Different effects of α- and β-adrenergic stimulation on cytosolic pH and myofilament responsiveness to Ca2+ in cardiac myocytes

Giovanni Gambassi, Harold A. Spurgeon, Edward G. Lakatta, Paul S. Blank, Maurizio C. Capogrossi

Research output: Contribution to journalArticle

Abstract

α-Adrenergic stimulation (α-AS) and β-adrenergic stimulation (α-AS) of the myocardium are associated respectively with an increase and a decrease in myofllament responsiveness to Ca2+. We hypothesized that changes in cytosolic pH (pHi) may modulate these opposite actions of α-AS and β-AS. The effects of α-AS (50 μM phenylephrine and 1 μM nadolol) and β-AS (0.05 μM isoproterenol) on contraction and either cytosolic Ca2+ (Cai) or pHi were assessed in adult rat ventricular myocytes bathed in bicarbonate buffer (pH 7.36±0.05). In cells loaded with the ester derivative (AM form) of indo-1, the 410/490-nm ratio of emitted fluorescence indexed Cai. Myofilament responsiveness to Ca2+ was assessed by the relaxation phase of the length-indo-1 fluorescence relation during a twitch. α-AS and β-AS shifted this relation in opposite directions, indicating that α-AS increased and β-AS decreased myofilament responsiveness to Ca2+. In addition, the positive inotropic action of α-AS was associated with an increased Cai transient amplitude in 50% of the myocytes (n=12), whereas β-AS always increased Cai (n=5). In cells loaded with the fluorescent pHi probe SNARF-1 AM, the emitted 590/640-nm fluorescence is a measure of pHi. The effect of α-AS on the extent of cell shortening during the twitch (ES) was expressed as the percentage of resting cell length. Both ES and pHi were assessed in myocytes bathed in 1.5 mM [Ca2+] and stimulated at 0.5 Hz (control ES, 7.4±1.5%; control pHi, 7.11±0.05; n =10). α-AS enhanced both ES (ΔEs, 1.8±0.6%; pi (ΔpHi, 0.06±0.01; pi (r=0.76, pi was observed in the absence of electrical stimulation (n=8). The α-AS-induced enhancement of ES and pHi was abolished by 10 μM ethylisopropylamiloride, a Na+-H+ exchange inhibitor (n=7). In additional experiments, myocytes were preincubated either with 0.2 μM 4β-phorbol 12-myristate 13-acetate (n=8) or with 5 nM staurosporine (n=8), which have been shown to downregulate and inhibit Ca2+-activated phospholipid-dependent protein kinase C, respectively. In either group, α-AS had no effect on pHi and decreased ES to ≈60% of control. In myocytes bathed in 0.5 mM [Ca2+] and stimulated at 0.2 Hz, β-AS enhanced ES (control ES as percentage of resting cell length, 2.8±1.0%; ΔES, 8.1±1.5%; n=7; pi either during electrical stimulation (n=7) or at rest (n=11). In summary, under the conditions of this study, α-AS enhances myofilament response to Ca2+ and increases pHi via protein kinase C-mediated activation of Na+-H+ exchange. Cytosolic alkalinization contributes to the effect of α-AS to augment contraction amplitude. An enhanced Cai transient occurs in 50% of the myocytes and is not an absolute requirement for the positive inotropic action of α-AS. In contrast, β-AS does not affect pHi, and cytosolic acidification is not the mechanism for the β-AS-induced decrease in myofilament responsiveness to Ca2+.

Original languageEnglish
Pages (from-to)870-882
Number of pages13
JournalCirculation Research
Volume71
Issue number4
Publication statusPublished - Oct 1992

Fingerprint

Myofibrils
Cardiac Myocytes
Adrenergic Agents
Muscle Cells
Fluorescence
Protein Kinase C
Electric Stimulation
Nadolol
Staurosporine
Phenylephrine
Bicarbonates
Fluorescent Dyes
Isoproterenol
Phospholipids
Myocardium
Buffers
Esters
Acetates
Down-Regulation

Keywords

  • α-adrenoceptors
  • β-adrenoceptors
  • Calcium
  • Cardiac myocytes
  • Cytosolic pH
  • Indo-1
  • Myocardial contraction
  • SNARF-1

ASJC Scopus subject areas

  • Physiology
  • Cardiology and Cardiovascular Medicine

Cite this

Gambassi, G., Spurgeon, H. A., Lakatta, E. G., Blank, P. S., & Capogrossi, M. C. (1992). Different effects of α- and β-adrenergic stimulation on cytosolic pH and myofilament responsiveness to Ca2+ in cardiac myocytes. Circulation Research, 71(4), 870-882.

