.025 by 2-tailed Student's t-test. Box plots represent the 10th to 90th percentile. Three mice in the high fat diet group (WT n = 1, AdipPanxKO n = 2) did not respond to diet as evidenced by AUC not being different from chow groups and thus were excluded from the analysis. One data point (WT n = 1) was greater than 2 standard deviations from the mean and thus was excluded. (B) Intraperitoneal insulin tolerance tests (0.75 U/kg) was performed on age-matched, male, WT and AdipPanxKO littermates that had been fed chow or high fat diet (60% fat) for 12 weeks. n = 6–7 mice per group. Data are expressed as mean + s.e.m. Box plots represent the 10th to 90th percentile. *p < 0.05 by Student's t-test. (C,D) Male WT and AdipPanx1KO littermates were fed a high fat diet (60% fat) for 12 weeks. Mice on a high fat diet were weighed weekly. Adiposity (%fat) and lean mass (%lean) were assessed by echo MRI. Data are representative of 3 independent experiments, n = 6 WT, n = 9 AdipPanx1KO. Data are expressed as mean ± s.e.m. (E) Serum free fatty acids (FFA) were measured after 12 weeks of high fat feeding in WT (n = 5) and AdipPanxKO (n = 9) mice using the colorimetric FFA kit from Wako. Data are expressed as mean ± s.d. (F) Serum insulin in WT and AdipPanxKO mice that were either fed chow diet or HFD for 12 weeks were measured after a 5 h fast. (n = 6 WT chow, n = 7 AdipPanxKO chow, n = 5 WT high fat diet, n = 9 AdipPanxKO high fat diet). Data are expressed as mean ± s.d. (G–I) High fat diet-fed (12 weeks) and chow-fed WT and AdipPanx1KO mice (n = 4 per group) were placed in metabolic cages for 72 h. Average VO2 consumption, VCO2, and respiratory exchange ratio (RER) by animal are shown for light and dark periods. The initial 4 h of readings were not part of average light values as this was time for mice to acclimate. Box plots represent the 10th to 90th percentile. (J) Locomotion was recorded as X- and Y-axis beam breaks during light and dark cycle. Total beam breaks were not different between WT and AdipPanx1KO mice indicating no overall difference in locomotion. Box plots represent the 10th to 90th percentile.
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fig4: Weight gain, energy expenditure, serum free fatty acids and insulin are not different between high fat fed WT and AdipPanx1KO mice. (A) Intraperitoneal glucose tolerance test was performed on age-matched, male, WT and AdipPanxKO littermates that had been fed chow or high fat diet (60% fat) for 12 weeks. Mice were injected i.p. with 1 g/kg glucose, and blood glucose was measured in tail vein blood via glucometer (One Touch Ultra). Data are presented as mean ± s.e.m and representative of 3 independent experiments (WT HFD n = 6, AdipPanxKO HFD n = 9, WT chow n = 7, AdipPanxKO chow n = 6). Combined area under the curve (AUC) analysis of glucose tolerance tests reveals that AdipPanxKO mice are significantly more glucose intolerant after high fat feeding compared to WT mice (WT HFD n = 18, AdipPanxKO HFD n = 14, WT chow n = 7, AdipPanxKO chow n = 4); *p = 0.025 by 2-tailed Student's t-test. Box plots represent the 10th to 90th percentile. Three mice in the high fat diet group (WT n = 1, AdipPanxKO n = 2) did not respond to diet as evidenced by AUC not being different from chow groups and thus were excluded from the analysis. One data point (WT n = 1) was greater than 2 standard deviations from the mean and thus was excluded. (B) Intraperitoneal insulin tolerance tests (0.75 U/kg) was performed on age-matched, male, WT and AdipPanxKO littermates that had been fed chow or high fat diet (60% fat) for 12 weeks. n = 6–7 mice per group. Data are expressed as mean + s.e.m. Box plots represent the 10th to 90th percentile. *p < 0.05 by Student's t-test. (C,D) Male WT and AdipPanx1KO littermates were fed a high fat diet (60% fat) for 12 weeks. Mice on a high fat diet were weighed weekly. Adiposity (%fat) and lean mass (%lean) were assessed by echo MRI. Data are representative of 3 independent experiments, n = 6 WT, n = 9 AdipPanx1KO. Data are expressed as mean ± s.e.m. (E) Serum free fatty acids (FFA) were measured after 12 weeks of high fat feeding in WT (n = 5) and AdipPanxKO (n = 9) mice using the colorimetric FFA kit from Wako. Data are expressed as mean ± s.d. (F) Serum insulin in WT and AdipPanxKO mice that were either fed chow diet or HFD for 12 weeks were measured after a 5 h fast. (n = 6 WT chow, n = 7 AdipPanxKO chow, n = 5 WT high fat diet, n = 9 AdipPanxKO high fat diet). Data are expressed as mean ± s.d. (G–I) High fat diet-fed (12 weeks) and chow-fed WT and AdipPanx1KO mice (n = 4 per group) were placed in metabolic cages for 72 h. Average VO2 consumption, VCO2, and respiratory exchange ratio (RER) by animal are shown for light and dark periods. The initial 4 h of readings were not part of average light values as this was time for mice to acclimate. Box plots represent the 10th to 90th percentile. (J) Locomotion was recorded as X- and Y-axis beam breaks during light and dark cycle. Total beam breaks were not different between WT and AdipPanx1KO mice indicating no overall difference in locomotion. Box plots represent the 10th to 90th percentile.

