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HCO 3--dependent volume regulation in alpha-cells of the rat endocrine pancreas.

Davies SL, Best L, Brown PD - Pflugers Arch. (2009)

Bottom Line: A RVI was observed, however, in cells that had first undergone a regulatory volume decrease (RVD), but only in HCO(3)(-)-buffered solutions.The post-RVD RVI and the glucose-induced RVI were both inhibited by 10 microM 5-(N-methyl-N-isobutyl) amiloride or 100 microM 2,2'-(1,2-ethenediyl) bis (5-isothio-cyanatobenzenesulfonic acid), but not by 10 microM benzamil nor 10 microM bumetanide.These data suggest that Na(+)-H(+) exchangers and Cl(-)-HCO(3)(-) exchangers contribute to volume regulation in alpha-cells.

View Article: PubMed Central - PubMed

Affiliation: Faculty of Life Sciences, University of Manchester, Second Floor CTF Building, Manchester, M13 9NT, UK.

ABSTRACT
Ion transport activity in pancreatic alpha-cells was assessed by studying cell volume regulation in response to anisotonic solutions. Cell volume was measured by a video imaging method, and cells were superfused with either 4-(2-hydroxyethyl)piperazine-1-ethanesulfonic acid-buffered or HCO(3)(-) -buffered solutions. alpha-Cells did not exhibit a regulatory volume increase (RVI) in response to cell shrinkage caused by hypertonic solutions. A RVI was observed, however, in cells that had first undergone a regulatory volume decrease (RVD), but only in HCO(3)(-)-buffered solutions. RVI was also observed in response to a HCO(3)(-) -buffered hypertonic solution in which the glucose concentration was increased from 4 to 20 mM. The post-RVD RVI and the glucose-induced RVI were both inhibited by 10 microM 5-(N-methyl-N-isobutyl) amiloride or 100 microM 2,2'-(1,2-ethenediyl) bis (5-isothio-cyanatobenzenesulfonic acid), but not by 10 microM benzamil nor 10 microM bumetanide. These data suggest that Na(+)-H(+) exchangers and Cl(-)-HCO(3)(-) exchangers contribute to volume regulation in alpha-cells.

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Isolated pancreatic α-cells behave as osmometers in anisotonic solutions. a Image of an isolated α-cell with a volume of 0.65 pl in HCO3−-buffered isotonic solution. The bar indicates 2 μm. b Boyle–van’t Hoff plot of α-cell relative cell volume as a function of the reciprocal of superfusate osmolality (1/osmolality). Cell volumes are the maximum or minimum recorded when cells were superfused with hypotonic or hypertonic solutions, respectively. Solutions were all buffered with HEPES and include the isotonic (285 mOsm. kg H2O−1; n = 1), hypotonic (182 mOsm. kg H2O−1; n = 6), and hypertonic (384 mOsm. kg H2O−1; n = 6) solutions described in Table 1. Two additional solutions were used: 433 mOsm. kg H2O−1 (the hypertonic solution but with 150 mM mannitol; n = 3) and 234 mOsm. kg H2O−1 (the hypotonic solution but with 120 mM Na Cl; n = 3). Data are mean ± SEM and the line through the data fitted by linear regression analysis (R2 = 0.982). Extrapolating this line to infinite osmotic strength (1/osmolality = 0) gives an osmotically inactive space of 0.24
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Fig1: Isolated pancreatic α-cells behave as osmometers in anisotonic solutions. a Image of an isolated α-cell with a volume of 0.65 pl in HCO3−-buffered isotonic solution. The bar indicates 2 μm. b Boyle–van’t Hoff plot of α-cell relative cell volume as a function of the reciprocal of superfusate osmolality (1/osmolality). Cell volumes are the maximum or minimum recorded when cells were superfused with hypotonic or hypertonic solutions, respectively. Solutions were all buffered with HEPES and include the isotonic (285 mOsm. kg H2O−1; n = 1), hypotonic (182 mOsm. kg H2O−1; n = 6), and hypertonic (384 mOsm. kg H2O−1; n = 6) solutions described in Table 1. Two additional solutions were used: 433 mOsm. kg H2O−1 (the hypertonic solution but with 150 mM mannitol; n = 3) and 234 mOsm. kg H2O−1 (the hypotonic solution but with 120 mM Na Cl; n = 3). Data are mean ± SEM and the line through the data fitted by linear regression analysis (R2 = 0.982). Extrapolating this line to infinite osmotic strength (1/osmolality = 0) gives an osmotically inactive space of 0.24

Mentions: Figure 1a shows the image of a pancreatic α-cell recorded 48 h after isolation. The cell is typical of the α-cells used in this study; it has a volume of 0.65 pl and is circular in appearance. The circular cross section suggests that the cells retain a spherical morphology, which is a major assumption in the volume measurement method. To further test this assumption, volume changes in response to a range of extracellular osmolalities were examined. Figure 1b shows a Boyle–van’t Hoff plot of α-cell volume as a function of osmolality (see [28]). Over the range of osmolalities used in this study, the α-cells behave as osmometers so that volume was linearly related to superfusate osmolality. These data indicate that the method is working as one would predict and suggest that the α-cells must be retaining a spherical shape during the experiments. An osmotically inactive space of 0.24 was predicted by the linear regression line fitted to the data. This value is similar to 0.26 measured in pancreatic β-cells using similar methods [20].Fig. 1


HCO 3--dependent volume regulation in alpha-cells of the rat endocrine pancreas.

