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Extracellular charge adsorption influences intracellular electrochemical homeostasis in amphibian skeletal muscle.

Mehta AR, Huang CL, Skepper JN, Fraser JA - Biophys. J. (2008)

Bottom Line: The membrane potential measured by intracellular electrodes, E(m), is the sum of the transmembrane potential difference (E(1)) between inner and outer cell membrane surfaces and a smaller potential difference (E(2)) between a volume containing fixed charges on or near the outer membrane surface and the bulk extracellular space.First, analytic equations were developed to calculate the influence of charges constrained within a three-dimensional glycocalyceal matrix enveloping the cell membrane outer surface upon local electrical potentials and ion concentrations.Electron microscopy confirmed predictions of these equations that extracellular charge adsorption influences glycocalyceal volume.

View Article: PubMed Central - PubMed

Affiliation: Physiological Laboratory, University of Cambridge, Cambridge, United Kingdom.

ABSTRACT
The membrane potential measured by intracellular electrodes, E(m), is the sum of the transmembrane potential difference (E(1)) between inner and outer cell membrane surfaces and a smaller potential difference (E(2)) between a volume containing fixed charges on or near the outer membrane surface and the bulk extracellular space. This study investigates the influence of E(2) upon transmembrane ion fluxes, and hence cellular electrochemical homeostasis, using an integrative approach that combines computational and experimental methods. First, analytic equations were developed to calculate the influence of charges constrained within a three-dimensional glycocalyceal matrix enveloping the cell membrane outer surface upon local electrical potentials and ion concentrations. Electron microscopy confirmed predictions of these equations that extracellular charge adsorption influences glycocalyceal volume. Second, the novel analytic glycocalyx formulation was incorporated into the charge-difference cellular model of Fraser and Huang to simulate the influence of extracellular fixed charges upon intracellular ionic homeostasis. Experimental measurements of E(m) supported the resulting predictions that an increased magnitude of extracellular fixed charge increases net transmembrane ionic leak currents, resulting in either a compensatory increase in Na(+)/K(+)-ATPase activity, or, in cells with reduced Na(+)/K(+)-ATPase activity, a partial dissipation of transmembrane ionic gradients and depolarization of E(m).

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The influence of extracellular [Ca2+] upon glycocalyceal thickness. Glycocalyceal thicknesses were measured from electron micrographs of fixed cutaneous pectoris muscles as described in Methods. Pairs of muscles obtained from the same frog then exposed to different experimental conditions are depicted with similar shading. Two muscles from different frogs were exposed to each extracellular Ca2+ concentration. There was no significant difference between glycocalyceal thicknesses within each condition (p > 0.05 in each case). In contrast, both increases (to 5 mM) and decreases (to nominally zero) from normal (1.8 mM) [Ca2+]e produced significant (p < 0.01) increases in glycocalyceal thickness. Thus, variation between conditions was significant even with muscles from the same animal, but there was no significant variation within conditions even with muscles obtained from different animals.
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fig5: The influence of extracellular [Ca2+] upon glycocalyceal thickness. Glycocalyceal thicknesses were measured from electron micrographs of fixed cutaneous pectoris muscles as described in Methods. Pairs of muscles obtained from the same frog then exposed to different experimental conditions are depicted with similar shading. Two muscles from different frogs were exposed to each extracellular Ca2+ concentration. There was no significant difference between glycocalyceal thicknesses within each condition (p > 0.05 in each case). In contrast, both increases (to 5 mM) and decreases (to nominally zero) from normal (1.8 mM) [Ca2+]e produced significant (p < 0.01) increases in glycocalyceal thickness. Thus, variation between conditions was significant even with muscles from the same animal, but there was no significant variation within conditions even with muscles obtained from different animals.

Mentions: Electron micrographic analyses of glycocalyceal thicknesses after alterations in extracellular Ca2+ were employed to test this hypothesis that glycocalyceal charge and volume are related. Fig. 5 demonstrates the influence of extracellular [Ca2+] upon glycocalyceal volume. Pairs of cutaneous pectoris muscles obtained from different frogs were exposed to each condition of [Ca2+]e. This demonstrated that there was no significant variation in glycocalyceal thickness, or in the relationship of this thickness to [Ca2+]e, between muscle fibers from different frogs (p > 0.05 in each case). In contrast, pairs of cutaneous pectoris muscles obtained from the same frogs were exposed to different conditions of [Ca2+]e and demonstrated increased glycocalyceal thicknesses after both increases (to 5 mM) and decreases (to nominally zero) in [Ca2+]e.