Mentions: Impaired glucose uptake in adipose tissue was shown to adversely affect high fat diet-induced insulin resistance [2,37]. To test whether the 20% reduction in glucose uptake rate that we observed in the adipose tissue of AdipPanx1KO mice (Figure 2C) would have an effect on the development of insulin resistance, we fed WT and AdipPanx1KO mice a diabetogenic high fat diet. The absence of Panx1 from adipocytes slightly, but significantly, exacerbated measures of insulin resistance, including glucose and insulin tolerance after 12 weeks of high fat diet feeding (Figure 4A and B). However, no difference in weight gain or adiposity was observed between the groups (Figure 4C and D). Serum free fatty acid levels were also similar between high fat diet-fed WT and AdipPanx1KO mice (Figure 4E), and high fat feeding increased serum insulin levels in both WT and AdipPanx1KO mice to the same extent (Figure 4F).


Pannexin 1 is required for full activation of insulin-stimulated glucose uptake in adipocytes.

Adamson SE, Meher AK, Chiu YH, Sandilos JK, Oberholtzer NP, Walker NN, Hargett SR, Seaman SA, Peirce-Cottler SM, Isakson BE, McNamara CA, Keller SR, Harris TE, Bayliss DA, Leitinger N - Mol Metab (2015)

Weight gain, energy expenditure, serum free fatty acids and insulin are not different between high fat fed WT and AdipPanx1KO mice. (A) Intraperitoneal glucose tolerance test was performed on age-matched, male, WT and AdipPanxKO littermates that had been fed chow or high fat diet (60% fat) for 12 weeks. Mice were injected i.p. with 1 g/kg glucose, and blood glucose was measured in tail vein blood via glucometer (One Touch Ultra). Data are presented as mean ± s.e.m and representative of 3 independent experiments (WT HFD n = 6, AdipPanxKO HFD n = 9, WT chow n = 7, AdipPanxKO chow n = 6). Combined area under the curve (AUC) analysis of glucose tolerance tests reveals that AdipPanxKO mice are significantly more glucose intolerant after high fat feeding compared to WT mice (WT HFD n = 18, AdipPanxKO HFD n = 14, WT chow n = 7, AdipPanxKO chow n = 4); *p = 0.025 by 2-tailed Student's t-test. Box plots represent the 10th to 90th percentile. Three mice in the high fat diet group (WT n = 1, AdipPanxKO n = 2) did not respond to diet as evidenced by AUC not being different from chow groups and thus were excluded from the analysis. One data point (WT n = 1) was greater than 2 standard deviations from the mean and thus was excluded. (B) Intraperitoneal insulin tolerance tests (0.75 U/kg) was performed on age-matched, male, WT and AdipPanxKO littermates that had been fed chow or high fat diet (60% fat) for 12 weeks. n = 6–7 mice per group. Data are expressed as mean + s.e.m. Box plots represent the 10th to 90th percentile. *p < 0.05 by Student's t-test. (C,D) Male WT and AdipPanx1KO littermates were fed a high fat diet (60% fat) for 12 weeks. Mice on a high fat diet were weighed weekly. Adiposity (%fat) and lean mass (%lean) were assessed by echo MRI. Data are representative of 3 independent experiments, n = 6 WT, n = 9 AdipPanx1KO. Data are expressed as mean ± s.e.m. (E) Serum free fatty acids (FFA) were measured after 12 weeks of high fat feeding in WT (n = 5) and AdipPanxKO (n = 9) mice using the colorimetric FFA kit from Wako. Data are expressed as mean ± s.d. (F) Serum insulin in WT and AdipPanxKO mice that were either fed chow diet or HFD for 12 weeks were measured after a 5 h fast. (n = 6 WT chow, n = 7 AdipPanxKO chow, n = 5 WT high fat diet, n = 9 AdipPanxKO high fat diet). Data are expressed as mean ± s.d. (G–I) High fat diet-fed (12 weeks) and chow-fed WT and AdipPanx1KO mice (n = 4 per group) were placed in metabolic cages for 72 h. Average VO2 consumption, VCO2, and respiratory exchange ratio (RER) by animal are shown for light and dark periods. The initial 4 h of readings were not part of average light values as this was time for mice to acclimate. Box plots represent the 10th to 90th percentile. (J) Locomotion was recorded as X- and Y-axis beam breaks during light and dark cycle. Total beam breaks were not different between WT and AdipPanx1KO mice indicating no overall difference in locomotion. Box plots represent the 10th to 90th percentile.
