Differential distribution and regulation of mouse cardiac Na+/K+-ATPase α1 and α2 subunits in T-tubule and surface sarcolemmal membranes

Roger G. Berry, Sanda Despa, William Fuller, Donald M Bers, Michael J. Shattock

Research output: Contribution to journalArticle

69 Citations (Scopus)

Abstract

Objectives: Two Na+/K+-ATPase (NKA) α-subunit isoforms, α1 and α2, are expressed in the adult mouse heart. The subcellular distribution of these isoforms in T-tubule and surface sarcolemmal (SSL) membranes and their regulation by cAMP-dependent protein kinase (PKA) is unclear. Methods: We used formamide-induced detubulation of mouse ventricular myocytes to investigate differential functional distribution and regulation by PKA of α1 and α2 in T-tubule versus SSL membranes by measuring NKA current (Ipump) and NKA-mediated Na+ efflux (- d[Na]i/dt). Results: Ipump is composed of 88% α1-mediated Ipump (Iα1) and 12% α2-mediated Ipump (Iα2). α1 and α2 subunits demonstrate distinct ouabain affinities (105 ± 6 and 0.3 ± 0.1 μmol/L respectively) but similar affinity for intracellular Na+ (K1/2Na+ of 16.6 ± 0.8 and 16.7±2.6 mmol/L respectively). Detubulation reduced (i) Ipump density (1.42 ± 0.1 to 1.20 ± 0.04 pA/pF), (ii) cell capacitance (181 ± 12 to 127±17 pF), and (iii) Iα2 contribution (12 to 6%). Total Ipump density was ∼ 60% higher in T-tubule (1.94 pA/pF, derived) vs. SSL membranes. Although T-tubule membranes represent only 30% of total surface area, they generate ∼ 70% of Iα2 and ∼ 37% of Iα1. Iα1 density was substantially higher than Iα2 in SSL (Iα1:Iα2 = 16:1) but this was markedly reduced in T-tubules (4:1). In addition to differential localisation, isoprenaline (ISO, 1 μmol/L) significantly increased α1-mediated NKA Na+ affinity (from 16.6 ± 0.8 to 13.3 ± 1.4 mmol/L) and caused a small increase in maximal NKA Na+ efflux rate. ISO had no effect on α2-mediated NKA activity. Conclusion: These data suggest that NKA α1 and α2 subunits are differentially localised and regulated by PKA in T-tubule and SSL membranes and may have distinct regulatory roles in cardiac excitation-contraction coupling.

Original languageEnglish (US)
Pages (from-to)92-100
Number of pages9
JournalCardiovascular Research
Volume73
Issue number1
DOIs
StatePublished - Jan 1 2007
Externally publishedYes

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Membranes
Protein Kinases
Protein Isoforms
Excitation Contraction Coupling
Ouabain
Cyclic AMP-Dependent Protein Kinases
sodium-translocating ATPase
Isoproterenol
Muscle Cells

Keywords

  • Adrenergic agonist
  • Ion pumps
  • Na/K-pump
  • Protein kinase A
  • Sarcolemmal

ASJC Scopus subject areas

  • Cardiology and Cardiovascular Medicine

Cite this

Differential distribution and regulation of mouse cardiac Na+/K+-ATPase α1 and α2 subunits in T-tubule and surface sarcolemmal membranes. / Berry, Roger G.; Despa, Sanda; Fuller, William; Bers, Donald M; Shattock, Michael J.

In: Cardiovascular Research, Vol. 73, No. 1, 01.01.2007, p. 92-100.

Research output: Contribution to journalArticle

@article{049d55ba291f47a18fa0782edd5c9240,
title = "Differential distribution and regulation of mouse cardiac Na+/K+-ATPase α1 and α2 subunits in T-tubule and surface sarcolemmal membranes",
abstract = "Objectives: Two Na+/K+-ATPase (NKA) α-subunit isoforms, α1 and α2, are expressed in the adult mouse heart. The subcellular distribution of these isoforms in T-tubule and surface sarcolemmal (SSL) membranes and their regulation by cAMP-dependent protein kinase (PKA) is unclear. Methods: We used formamide-induced detubulation of mouse ventricular myocytes to investigate differential functional distribution and regulation by PKA of α1 and α2 in T-tubule versus SSL membranes by measuring NKA current (Ipump) and NKA-mediated Na+ efflux (- d[Na]i/dt). Results: Ipump is composed of 88{\%} α1-mediated Ipump (Iα1) and 12{\%} α2-mediated Ipump (Iα2). α1 and α2 subunits demonstrate distinct ouabain affinities (105 ± 6 and 0.3 ± 0.1 μmol/L respectively) but similar affinity for intracellular Na+ (K1/2Na+ of 16.6 ± 0.8 and 16.7±2.6 mmol/L respectively). Detubulation reduced (i) Ipump density (1.42 ± 0.1 to 1.20 ± 0.04 pA/pF), (ii) cell capacitance (181 ± 12 to 127±17 pF), and (iii) Iα2 contribution (12 to 6{\%}). Total Ipump density was ∼ 60{\%} higher in T-tubule (1.94 pA/pF, derived) vs. SSL membranes. Although T-tubule membranes represent only 30{\%} of total surface area, they generate ∼ 70{\%} of Iα2 and ∼ 37{\%} of Iα1. Iα1 density was substantially higher than Iα2 in SSL (Iα1:Iα2 = 16:1) but this was markedly reduced in T-tubules (4:1). In addition to differential localisation, isoprenaline (ISO, 1 μmol/L) significantly increased α1-mediated NKA Na+ affinity (from 16.6 ± 0.8 to 13.3 ± 1.4 mmol/L) and caused a small increase in maximal NKA Na+ efflux rate. ISO had no effect on α2-mediated NKA activity. Conclusion: These data suggest that NKA α1 and α2 subunits are differentially localised and regulated by PKA in T-tubule and SSL membranes and may have distinct regulatory roles in cardiac excitation-contraction coupling.",
keywords = "Adrenergic agonist, Ion pumps, Na/K-pump, Protein kinase A, Sarcolemmal",
author = "Berry, {Roger G.} and Sanda Despa and William Fuller and Bers, {Donald M} and Shattock, {Michael J.}",
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T1 - Differential distribution and regulation of mouse cardiac Na+/K+-ATPase α1 and α2 subunits in T-tubule and surface sarcolemmal membranes

