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Gap junction channels exhibit connexin-specific permeability to cyclic nucleotides.

Kanaporis G, Mese G, Valiuniene L, White TW, Brink PR, Valiunas V - J. Gen. Physiol. (2008)

Bottom Line: However, homotypic Cx40 and homotypic Cx26 exhibited reduced cAMP permeability in comparison to Cx43.These data suggest that Cx43 permeability to cAMP results in a rapid delivery of cAMP from cell to cell in sufficient quantity before degradation by phosphodiesterase to trigger relevant intracellular responses.The data also suggest that the reduced permeability of Cx26 and Cx40 might compromise their ability to deliver cAMP rapidly enough to cause functional changes in a recipient cell.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology and Biophysics, Stony Brook University, Stony Brook, NY 11794, USA.

ABSTRACT
Gap junction channels exhibit connexin dependent biophysical properties, including selective intercellular passage of larger solutes, such as second messengers and siRNA. Here, we report the determination of cyclic nucleotide (cAMP) permeability through gap junction channels composed of Cx43, Cx40, or Cx26 using simultaneous measurements of junctional conductance and intercellular transfer of cAMP. For cAMP detection the recipient cells were transfected with a reporter gene, the cyclic nucleotide-modulated channel from sea urchin sperm (SpIH). cAMP was introduced via patch pipette into the cell of the pair that did not express SpIH. SpIH-derived currents (I(h)) were recorded from the other cell of a pair that expressed SpIH. cAMP diffusion through gap junction channels to the neighboring SpIH-transfected cell resulted in a five to sixfold increase in I(h) current over time. Cyclic AMP transfer was observed for homotypic Cx43 channels over a wide range of conductances. However, homotypic Cx40 and homotypic Cx26 exhibited reduced cAMP permeability in comparison to Cx43. The cAMP/K(+) permeability ratios were 0.18, 0.027, and 0.018 for Cx43, Cx26, and Cx40, respectively. Cx43 channels were approximately 10 to 7 times more permeable to cAMP than Cx40 or Cx26 (Cx43 > Cx26 > or = Cx40), suggesting that these channels have distinctly different selectivity for negatively charged larger solutes involved in metabolic/biochemical coupling. These data suggest that Cx43 permeability to cAMP results in a rapid delivery of cAMP from cell to cell in sufficient quantity before degradation by phosphodiesterase to trigger relevant intracellular responses. The data also suggest that the reduced permeability of Cx26 and Cx40 might compromise their ability to deliver cAMP rapidly enough to cause functional changes in a recipient cell.

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Levitt model plotting D(x)/Do versus pore diameter for cAMP (○) and LY (•), where D(x) is the diffusion coefficient for a solute within the channel and Do is the equivalent within the cytoplasm. D(x)/Do is assumed to be approximated by the calculated flux ratios for cAMP/K+ and LY/K+. The continuous and dashed lines represent cAMP/K and LY/K+ ratios for Cx43 and Cx40, respectively.
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fig9: Levitt model plotting D(x)/Do versus pore diameter for cAMP (○) and LY (•), where D(x) is the diffusion coefficient for a solute within the channel and Do is the equivalent within the cytoplasm. D(x)/Do is assumed to be approximated by the calculated flux ratios for cAMP/K+ and LY/K+. The continuous and dashed lines represent cAMP/K and LY/K+ ratios for Cx43 and Cx40, respectively.

Mentions: Does the data generated in this study illustrate apparent differences in pore diameter for LY and cAMP? Using the same analysis to determine pore diameter for cAMP where its minor diameter is ∼0.52 nm (Hernandez et al., 2007) yields a channel diameter of ∼1.12 nm for Cx43, which is consistent with the value determined by Valiunas et al. (2002) for LY. The calculated pore diameter determined from the cAMP data for Cx40 is not consistent with the value calculated for LY determined by Valiunas et al. (2002). Instead, a calculated pore diameter of ∼0.65 nm for Cx40 is obtained from the cAMP data while a value of ∼1.1 nm is obtained from the LY data. The data for LY and cAMP when applied to the Levitt model (Levitt, 1991b) for Cx26 also generate two different apparent pore diameters, 0.69 nm for cAMP and 1.1 nm for LY. The estimates of pore size determined from the Levitt model (Levitt, 1991b) assume that the ratio D(x)/D(o) is roughly equivalent to the flux ratios for cAMP/K+ or LY/K+ by assuming that the diffusion coefficients for cAMP, LY, and K+ in the cytoplasm are similar. D(x) is the diffusion coefficient of a solute within the pore, and D(o) indicates the diffusion coefficient in cytoplasm. Fig. 9 shows plots for cAMP and LY derived from the Levitt equation (Levitt, 1991b) where the ordinate is the ratio D(x)/D(o) and the abscissa is diameter of the pore. As the minor diameter of the solute and the diameter of the pore approximate each other, the diffusion coefficient within the channel declines. Table II also includes the dimensions of the solutes tested and the calculated pore size from the Levitt equation based on the cAMP/K+ or LY/K+ ratios.