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fig4: Weight gain, energy expenditure, serum free fatty acids and insulin are not different between high fat fed WT and AdipPanx1KO mice. (A) Intraperitoneal glucose tolerance test was performed on age-matched, male, WT and AdipPanxKO littermates that had been fed chow or high fat diet (60% fat) for 12 weeks. Mice were injected i.p. with 1 g/kg glucose, and blood glucose was measured in tail vein blood via glucometer (One Touch Ultra). Data are presented as mean ± s.e.m and representative of 3 independent experiments (WT HFD n = 6, AdipPanxKO HFD n = 9, WT chow n = 7, AdipPanxKO chow n = 6). Combined area under the curve (AUC) analysis of glucose tolerance tests reveals that AdipPanxKO mice are significantly more glucose intolerant after high fat feeding compared to WT mice (WT HFD n = 18, AdipPanxKO HFD n = 14, WT chow n = 7, AdipPanxKO chow n = 4); *p = 0.025 by 2-tailed Student's t-test. Box plots represent the 10th to 90th percentile. Three mice in the high fat diet group (WT n = 1, AdipPanxKO n = 2) did not respond to diet as evidenced by AUC not being different from chow groups and thus were excluded from the analysis. One data point (WT n = 1) was greater than 2 standard deviations from the mean and thus was excluded. (B) Intraperitoneal insulin tolerance tests (0.75 U/kg) was performed on age-matched, male, WT and AdipPanxKO littermates that had been fed chow or high fat diet (60% fat) for 12 weeks. n = 6–7 mice per group. Data are expressed as mean + s.e.m. Box plots represent the 10th to 90th percentile. *p < 0.05 by Student's t-test. (C,D) Male WT and AdipPanx1KO littermates were fed a high fat diet (60% fat) for 12 weeks. Mice on a high fat diet were weighed weekly. Adiposity (%fat) and lean mass (%lean) were assessed by echo MRI. Data are representative of 3 independent experiments, n = 6 WT, n = 9 AdipPanx1KO. Data are expressed as mean ± s.e.m. (E) Serum free fatty acids (FFA) were measured after 12 weeks of high fat feeding in WT (n = 5) and AdipPanxKO (n = 9) mice using the colorimetric FFA kit from Wako. Data are expressed as mean ± s.d. (F) Serum insulin in WT and AdipPanxKO mice that were either fed chow diet or HFD for 12 weeks were measured after a 5 h fast. (n = 6 WT chow, n = 7 AdipPanxKO chow, n = 5 WT high fat diet, n = 9 AdipPanxKO high fat diet). Data are expressed as mean ± s.d. (G–I) High fat diet-fed (12 weeks) and chow-fed WT and AdipPanx1KO mice (n = 4 per group) were placed in metabolic cages for 72 h. Average VO2 consumption, VCO2, and respiratory exchange ratio (RER) by animal are shown for light and dark periods. The initial 4 h of readings were not part of average light values as this was time for mice to acclimate. Box plots represent the 10th to 90th percentile. (J) Locomotion was recorded as X- and Y-axis beam breaks during light and dark cycle. Total beam breaks were not different between WT and AdipPanx1KO mice indicating no overall difference in locomotion. Box plots represent the 10th to 90th percentile.
Mentions: Impaired glucose uptake in adipose tissue was shown to adversely affect high fat diet-induced insulin resistance [2,37]. To test whether the 20% reduction in glucose uptake rate that we observed in the adipose tissue of AdipPanx1KO mice (Figure 2C) would have an effect on the development of insulin resistance, we fed WT and AdipPanx1KO mice a diabetogenic high fat diet. The absence of Panx1 from adipocytes slightly, but significantly, exacerbated measures of insulin resistance, including glucose and insulin tolerance after 12 weeks of high fat diet feeding (Figure 4A and B). However, no difference in weight gain or adiposity was observed between the groups (Figure 4C and D). Serum free fatty acid levels were also similar between high fat diet-fed WT and AdipPanx1KO mice (Figure 4E), and high fat feeding increased serum insulin levels in both WT and AdipPanx1KO mice to the same extent (Figure 4F).

