ABSTRACT
The human ventricle contains both beta(1)- and beta(1)-adrenoceptors (AR) and both have been shown to be present on a single myocyte. In animal ventricular myocardium there is evidence that beta(1)ARs increase cardiac contraction by non-cAMP-dependent mechanisms. We have used the anti-adrenergic effects of carbachol and the cAMP antagonist Rp -cAMPS to investigate the functional contribution of cAMP to beta(2)AR responses in human ventricular myocytes isolated from cardiac biopsies or explants. Concentration-response curves to isoproterenol (Iso) were constructed in the absence and presence of a beta(1)AR antagonist, CGP 207 12A (300 nmol/l) to determine the contribution of the beta(2)AR to contraction. The cells were rechallenged with sub-maximal dose of Iso under beta(2)AR-specific conditions and Rp -cAMPS (100-200 micromol/l) or carbachol (1-3 microm/l) added. Rp -cAMPS significantly decreased contraction amplitude (% shortening; Iso 7.1+/-0.7, Iso+Rp -cAMPS 3.5+/-0.5, n=7, P<0.001) though not completely to the baseline (2.2+/-0.6, n=7). Rechallenge with Iso alone reversed the effects of Rp -cAMPS, and subsequent addition of the beta(1)AR antagonist ICI 118,551 reduced the response to baseline (1.6+/-0.3, n=4) confirming beta(2)AR involvement. Similarly, carbachol decreased Iso-stimulated contraction from 7.5+/-1.2% to 3.2+/-0.9% (P<0.05, n=4), but not completely to basal levels (1.6+/-0.3%). These results provide functional evidence for a predominantly cAMP-mediated mechanism of contractile stimulation by beta(1)ARs in human ventricular myocardium, although a small contribution from a non-cAMP dependent pathway may occur.
Subject(s)
Cyclic AMP/metabolism , Heart Ventricles/metabolism , Receptors, Adrenergic, beta-2/metabolism , Adrenergic beta-Antagonists/pharmacology , Adult , Aged , Carbachol/pharmacology , Cardiotonic Agents/pharmacology , Cells, Cultured , Cyclic AMP/physiology , Dose-Response Relationship, Drug , Female , Humans , Imidazoles/pharmacology , Isoproterenol/pharmacology , Male , Middle Aged , Myocardial Contraction/drug effects , Myocardial Contraction/physiology , Receptors, Adrenergic, beta-2/physiologyABSTRACT
To determine whether there is a defect in the surviving muscle cells of the failing human heart, studies have been performed on individual myocytes isolated from normal and failing human myocardium. Myocytes from the failing ventricle contract and relax more slowly, and have a reduced contraction amplitude at physiological (but not low) stimulation frequencies. Slow relaxation is seen irrespective of the aetiology of the heart disease studied, and is more pronounced in myocytes from hypertrophied ventricles. Myocytes from hypertrophied ventricles are larger than normal, but the relaxation deficit is independent of cell size. Beta-adrenoceptor desensitization is evident in myocytes and it varies according to the severity of disease and with the age of the patient. Action potentials are longer in myocytes from failing human heart, probably because of an alteration in K+ current density. Many of the functional changes identified in failing human myocardium are seen at the level of the single cardiac myocyte, which implies that pharmacological or genetic manipulation of surviving cells is a logical therapeutic strategy.
Subject(s)
Cardiomyopathy, Hypertrophic/physiopathology , Heart Failure/physiopathology , Myocardial Contraction/physiology , Myocardium/metabolism , Action Potentials , Animals , Cells, Cultured , Disease Models, Animal , Humans , Myocardium/cytology , Reaction TimeABSTRACT
OBJECTIVES: beta-Receptor desensitisation, low basal cAMP, and a negative force-frequency relationship are characteristic changes in human heart failure. Isolated cardiomyocytes from noradrenaline-treated guinea pigs also show these features. We tested the hypothesis that low basal cAMP underlies the loss of contractile response to increasing stimulation frequency in this model. METHODS: Isolated cardiomyocytes were obtained from noradrenaline-treated (NA) and sham-operated (SHAM) guinea pigs. They were stimulated from 0.1-2 Hz and contraction amplitude was monitored with a video edge-detection system. RESULTS: NA cells had less positive amplitude-frequency responses (AFR) compared to SHAMs at 2 mM (P = 0.002, n = 17), or midrange Ca2+ concentrations (EC40-EC60) (P < 0.001, n = 13). When the cAMP agonist, 8-CPT-cAMP (CPT, 10 microM) or high Ca2+ (above EC75) was added to NA cells the AFR was normalised to that of SHAM myocytes (NA vs. SHAM P = ns). In control experiments the cAMP antagonists, Rp-cAMPS (Rpc) and Rp-8-CPT-cAMPS (Rp8, 100 microM), blocked the positive inotropic effects of CPT at 0.5 Hz (control pD2 = 4.36 +/- 0.06, Rp8 pD2 = 3.68 +/- 0.08, P < 0.0001), n = 6 paired). Rpc (100 microM) completely but reversibly blocked the effect of maximal isoprenaline in control experiments (P < 0.0001). Neither antagonist reduced the AFR compared to time-matched controls (P = ns, n = 6). Blockade of SERCA2a with thapsigargin resulted in a significant reduction in the AFR (ANOVA P < 0.0001). CONCLUSIONS: The results are consistent with sarcoplasmic reticulum (SR) function being a more important determinant of the amplitude-frequency relationship than tonic levels of cAMP under basal conditions. Reversal of AFR depression by CPT may result from stimulation of SR Ca2+ uptake.
