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.

Gram negative endotoxemia is associated with an intrinsic impairment of cardiomyocyte contraction, in part due to a reduction in myofilament Ca2+ responsiveness. Endotoxemic rat hearts show increased cardiac troponin I (cTnI) phosphorylation at serines 23 and 24, residues required for the protein kinase A (PKA)-dependent reduction of myofilament Ca2+ sensitivity after beta-adrenoceptor stimulation. To investigate the functional significance of increased TnI phosphorylation in endotoxemia, we studied the contractile effects of systemic bacterial lipopolysaccharide (LPS) treatment in transgenic mice (TG) with cardiac-specific replacement of cTnI by slow skeletal TnI (ssTnI, which lacks the PKA phosphorylation sites) and matched nontransgenic littermates (NTG) on a CD1 background. In wild-type CD1 mice treated with LPS (6 mg/kg ip), after 16-18 h there was a significant reduction in the maximum rates of left ventricular pressure development and pressure decline in isolated Langendorff-perfused hearts compared with saline-treated controls and a decrease in isolated myocyte unloaded sarcomere shortening from 6.1 +/- 0.2 to 3.9 +/- 0.2% (1 Hz, 32 degrees C, P<0.05). Similarly, in NTG myocytes, endotoxemia reduced myocyte shortening by 42% from 6.7 +/- 0.2 to 3.9 +/- 0.1% (P<0.05) with no change in intracellular Ca2+ transients. However, in the TG group, LPS reduced myocyte shortening by only 13% from 7.5 +/- 0.2 to 6.5 +/- 0.2% (P<0.05). LPS treatment significantly reduced the positive inotropic effect of isoproterenol in NTG myocytes but not in TG myocytes, even though isoproterenol-induced increases in Ca2+ transient amplitude were similar in both groups. Only LPS-treated NTG hearts showed a significant increase in cTnI phosphorylation. Investigation of the sarcomere shortening-Ca2+ relationship in Triton-skinned cardiomyocytes revealed a significant reduction in myofilament Ca2+ sensitivity after LPS treatment in NTG myocytes, an effect that was substantially attenuated in TG myocytes. In conclusion, the replacement of cTnI with ssTnI in the heart provides significant protection against endotoxemia-induced cardiac contractile dysfunction, most probably by preserving myofilament Ca2+ responsiveness due to prevention of phosphorylation of TnI at PKA-sensitive sites.

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BackgroundPaediatric mental health patients frequently use natural health products (NHP) in addition to prescription medications, but very little is known about adverse events and possible NHP–drug interactions.ObjectiveTo determine: (1) the prevalence of paediatric mental health patients taking prescription medications only, NHP only, both NHP and prescription medications concurrently or neither; (2) which prescription medications and NHP are most commonly used in paediatric mental health populations and (3) adverse events experienced in the last 30 days (serious and non-serious).DesignCross-sectional surveillance study.SettingPaediatric mental health clinics.Population/interventionOn their first clinic visit, paediatric mental health patients were provided with a form inquiring about prescription drug use, NHP use and any undesirable event experienced in the last month.ResultsOf the 536 patients included in this study, 23% (n=120) reported taking only prescription medication(s), 21% (n=109) reported only NHP use, 21% (n=112) reported using both NHP and prescription drugs concurrently, and 36% (n=191) reported using neither. Overall, there were 23 adverse events reported; this represents 6.3%, 2.8%, 10.8% and 0.6% of each population, respectively. The majority of patients who experienced an adverse event reported taking more than one NHP or prescription drug. No serious adverse events were reported.ConclusionNearly half of the paediatric mental health patients in this study were taking NHPs alone or in addition to prescription medications. Active surveillance identified multiple adverse events associated with NHP and prescription drug use; none were serious. Healthcare professionals were encouraged to initiate conversations regarding NHP use.

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