Ouabain sensitive ion fluxes in the smooth muscle of the guinea pig's taenia coli

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Abstract

Tissues with raised intracellular Na levels, produced by incubation in K free media, were used throughout. The uptake of 42K by these Na loaded tissues was followed for 10 min in the presence and absence of 1.37 x 10-4M ouabain, this being sufficient to inhibit Na pumping maximally. Subtraction of the uptake seen in the presence from that seen in the absence of ouabain gave estimates of the pumped ouabain sensitive uptake. In Na free (MgCl2) medium this depended on the [K]0 in a sigmoidal fashion with a half maximal [K]0 for activation of some 4 mM. The maximal uptake of K was 3 m mole/kg.min corresponding to a transmembrane flux of some 12.5 p mole.cm-2.sec-1. In the presence of Na the K activation curve became more obviously sigmoid and higher concentrations of K were needed to achieve a given active K influx. The results were well fitted by assuming that Na and K competed for two identical, non interacting sites on the external pump face. Addition of K during the efflux of 24Na into a Na free (MgCl2) medium led to an increased rate of tracer loss. The magnitude of this increase depended on the [K] used in a hyperbolic fashion and it was abolished by addition of ouabain. The [K] causing half maximal activation of ouabain sensitive Na efflux was in the order of 1-2 mM. When the [K] in the uptake media was 1.5 mM, Na, Li, Rb and Cs all inhibited ouabain sensitive K uptake the order of effectiveness being Rb > Cs > Na > Li. With a [K]0 of 0.15 mM low concentrations of Cs and Rb were shown to stimulate uptake. Such an effect is predicted by assuming two ion binding sites on the pump's outer face, and that the pump can translocate mixtures of K and either Rb or Cs. As well as K, Rb, Cs and Li were all shown to stimulate ouabain sensitive 24Na efflux, the relative potencies being in the order K ∞ Rb > Cs > Li. External Na stimulated 24Na efflux but in an ouabain insensitive fashion. Li also showed ouabain insensitive exchange with internal Na. Na loaded tissues were placed into normal Krebs solution or into similar solutions in which Cs, Rb or Li replaced the K, and the changes in ion content followed. Tissues placed in K- or Rb- Krebs solution lost Na and gained equal amounts of the other ion, complete recovery to normal Na levels occurring after about 1.5 hr. When placed in Cs Krebs solution Na was lost and Cs gained but at a slower rate. Even after 5.5 hr in Krebs solution no more than a small passive uptake of Li occurred. The ouabain sensitive uptakes of 82Rb and 134Cs were determined in the same manner as for 42K. The activation curves for these ions were hyperbolic. The half maximal concentrations for ouabain sensitive uptake of the three ions were in the order K ∞ Rb > Cs. The maximal influxes were in the order K > Rb > Cs, this being the same order as for the passive uptakes of these ions in the presence of 1.37 x 10-4M ouabain. It is concluded that the Na pump in smooth muscle is similar to that of other tissues. It appears to have two external activation sites, is activated by external Rb, K, Cs and Li, in the potency sequence K - Rb > Cs > Li, and is inhibited by external Na.

Original languageEnglish (US)
Pages (from-to)235-254
Number of pages20
JournalJournal of Physiology
Volume266
Issue number2
StatePublished - 1977
Externally publishedYes

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Ouabain
Smooth Muscle
Guinea Pigs
Colon
Ions
Magnesium Chloride
Sigmoid Colon
Binding Sites

ASJC Scopus subject areas

  • Physiology

Cite this

Ouabain sensitive ion fluxes in the smooth muscle of the guinea pig's taenia coli. / Widdicombe, Jonathan.

In: Journal of Physiology, Vol. 266, No. 2, 1977, p. 235-254.

