Essential role of troponin I in the positive inotropic response to isoprenaline in mouse hearts contracting auxotonically

Abstract

PKA-dependent phosphorylation of cardiac troponin I (cTnI) contributes significantly to β-adrenergic agonist-induced acceleration of myocardial relaxation (lusitropy). However, the role of PKA-dependent cTnI phosphorylation in the positive inotropic response to β-adrenergic stimulation is unclear. We studied the contractile response to isoprenaline (10 nM) in isolated hearts and isolated cardiomyocytes from transgenic mice with cardiacspecific expression of slow skeletal TnI (ssTnI, which lacks the N-terminal protein extension containing PKA-sensitive phosphorylation sites in cTnI) and matched wild-type littermate controls. As expected, the lusitropic effect of isoprenaline was significantly blunted in ssTnI hearts. However, the positive inotropic response to isoprenaline was also blunted in ssTnI hearts. This effect was especially prominent for ejection-phase indices in isolated auxotonically loaded ssTnI hearts whereas the positive inotropic response of isovolumic hearts or unloaded isolated myocytes was much less affected. Isoprenaline decreased left ventricular end-systolic volume in wild-type hearts (10.6 ± 1.6 to 6.2 ± 0.4 µl at a preload of 20 cmH 2 O; P < 0.05) but not transgenic hearts (11.4 ± 1.3 to 10.9 ± 1.3 µl; P = n.s.). Likewise, isoprenaline increased stroke work in control hearts (14.5 ± 1.0 to 22.5 ± 1.8 mmHg µl mg −1 ; P < 0.05) but not transgenic hearts (15.4 ± 1.3 to 18.3 ± 1.2 mmHg µl mg −1 ; P = n.s.). The end-systolic pressurevolume relation was increased by isoprenaline to a greater extent in control than transgenic hearts. However, isoprenaline induced a similar rise in intracellular Ca 2+ transients in transgenic and non-transgenic cardiomyocytes. These results indicate that cTnI has a pivotal role in the positive inotropic response of the murine heart to β-adrenergic stimulation, an effect that is highly dependent on loading conditions and is most evident in the auxotonically loaded ejecting heart.

Expression of Slow Skeletal Troponin I in Adult Mouse Heart Helps to Maintain the Left Ventricular Systolic Function During Respiratory Hypercapnia

“…An important rationale for our experiments comes from our recent data indicating that the role of TnI on cardiac function is best measured in ejecting, afterloaded hearts. [11][12][13] Our experiments provide the first evidence that specific replacement of cTnI with ssTnI has a protective effect on the left ventricular (LV) systolic function during hypercapnic acidosis in the intact animal.…”

The unique functions of cardiac troponin I in the control of cardiac muscle contraction and relaxation

“…The role of cTnI phosphorylation in the relaxant effect of adrenergic stimulation has also been emphasized in work reported by Stelzer et al [15] in studies of mice expressing cTnI-S23D/S24D against a cTnI and myosin binding protein C null background. These studies together with others provide compelling evidence for a prominent role of cTnI phosphorylation in the maintenance of power and frequency response in ejecting ventricles [13,[16][17][18][19]. Apart from their significant effects on cardiac dynamics, cTnI phosphorylation at the PKA sites affects length dependence of activation (LDA) [20].…”

“…While the inhibitory region in both skeletal and cardiac TnI has been shown to be helical in the absence of Ca 2+ -binding to the regulatory domain [36], bioinformatics analyses supports a β-turn type conformation for this region when bound to actin [29], as mentioned. Recently, the determined structure for the cardiac N-extension bisphosphorylated Ser-23/Ser-24 was found to contain a C-terminal helix (residues 21-30) containing the phosphorylation motif, an extended poly(L-proline)II helix (residues [11][12][13][14][15][16][17][18][19], and an acidic N-terminus with some propensity for helical structure [47]. Using this structure, the X-ray crystal structure of the cTn core, and uniform density models of the cTn subunits derived from neutron contrast variation data, atomic models were built that show the conformational transition induced by PKA phosphorylation at Ser-23/Ser-24 of cTnI and suggest a molecular linkage between the cardiac N-extension and the inhibitory region of cTnI [47].…”

Another New Kinase Targets Troponin I

“…More recent data however suggest TnI phosphorylation at PKA sites also has effects on myocyte power output, 19 as well as systolic performance, force frequency relationship, and the response to afterload in vivo. 20,21 Although phosphorylation by PKC has been less intensively studied, data indicate phosphorylation of TnI by PKC is associated with a decreased maximal myofibrillar MgATPase. 22 The primary PKC phosphorylation sites of mouse TnI are Ser-42, Ser-44, and Thr-143; however, in vitro studies with site-specific mutants indicated that the Ser-42 and Ser-44 sites (referred to as Ser-43 and Ser-45 because of including the initiating methionine) are primarily involved in the functional effect.…”