AU - Berry, Roger G.

AU - Despa, Sanda

AU - Fuller, William

AU - Bers, Donald M

AU - Shattock, Michael J.

PY - 2007/1/1

Y1 - 2007/1/1

N2 - Objectives: Two Na+/K+-ATPase (NKA) α-subunit isoforms, α1 and α2, are expressed in the adult mouse heart. The subcellular distribution of these isoforms in T-tubule and surface sarcolemmal (SSL) membranes and their regulation by cAMP-dependent protein kinase (PKA) is unclear. Methods: We used formamide-induced detubulation of mouse ventricular myocytes to investigate differential functional distribution and regulation by PKA of α1 and α2 in T-tubule versus SSL membranes by measuring NKA current (Ipump) and NKA-mediated Na+ efflux (- d[Na]i/dt). Results: Ipump is composed of 88% α1-mediated Ipump (Iα1) and 12% α2-mediated Ipump (Iα2). α1 and α2 subunits demonstrate distinct ouabain affinities (105 ± 6 and 0.3 ± 0.1 μmol/L respectively) but similar affinity for intracellular Na+ (K1/2Na+ of 16.6 ± 0.8 and 16.7±2.6 mmol/L respectively). Detubulation reduced (i) Ipump density (1.42 ± 0.1 to 1.20 ± 0.04 pA/pF), (ii) cell capacitance (181 ± 12 to 127±17 pF), and (iii) Iα2 contribution (12 to 6%). Total Ipump density was ∼ 60% higher in T-tubule (1.94 pA/pF, derived) vs. SSL membranes. Although T-tubule membranes represent only 30% of total surface area, they generate ∼ 70% of Iα2 and ∼ 37% of Iα1. Iα1 density was substantially higher than Iα2 in SSL (Iα1:Iα2 = 16:1) but this was markedly reduced in T-tubules (4:1). In addition to differential localisation, isoprenaline (ISO, 1 μmol/L) significantly increased α1-mediated NKA Na+ affinity (from 16.6 ± 0.8 to 13.3 ± 1.4 mmol/L) and caused a small increase in maximal NKA Na+ efflux rate. ISO had no effect on α2-mediated NKA activity. Conclusion: These data suggest that NKA α1 and α2 subunits are differentially localised and regulated by PKA in T-tubule and SSL membranes and may have distinct regulatory roles in cardiac excitation-contraction coupling.

AB - Objectives: Two Na+/K+-ATPase (NKA) α-subunit isoforms, α1 and α2, are expressed in the adult mouse heart. The subcellular distribution of these isoforms in T-tubule and surface sarcolemmal (SSL) membranes and their regulation by cAMP-dependent protein kinase (PKA) is unclear. Methods: We used formamide-induced detubulation of mouse ventricular myocytes to investigate differential functional distribution and regulation by PKA of α1 and α2 in T-tubule versus SSL membranes by measuring NKA current (Ipump) and NKA-mediated Na+ efflux (- d[Na]i/dt). Results: Ipump is composed of 88% α1-mediated Ipump (Iα1) and 12% α2-mediated Ipump (Iα2). α1 and α2 subunits demonstrate distinct ouabain affinities (105 ± 6 and 0.3 ± 0.1 μmol/L respectively) but similar affinity for intracellular Na+ (K1/2Na+ of 16.6 ± 0.8 and 16.7±2.6 mmol/L respectively). Detubulation reduced (i) Ipump density (1.42 ± 0.1 to 1.20 ± 0.04 pA/pF), (ii) cell capacitance (181 ± 12 to 127±17 pF), and (iii) Iα2 contribution (12 to 6%). Total Ipump density was ∼ 60% higher in T-tubule (1.94 pA/pF, derived) vs. SSL membranes. Although T-tubule membranes represent only 30% of total surface area, they generate ∼ 70% of Iα2 and ∼ 37% of Iα1. Iα1 density was substantially higher than Iα2 in SSL (Iα1:Iα2 = 16:1) but this was markedly reduced in T-tubules (4:1). In addition to differential localisation, isoprenaline (ISO, 1 μmol/L) significantly increased α1-mediated NKA Na+ affinity (from 16.6 ± 0.8 to 13.3 ± 1.4 mmol/L) and caused a small increase in maximal NKA Na+ efflux rate. ISO had no effect on α2-mediated NKA activity. Conclusion: These data suggest that NKA α1 and α2 subunits are differentially localised and regulated by PKA in T-tubule and SSL membranes and may have distinct regulatory roles in cardiac excitation-contraction coupling.

KW - Adrenergic agonist

KW - Ion pumps

KW - Na/K-pump

KW - Protein kinase A

KW - Sarcolemmal

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