Research output: Contribution to journalArticle

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title = "Ouabain sensitive ion fluxes in the smooth muscle of the guinea pig's taenia coli",
abstract = "Tissues with raised intracellular Na levels, produced by incubation in K free media, were used throughout. The uptake of 42K by these Na loaded tissues was followed for 10 min in the presence and absence of 1.37 x 10-4M ouabain, this being sufficient to inhibit Na pumping maximally. Subtraction of the uptake seen in the presence from that seen in the absence of ouabain gave estimates of the pumped ouabain sensitive uptake. In Na free (MgCl2) medium this depended on the [K]0 in a sigmoidal fashion with a half maximal [K]0 for activation of some 4 mM. The maximal uptake of K was 3 m mole/kg.min corresponding to a transmembrane flux of some 12.5 p mole.cm-2.sec-1. In the presence of Na the K activation curve became more obviously sigmoid and higher concentrations of K were needed to achieve a given active K influx. The results were well fitted by assuming that Na and K competed for two identical, non interacting sites on the external pump face. Addition of K during the efflux of 24Na into a Na free (MgCl2) medium led to an increased rate of tracer loss. The magnitude of this increase depended on the [K] used in a hyperbolic fashion and it was abolished by addition of ouabain. The [K] causing half maximal activation of ouabain sensitive Na efflux was in the order of 1-2 mM. When the [K] in the uptake media was 1.5 mM, Na, Li, Rb and Cs all inhibited ouabain sensitive K uptake the order of effectiveness being Rb > Cs > Na > Li. With a [K]0 of 0.15 mM low concentrations of Cs and Rb were shown to stimulate uptake. Such an effect is predicted by assuming two ion binding sites on the pump's outer face, and that the pump can translocate mixtures of K and either Rb or Cs. As well as K, Rb, Cs and Li were all shown to stimulate ouabain sensitive 24Na efflux, the relative potencies being in the order K ∞ Rb > Cs > Li. External Na stimulated 24Na efflux but in an ouabain insensitive fashion. Li also showed ouabain insensitive exchange with internal Na. Na loaded tissues were placed into normal Krebs solution or into similar solutions in which Cs, Rb or Li replaced the K, and the changes in ion content followed. Tissues placed in K- or Rb- Krebs solution lost Na and gained equal amounts of the other ion, complete recovery to normal Na levels occurring after about 1.5 hr. When placed in Cs Krebs solution Na was lost and Cs gained but at a slower rate. Even after 5.5 hr in Krebs solution no more than a small passive uptake of Li occurred. The ouabain sensitive uptakes of 82Rb and 134Cs were determined in the same manner as for 42K. The activation curves for these ions were hyperbolic. The half maximal concentrations for ouabain sensitive uptake of the three ions were in the order K ∞ Rb > Cs. The maximal influxes were in the order K > Rb > Cs, this being the same order as for the passive uptakes of these ions in the presence of 1.37 x 10-4M ouabain. It is concluded that the Na pump in smooth muscle is similar to that of other tissues. It appears to have two external activation sites, is activated by external Rb, K, Cs and Li, in the potency sequence K - Rb > Cs > Li, and is inhibited by external Na.",
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N2 - Tissues with raised intracellular Na levels, produced by incubation in K free media, were used throughout. The uptake of 42K by these Na loaded tissues was followed for 10 min in the presence and absence of 1.37 x 10-4M ouabain, this being sufficient to inhibit Na pumping maximally. Subtraction of the uptake seen in the presence from that seen in the absence of ouabain gave estimates of the pumped ouabain sensitive uptake. In Na free (MgCl2) medium this depended on the [K]0 in a sigmoidal fashion with a half maximal [K]0 for activation of some 4 mM. The maximal uptake of K was 3 m mole/kg.min corresponding to a transmembrane flux of some 12.5 p mole.cm-2.sec-1. In the presence of Na the K activation curve became more obviously sigmoid and higher concentrations of K were needed to achieve a given active K influx. The results were well fitted by assuming that Na and K competed for two identical, non interacting sites on the external pump face. Addition of K during the efflux of 24Na into a Na free (MgCl2) medium led to an increased rate of tracer loss. The magnitude of this increase depended on the [K] used in a hyperbolic fashion and it was abolished by addition of ouabain. The [K] causing half maximal activation of ouabain sensitive Na efflux was in the order of 1-2 mM. When the [K] in the uptake media was 1.5 mM, Na, Li, Rb and Cs all inhibited ouabain sensitive K uptake the order of effectiveness being Rb > Cs > Na > Li. With a [K]0 of 0.15 mM low concentrations of Cs and Rb were shown to stimulate uptake. Such an effect is predicted by assuming two ion binding sites on the pump's outer face, and that the pump can translocate mixtures of K and either Rb or Cs. As well as K, Rb, Cs and Li were all shown to stimulate ouabain sensitive 24Na efflux, the relative potencies being in the order K ∞ Rb > Cs > Li. External Na stimulated 24Na efflux but in an ouabain insensitive fashion. Li also showed ouabain insensitive exchange with internal Na. Na loaded tissues were placed into normal Krebs solution or into similar solutions in which Cs, Rb or Li replaced the K, and the changes in ion content followed. Tissues placed in K- or Rb- Krebs solution lost Na and gained equal amounts of the other ion, complete recovery to normal Na levels occurring after about 1.5 hr. When placed in Cs Krebs solution Na was lost and Cs gained but at a slower rate. Even after 5.5 hr in Krebs solution no more than a small passive uptake of Li occurred. The ouabain sensitive uptakes of 82Rb and 134Cs were determined in the same manner as for 42K. The activation curves for these ions were hyperbolic. The half maximal concentrations for ouabain sensitive uptake of the three ions were in the order K ∞ Rb > Cs. The maximal influxes were in the order K > Rb > Cs, this being the same order as for the passive uptakes of these ions in the presence of 1.37 x 10-4M ouabain. It is concluded that the Na pump in smooth muscle is similar to that of other tissues. It appears to have two external activation sites, is activated by external Rb, K, Cs and Li, in the potency sequence K - Rb > Cs > Li, and is inhibited by external Na.

