Functional analysis of Na+/K+-ATPase isoform distribution in rat ventricular myocytes

Sanda Despa, Donald M Bers

Research output: Contribution to journalArticle

47 Citations (Scopus)

Abstract

The Na+/K+-ATPase (NKA) is the main route for Na + extrusion from cardiac myocytes. Different NKA α-subunit isoforms are present in the heart. NKA-α1 is predominant, although there is a variable amount of NKA-α2 in adult ventricular myocytes of most species. It has been proposed that NKA-α2 is localized mainly in T-tubules (TT), where it could regulate local Na+/Ca2+ exchange and thus cardiac myocyte Ca2+. However, there is controversy as to where NKA-α1 vs. NKA-α2 are localized in ventricular myocytes. Here, we assess the TT vs. external sarcolemma (ESL) distribution functionally using formamide-induced detubulation of rat ventricular myocytes, NKA current (I Pump) measurements and the different ouabain sensitivity of NKA-α1 (low) and NKA-α2 (high) in rat heart. Ouabain-dependent IPump inhibition in control myocytes indicates a high-affinity NKA isoform (NKA-α2, K1/2 = 0.38 ± 0.16 μM) that accounts for 29.5 ± 1.3% of IPump and a low-affinity isoform (NKA-α1, K1/2 = 141 ± 17 μM) that accounts for 70.5% of IPump. Detubulation decreased cell capacitance from 164 ± 6 to 120 ± 8 pF and reduced IPump density from 1.24 ± 0.05 to 1.02 ± 0.05 pA/pF, indicating that the functional density of NKA is significantly higher in TT vs. ESL. In detubulated myocytes, NKA-α2 accounted for only 18.2 ± 1.1% of IPump. Thus, -∼63% of IPump generated by NKA-α2 is from the TT (although TT are only 27% of the total sarcolemma), and the NKA-α2/NKA-α1 ratio in TT is significantly higher than in the ESL. The functional density of NKA-α2 is ∼-4.5 times higher in the T-tubules vs. ESL, whereas NKA-α1 is almost uniformly distributed between the TT and ESL.

Original languageEnglish (US)
JournalAmerican Journal of Physiology - Cell Physiology
Volume293
Issue number1
DOIs
StatePublished - Jul 2007
Externally publishedYes

Fingerprint

Functional analysis
Muscle Cells
Adenosine Triphosphatases
Rats
Protein Isoforms
Sarcolemma
sodium-translocating ATPase
Ouabain
Cardiac Myocytes

Keywords

  • Detubulation
  • External sarcolemma
  • Na/K pump current
  • Ouabain
  • T-tubules

ASJC Scopus subject areas

  • Clinical Biochemistry
  • Cell Biology
  • Physiology

Cite this

Functional analysis of Na+/K+-ATPase isoform distribution in rat ventricular myocytes. / Despa, Sanda; Bers, Donald M.

In: American Journal of Physiology - Cell Physiology, Vol. 293, No. 1, 07.2007.

Research output: Contribution to journalArticle

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abstract = "The Na+/K+-ATPase (NKA) is the main route for Na + extrusion from cardiac myocytes. Different NKA α-subunit isoforms are present in the heart. NKA-α1 is predominant, although there is a variable amount of NKA-α2 in adult ventricular myocytes of most species. It has been proposed that NKA-α2 is localized mainly in T-tubules (TT), where it could regulate local Na+/Ca2+ exchange and thus cardiac myocyte Ca2+. However, there is controversy as to where NKA-α1 vs. NKA-α2 are localized in ventricular myocytes. Here, we assess the TT vs. external sarcolemma (ESL) distribution functionally using formamide-induced detubulation of rat ventricular myocytes, NKA current (I Pump) measurements and the different ouabain sensitivity of NKA-α1 (low) and NKA-α2 (high) in rat heart. Ouabain-dependent IPump inhibition in control myocytes indicates a high-affinity NKA isoform (NKA-α2, K1/2 = 0.38 ± 0.16 μM) that accounts for 29.5 ± 1.3{\%} of IPump and a low-affinity isoform (NKA-α1, K1/2 = 141 ± 17 μM) that accounts for 70.5{\%} of IPump. Detubulation decreased cell capacitance from 164 ± 6 to 120 ± 8 pF and reduced IPump density from 1.24 ± 0.05 to 1.02 ± 0.05 pA/pF, indicating that the functional density of NKA is significantly higher in TT vs. ESL. In detubulated myocytes, NKA-α2 accounted for only 18.2 ± 1.1{\%} of IPump. Thus, -∼63{\%} of IPump generated by NKA-α2 is from the TT (although TT are only 27{\%} of the total sarcolemma), and the NKA-α2/NKA-α1 ratio in TT is significantly higher than in the ESL. The functional density of NKA-α2 is ∼-4.5 times higher in the T-tubules vs. ESL, whereas NKA-α1 is almost uniformly distributed between the TT and ESL.",
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N2 - The Na+/K+-ATPase (NKA) is the main route for Na + extrusion from cardiac myocytes. Different NKA α-subunit isoforms are present in the heart. NKA-α1 is predominant, although there is a variable amount of NKA-α2 in adult ventricular myocytes of most species. It has been proposed that NKA-α2 is localized mainly in T-tubules (TT), where it could regulate local Na+/Ca2+ exchange and thus cardiac myocyte Ca2+. However, there is controversy as to where NKA-α1 vs. NKA-α2 are localized in ventricular myocytes. Here, we assess the TT vs. external sarcolemma (ESL) distribution functionally using formamide-induced detubulation of rat ventricular myocytes, NKA current (I Pump) measurements and the different ouabain sensitivity of NKA-α1 (low) and NKA-α2 (high) in rat heart. Ouabain-dependent IPump inhibition in control myocytes indicates a high-affinity NKA isoform (NKA-α2, K1/2 = 0.38 ± 0.16 μM) that accounts for 29.5 ± 1.3% of IPump and a low-affinity isoform (NKA-α1, K1/2 = 141 ± 17 μM) that accounts for 70.5% of IPump. Detubulation decreased cell capacitance from 164 ± 6 to 120 ± 8 pF and reduced IPump density from 1.24 ± 0.05 to 1.02 ± 0.05 pA/pF, indicating that the functional density of NKA is significantly higher in TT vs. ESL. In detubulated myocytes, NKA-α2 accounted for only 18.2 ± 1.1% of IPump. Thus, -∼63% of IPump generated by NKA-α2 is from the TT (although TT are only 27% of the total sarcolemma), and the NKA-α2/NKA-α1 ratio in TT is significantly higher than in the ESL. The functional density of NKA-α2 is ∼-4.5 times higher in the T-tubules vs. ESL, whereas NKA-α1 is almost uniformly distributed between the TT and ESL.

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KW - Ouabain

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