Davies SL, Best L, Brown PD - Pflugers Arch. (2009)

Isolated pancreatic α-cells behave as osmometers in anisotonic solutions. a Image of an isolated α-cell with a volume of 0.65 pl in HCO3−-buffered isotonic solution. The bar indicates 2 μm. b Boyle–van’t Hoff plot of α-cell relative cell volume as a function of the reciprocal of superfusate osmolality (1/osmolality). Cell volumes are the maximum or minimum recorded when cells were superfused with hypotonic or hypertonic solutions, respectively. Solutions were all buffered with HEPES and include the isotonic (285 mOsm. kg H2O−1; n = 1), hypotonic (182 mOsm. kg H2O−1; n = 6), and hypertonic (384 mOsm. kg H2O−1; n = 6) solutions described in Table 1. Two additional solutions were used: 433 mOsm. kg H2O−1 (the hypertonic solution but with 150 mM mannitol; n = 3) and 234 mOsm. kg H2O−1 (the hypotonic solution but with 120 mM Na Cl; n = 3). Data are mean ± SEM and the line through the data fitted by linear regression analysis (R2 = 0.982). Extrapolating this line to infinite osmotic strength (1/osmolality = 0) gives an osmotically inactive space of 0.24
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Related In: Results  -  Collection

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Fig1: Isolated pancreatic α-cells behave as osmometers in anisotonic solutions. a Image of an isolated α-cell with a volume of 0.65 pl in HCO3−-buffered isotonic solution. The bar indicates 2 μm. b Boyle–van’t Hoff plot of α-cell relative cell volume as a function of the reciprocal of superfusate osmolality (1/osmolality). Cell volumes are the maximum or minimum recorded when cells were superfused with hypotonic or hypertonic solutions, respectively. Solutions were all buffered with HEPES and include the isotonic (285 mOsm. kg H2O−1; n = 1), hypotonic (182 mOsm. kg H2O−1; n = 6), and hypertonic (384 mOsm. kg H2O−1; n = 6) solutions described in Table 1. Two additional solutions were used: 433 mOsm. kg H2O−1 (the hypertonic solution but with 150 mM mannitol; n = 3) and 234 mOsm. kg H2O−1 (the hypotonic solution but with 120 mM Na Cl; n = 3). Data are mean ± SEM and the line through the data fitted by linear regression analysis (R2 = 0.982). Extrapolating this line to infinite osmotic strength (1/osmolality = 0) gives an osmotically inactive space of 0.24
Mentions: Figure 1a shows the image of a pancreatic α-cell recorded 48 h after isolation. The cell is typical of the α-cells used in this study; it has a volume of 0.65 pl and is circular in appearance. The circular cross section suggests that the cells retain a spherical morphology, which is a major assumption in the volume measurement method. To further test this assumption, volume changes in response to a range of extracellular osmolalities were examined. Figure 1b shows a Boyle–van’t Hoff plot of α-cell volume as a function of osmolality (see [28]). Over the range of osmolalities used in this study, the α-cells behave as osmometers so that volume was linearly related to superfusate osmolality. These data indicate that the method is working as one would predict and suggest that the α-cells must be retaining a spherical shape during the experiments. An osmotically inactive space of 0.24 was predicted by the linear regression line fitted to the data. This value is similar to 0.26 measured in pancreatic β-cells using similar methods [20].Fig. 1

Bottom Line: A RVI was observed, however, in cells that had first undergone a regulatory volume decrease (RVD), but only in HCO(3)(-)-buffered solutions.The post-RVD RVI and the glucose-induced RVI were both inhibited by 10 microM 5-(N-methyl-N-isobutyl) amiloride or 100 microM 2,2'-(1,2-ethenediyl) bis (5-isothio-cyanatobenzenesulfonic acid), but not by 10 microM benzamil nor 10 microM bumetanide.These data suggest that Na(+)-H(+) exchangers and Cl(-)-HCO(3)(-) exchangers contribute to volume regulation in alpha-cells.

View Article: PubMed Central - PubMed

Affiliation: Faculty of Life Sciences, University of Manchester, Second Floor CTF Building, Manchester, M13 9NT, UK.

ABSTRACT
Ion transport activity in pancreatic alpha-cells was assessed by studying cell volume regulation in response to anisotonic solutions. Cell volume was measured by a video imaging method, and cells were superfused with either 4-(2-hydroxyethyl)piperazine-1-ethanesulfonic acid-buffered or HCO(3)(-) -buffered solutions. alpha-Cells did not exhibit a regulatory volume increase (RVI) in response to cell shrinkage caused by hypertonic solutions. A RVI was observed, however, in cells that had first undergone a regulatory volume decrease (RVD), but only in HCO(3)(-)-buffered solutions. RVI was also observed in response to a HCO(3)(-) -buffered hypertonic solution in which the glucose concentration was increased from 4 to 20 mM. The post-RVD RVI and the glucose-induced RVI were both inhibited by 10 microM 5-(N-methyl-N-isobutyl) amiloride or 100 microM 2,2'-(1,2-ethenediyl) bis (5-isothio-cyanatobenzenesulfonic acid), but not by 10 microM benzamil nor 10 microM bumetanide. These data suggest that Na(+)-H(+) exchangers and Cl(-)-HCO(3)(-) exchangers contribute to volume regulation in alpha-cells.

Show MeSH
Related in: MedlinePlus