AB - Tissues with raised intracellular Na levels, produced by incubation in K free media, were used throughout. The uptake of 42K by these Na loaded tissues was followed for 10 min in the presence and absence of 1.37 x 10-4M ouabain, this being sufficient to inhibit Na pumping maximally. Subtraction of the uptake seen in the presence from that seen in the absence of ouabain gave estimates of the pumped ouabain sensitive uptake. In Na free (MgCl2) medium this depended on the [K]0 in a sigmoidal fashion with a half maximal [K]0 for activation of some 4 mM. The maximal uptake of K was 3 m mole/kg.min corresponding to a transmembrane flux of some 12.5 p mole.cm-2.sec-1. In the presence of Na the K activation curve became more obviously sigmoid and higher concentrations of K were needed to achieve a given active K influx. The results were well fitted by assuming that Na and K competed for two identical, non interacting sites on the external pump face. Addition of K during the efflux of 24Na into a Na free (MgCl2) medium led to an increased rate of tracer loss. The magnitude of this increase depended on the [K] used in a hyperbolic fashion and it was abolished by addition of ouabain. The [K] causing half maximal activation of ouabain sensitive Na efflux was in the order of 1-2 mM. When the [K] in the uptake media was 1.5 mM, Na, Li, Rb and Cs all inhibited ouabain sensitive K uptake the order of effectiveness being Rb > Cs > Na > Li. With a [K]0 of 0.15 mM low concentrations of Cs and Rb were shown to stimulate uptake. Such an effect is predicted by assuming two ion binding sites on the pump's outer face, and that the pump can translocate mixtures of K and either Rb or Cs. As well as K, Rb, Cs and Li were all shown to stimulate ouabain sensitive 24Na efflux, the relative potencies being in the order K ∞ Rb > Cs > Li. External Na stimulated 24Na efflux but in an ouabain insensitive fashion. Li also showed ouabain insensitive exchange with internal Na. Na loaded tissues were placed into normal Krebs solution or into similar solutions in which Cs, Rb or Li replaced the K, and the changes in ion content followed. Tissues placed in K- or Rb- Krebs solution lost Na and gained equal amounts of the other ion, complete recovery to normal Na levels occurring after about 1.5 hr. When placed in Cs Krebs solution Na was lost and Cs gained but at a slower rate. Even after 5.5 hr in Krebs solution no more than a small passive uptake of Li occurred. The ouabain sensitive uptakes of 82Rb and 134Cs were determined in the same manner as for 42K. The activation curves for these ions were hyperbolic. The half maximal concentrations for ouabain sensitive uptake of the three ions were in the order K ∞ Rb > Cs. The maximal influxes were in the order K > Rb > Cs, this being the same order as for the passive uptakes of these ions in the presence of 1.37 x 10-4M ouabain. It is concluded that the Na pump in smooth muscle is similar to that of other tissues. It appears to have two external activation sites, is activated by external Rb, K, Cs and Li, in the potency sequence K - Rb > Cs > Li, and is inhibited by external Na.

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