Extracellular charge adsorption influences intracellular electrochemical homeostasis in amphibian skeletal muscle.

Mehta AR, Huang CL, Skepper JN, Fraser JA - Biophys. J. (2008)

The influence of extracellular [Ca2+] upon glycocalyceal thickness. Glycocalyceal thicknesses were measured from electron micrographs of fixed cutaneous pectoris muscles as described in Methods. Pairs of muscles obtained from the same frog then exposed to different experimental conditions are depicted with similar shading. Two muscles from different frogs were exposed to each extracellular Ca2+ concentration. There was no significant difference between glycocalyceal thicknesses within each condition (p > 0.05 in each case). In contrast, both increases (to 5 mM) and decreases (to nominally zero) from normal (1.8 mM) [Ca2+]e produced significant (p < 0.01) increases in glycocalyceal thickness. Thus, variation between conditions was significant even with muscles from the same animal, but there was no significant variation within conditions even with muscles obtained from different animals.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2480687&req=5

fig5: The influence of extracellular [Ca2+] upon glycocalyceal thickness. Glycocalyceal thicknesses were measured from electron micrographs of fixed cutaneous pectoris muscles as described in Methods. Pairs of muscles obtained from the same frog then exposed to different experimental conditions are depicted with similar shading. Two muscles from different frogs were exposed to each extracellular Ca2+ concentration. There was no significant difference between glycocalyceal thicknesses within each condition (p > 0.05 in each case). In contrast, both increases (to 5 mM) and decreases (to nominally zero) from normal (1.8 mM) [Ca2+]e produced significant (p < 0.01) increases in glycocalyceal thickness. Thus, variation between conditions was significant even with muscles from the same animal, but there was no significant variation within conditions even with muscles obtained from different animals.
Mentions: Electron micrographic analyses of glycocalyceal thicknesses after alterations in extracellular Ca2+ were employed to test this hypothesis that glycocalyceal charge and volume are related. Fig. 5 demonstrates the influence of extracellular [Ca2+] upon glycocalyceal volume. Pairs of cutaneous pectoris muscles obtained from different frogs were exposed to each condition of [Ca2+]e. This demonstrated that there was no significant variation in glycocalyceal thickness, or in the relationship of this thickness to [Ca2+]e, between muscle fibers from different frogs (p > 0.05 in each case). In contrast, pairs of cutaneous pectoris muscles obtained from the same frogs were exposed to different conditions of [Ca2+]e and demonstrated increased glycocalyceal thicknesses after both increases (to 5 mM) and decreases (to nominally zero) in [Ca2+]e.

Bottom Line: The membrane potential measured by intracellular electrodes, E(m), is the sum of the transmembrane potential difference (E(1)) between inner and outer cell membrane surfaces and a smaller potential difference (E(2)) between a volume containing fixed charges on or near the outer membrane surface and the bulk extracellular space.First, analytic equations were developed to calculate the influence of charges constrained within a three-dimensional glycocalyceal matrix enveloping the cell membrane outer surface upon local electrical potentials and ion concentrations.Electron microscopy confirmed predictions of these equations that extracellular charge adsorption influences glycocalyceal volume.

View Article: PubMed Central - PubMed

Affiliation: Physiological Laboratory, University of Cambridge, Cambridge, United Kingdom.

ABSTRACT
The membrane potential measured by intracellular electrodes, E(m), is the sum of the transmembrane potential difference (E(1)) between inner and outer cell membrane surfaces and a smaller potential difference (E(2)) between a volume containing fixed charges on or near the outer membrane surface and the bulk extracellular space. This study investigates the influence of E(2) upon transmembrane ion fluxes, and hence cellular electrochemical homeostasis, using an integrative approach that combines computational and experimental methods. First, analytic equations were developed to calculate the influence of charges constrained within a three-dimensional glycocalyceal matrix enveloping the cell membrane outer surface upon local electrical potentials and ion concentrations. Electron microscopy confirmed predictions of these equations that extracellular charge adsorption influences glycocalyceal volume. Second, the novel analytic glycocalyx formulation was incorporated into the charge-difference cellular model of Fraser and Huang to simulate the influence of extracellular fixed charges upon intracellular ionic homeostasis. Experimental measurements of E(m) supported the resulting predictions that an increased magnitude of extracellular fixed charge increases net transmembrane ionic leak currents, resulting in either a compensatory increase in Na(+)/K(+)-ATPase activity, or, in cells with reduced Na(+)/K(+)-ATPase activity, a partial dissipation of transmembrane ionic gradients and depolarization of E(m).

Show MeSH
Related in: MedlinePlus