Gap junction channels exhibit connexin-specific permeability to cyclic nucleotides.

Kanaporis G, Mese G, Valiuniene L, White TW, Brink PR, Valiunas V - J. Gen. Physiol. (2008)

Levitt model plotting D(x)/Do versus pore diameter for cAMP (○) and LY (•), where D(x) is the diffusion coefficient for a solute within the channel and Do is the equivalent within the cytoplasm. D(x)/Do is assumed to be approximated by the calculated flux ratios for cAMP/K+ and LY/K+. The continuous and dashed lines represent cAMP/K and LY/K+ ratios for Cx43 and Cx40, respectively.
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Related In: Results  -  Collection

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fig9: Levitt model plotting D(x)/Do versus pore diameter for cAMP (○) and LY (•), where D(x) is the diffusion coefficient for a solute within the channel and Do is the equivalent within the cytoplasm. D(x)/Do is assumed to be approximated by the calculated flux ratios for cAMP/K+ and LY/K+. The continuous and dashed lines represent cAMP/K and LY/K+ ratios for Cx43 and Cx40, respectively.
Mentions: Does the data generated in this study illustrate apparent differences in pore diameter for LY and cAMP? Using the same analysis to determine pore diameter for cAMP where its minor diameter is ∼0.52 nm (Hernandez et al., 2007) yields a channel diameter of ∼1.12 nm for Cx43, which is consistent with the value determined by Valiunas et al. (2002) for LY. The calculated pore diameter determined from the cAMP data for Cx40 is not consistent with the value calculated for LY determined by Valiunas et al. (2002). Instead, a calculated pore diameter of ∼0.65 nm for Cx40 is obtained from the cAMP data while a value of ∼1.1 nm is obtained from the LY data. The data for LY and cAMP when applied to the Levitt model (Levitt, 1991b) for Cx26 also generate two different apparent pore diameters, 0.69 nm for cAMP and 1.1 nm for LY. The estimates of pore size determined from the Levitt model (Levitt, 1991b) assume that the ratio D(x)/D(o) is roughly equivalent to the flux ratios for cAMP/K+ or LY/K+ by assuming that the diffusion coefficients for cAMP, LY, and K+ in the cytoplasm are similar. D(x) is the diffusion coefficient of a solute within the pore, and D(o) indicates the diffusion coefficient in cytoplasm. Fig. 9 shows plots for cAMP and LY derived from the Levitt equation (Levitt, 1991b) where the ordinate is the ratio D(x)/D(o) and the abscissa is diameter of the pore. As the minor diameter of the solute and the diameter of the pore approximate each other, the diffusion coefficient within the channel declines. Table II also includes the dimensions of the solutes tested and the calculated pore size from the Levitt equation based on the cAMP/K+ or LY/K+ ratios.

Bottom Line: However, homotypic Cx40 and homotypic Cx26 exhibited reduced cAMP permeability in comparison to Cx43.These data suggest that Cx43 permeability to cAMP results in a rapid delivery of cAMP from cell to cell in sufficient quantity before degradation by phosphodiesterase to trigger relevant intracellular responses.The data also suggest that the reduced permeability of Cx26 and Cx40 might compromise their ability to deliver cAMP rapidly enough to cause functional changes in a recipient cell.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology and Biophysics, Stony Brook University, Stony Brook, NY 11794, USA.

ABSTRACT
Gap junction channels exhibit connexin dependent biophysical properties, including selective intercellular passage of larger solutes, such as second messengers and siRNA. Here, we report the determination of cyclic nucleotide (cAMP) permeability through gap junction channels composed of Cx43, Cx40, or Cx26 using simultaneous measurements of junctional conductance and intercellular transfer of cAMP. For cAMP detection the recipient cells were transfected with a reporter gene, the cyclic nucleotide-modulated channel from sea urchin sperm (SpIH). cAMP was introduced via patch pipette into the cell of the pair that did not express SpIH. SpIH-derived currents (I(h)) were recorded from the other cell of a pair that expressed SpIH. cAMP diffusion through gap junction channels to the neighboring SpIH-transfected cell resulted in a five to sixfold increase in I(h) current over time. Cyclic AMP transfer was observed for homotypic Cx43 channels over a wide range of conductances. However, homotypic Cx40 and homotypic Cx26 exhibited reduced cAMP permeability in comparison to Cx43. The cAMP/K(+) permeability ratios were 0.18, 0.027, and 0.018 for Cx43, Cx26, and Cx40, respectively. Cx43 channels were approximately 10 to 7 times more permeable to cAMP than Cx40 or Cx26 (Cx43 > Cx26 > or = Cx40), suggesting that these channels have distinctly different selectivity for negatively charged larger solutes involved in metabolic/biochemical coupling. These data suggest that Cx43 permeability to cAMP results in a rapid delivery of cAMP from cell to cell in sufficient quantity before degradation by phosphodiesterase to trigger relevant intracellular responses. The data also suggest that the reduced permeability of Cx26 and Cx40 might compromise their ability to deliver cAMP rapidly enough to cause functional changes in a recipient cell.

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