Different effects of α- and β-adrenergic stimulation on cytosolic pH and myofilament responsiveness to Ca2+ in cardiac myocytes. / Gambassi, Giovanni; Spurgeon, Harold A.; Lakatta, Edward G.; Blank, Paul S.; Capogrossi, Maurizio C.

In: Circulation Research, Vol. 71, No. 4, 10.1992, p. 870-882.

Research output: Contribution to journalArticle

Gambassi, G, Spurgeon, HA, Lakatta, EG, Blank, PS & Capogrossi, MC 1992, 'Different effects of α- and β-adrenergic stimulation on cytosolic pH and myofilament responsiveness to Ca2+ in cardiac myocytes', Circulation Research, vol. 71, no. 4, pp. 870-882.
Gambassi, Giovanni ; Spurgeon, Harold A. ; Lakatta, Edward G. ; Blank, Paul S. ; Capogrossi, Maurizio C. / Different effects of α- and β-adrenergic stimulation on cytosolic pH and myofilament responsiveness to Ca2+ in cardiac myocytes. In: Circulation Research. 1992 ; Vol. 71, No. 4. pp. 870-882.
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T1 - Different effects of α- and β-adrenergic stimulation on cytosolic pH and myofilament responsiveness to Ca2+ in cardiac myocytes

AU - Gambassi, Giovanni

AU - Spurgeon, Harold A.

AU - Lakatta, Edward G.

AU - Blank, Paul S.

AU - Capogrossi, Maurizio C.

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N2 - α-Adrenergic stimulation (α-AS) and β-adrenergic stimulation (α-AS) of the myocardium are associated respectively with an increase and a decrease in myofllament responsiveness to Ca2+. We hypothesized that changes in cytosolic pH (pHi) may modulate these opposite actions of α-AS and β-AS. The effects of α-AS (50 μM phenylephrine and 1 μM nadolol) and β-AS (0.05 μM isoproterenol) on contraction and either cytosolic Ca2+ (Cai) or pHi were assessed in adult rat ventricular myocytes bathed in bicarbonate buffer (pH 7.36±0.05). In cells loaded with the ester derivative (AM form) of indo-1, the 410/490-nm ratio of emitted fluorescence indexed Cai. Myofilament responsiveness to Ca2+ was assessed by the relaxation phase of the length-indo-1 fluorescence relation during a twitch. α-AS and β-AS shifted this relation in opposite directions, indicating that α-AS increased and β-AS decreased myofilament responsiveness to Ca2+. In addition, the positive inotropic action of α-AS was associated with an increased Cai transient amplitude in 50% of the myocytes (n=12), whereas β-AS always increased Cai (n=5). In cells loaded with the fluorescent pHi probe SNARF-1 AM, the emitted 590/640-nm fluorescence is a measure of pHi. The effect of α-AS on the extent of cell shortening during the twitch (ES) was expressed as the percentage of resting cell length. Both ES and pHi were assessed in myocytes bathed in 1.5 mM [Ca2+] and stimulated at 0.5 Hz (control ES, 7.4±1.5%; control pHi, 7.11±0.05; n =10). α-AS enhanced both ES (ΔEs, 1.8±0.6%; pi (ΔpHi, 0.06±0.01; pi (r=0.76, pi was observed in the absence of electrical stimulation (n=8). The α-AS-induced enhancement of ES and pHi was abolished by 10 μM ethylisopropylamiloride, a Na+-H+ exchange inhibitor (n=7). In additional experiments, myocytes were preincubated either with 0.2 μM 4β-phorbol 12-myristate 13-acetate (n=8) or with 5 nM staurosporine (n=8), which have been shown to downregulate and inhibit Ca2+-activated phospholipid-dependent protein kinase C, respectively. In either group, α-AS had no effect on pHi and decreased ES to ≈60% of control. In myocytes bathed in 0.5 mM [Ca2+] and stimulated at 0.2 Hz, β-AS enhanced ES (control ES as percentage of resting cell length, 2.8±1.0%; ΔES, 8.1±1.5%; n=7; pi either during electrical stimulation (n=7) or at rest (n=11). In summary, under the conditions of this study, α-AS enhances myofilament response to Ca2+ and increases pHi via protein kinase C-mediated activation of Na+-H+ exchange. Cytosolic alkalinization contributes to the effect of α-AS to augment contraction amplitude. An enhanced Cai transient occurs in 50% of the myocytes and is not an absolute requirement for the positive inotropic action of α-AS. In contrast, β-AS does not affect pHi, and cytosolic acidification is not the mechanism for the β-AS-induced decrease in myofilament responsiveness to Ca2+.

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