Bottom Line: Finally, we measured Panx1 mRNA in human visceral adipose tissue samples by qRT-PCR and compared expression levels with glucose levels and HOMA-IR measurements in patients.Our data show that adipocytes express functional Pannexin 1 (Panx1) channels that can be activated to release ATP.We show that Panx1 channel activity regulates insulin-stimulated glucose uptake in adipocytes and thus contributes to control of metabolic homeostasis.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology, University of Virginia, Charlottesville, VA 22908, USA ; Cardiovascular Research Center, University of Virginia, Charlottesville, VA 22908, USA.

ABSTRACT

Objective: Defective glucose uptake in adipocytes leads to impaired metabolic homeostasis and insulin resistance, hallmarks of type 2 diabetes. Extracellular ATP-derived nucleotides and nucleosides are important regulators of adipocyte function, but the pathway for controlled ATP release from adipocytes is unknown. Here, we investigated whether Pannexin 1 (Panx1) channels control ATP release from adipocytes and contribute to metabolic homeostasis.

Methods: We assessed Panx1 functionality in cultured 3T3-L1 adipocytes and in adipocytes isolated from murine white adipose tissue by measuring ATP release in response to known activators of Panx1 channels. Glucose uptake in cultured 3T3-L1 adipocytes was measured in the presence of Panx1 pharmacologic inhibitors and in adipocytes isolated from white adipose tissue from wildtype (WT) or adipocyte-specific Panx1 knockout (AdipPanx1 KO) mice generated in our laboratory. We performed in vivo glucose uptake studies in chow fed WT and AdipPanx1 KO mice and assessed insulin resistance in WT and AdipPanx1 KO mice fed a high fat diet for 12 weeks. Panx1 channel function was assessed in response to insulin by performing electrophysiologic recordings in a heterologous expression system. Finally, we measured Panx1 mRNA in human visceral adipose tissue samples by qRT-PCR and compared expression levels with glucose levels and HOMA-IR measurements in patients.

Results: Our data show that adipocytes express functional Pannexin 1 (Panx1) channels that can be activated to release ATP. Pharmacologic inhibition or selective genetic deletion of Panx1 from adipocytes decreased insulin-induced glucose uptake in vitro and in vivo and exacerbated diet-induced insulin resistance in mice. Further, we identify insulin as a novel activator of Panx1 channels. In obese humans Panx1 expression in adipose tissue is increased and correlates with the degree of insulin resistance.

Conclusions: We show that Panx1 channel activity regulates insulin-stimulated glucose uptake in adipocytes and thus contributes to control of metabolic homeostasis.

No MeSH data available.


Related in: MedlinePlus