Decrease In Developed Tension Research Articles

Suppression of myofilament cross-bridge kinetic in the heart of orchidectomized rats

AimsThe increased incidence of heart failure with reduced ejection fraction in men compared with women suggests that male sex hormones significantly impact myocardial contractile activation. This study aims to examine associations among molecular alterations, cellular modulations and in vivo cardiac contractile function upon deprivation of testicular hormones. Main methodsMyocardial structure and functions were compared among sham-operated control and twelve-week orchidectomized (ORX) male rats with and without testosterone supplementation. Key findingsEchocardiography and pressure-volume relationships demonstrated a decreased left ventricular ejection fraction compared with sham-operated controls. The percentage of contractility reduction was generally similar to the decrease in tension development detected in both right ventricular trabeculae and skinned isolated left ventricular cardiomyocytes of ORX rats. Reductions in tension cost and the rate constant of tension redevelopment (ktr) in ORX samples suggested a decrease in the rate of cross-bridge formation, reflecting a reduced number of cross-bridges. Slow cross-bridge detachment in ORX rat hearts could result from a shift of myosin heavy chain isoforms towards a slower ATPase activity β-isoform and reductions in the phosphorylation levels of cardiac troponin I and myosin binding protein-C. All the changes in the ORX rat heart, including ejection fractions and myofilament protein expression and phosphorylation, were completed attenuated by a physiological dose of testosterone. SignificanceTestosterone plays a critical role in regulating the mechanical and contractile dynamics of the heart. Deprivation of male sex hormones cause the loss of normal preserved cardiac contractile function leading to a high risk of severe cardiomyopathy progression.

Biphasic effect of hypercapnia on myocardial contractility.

AbstractThe effects of hypercapnia were studied in cat papillary muscles driven at a frequency of 12 beats/min. The Pco2 of the medium was changed from ⋍ 25 mm to 95 mmHg. Developed tension (DT) and maximal rate of tension development (dT/dt max) were recorded during a 30-minute period. Under isometric conditions and at 30 °C, increasing Pco2 resulted in a biphasic phenomenon Characterized by a decrease in DT and dT/dt max, followed by a partial recovery. Similar results were obtained under isotonic conditions, reserpine pretreatment or addition of propranolol. In the latter experiments, however, values of maximum depression and after 30 minutes of hypercapnia, were significantly lower than without the β-blocking agent.At 22 °C and 22 °C plus reserpine, maximum contractility depression was greater and occurred later during the hypercapnic period than at 30 °C. In addition, at 22 °C plus reserpine, the degree and rate of recovery were significantly lower than at 30 °C.High calcium concentration diminished ...

Negative inotropic effect induced by diethylamiloride (DEA) in rabbit myocardium

The effects of the Na+/H+ exchange blocking drug diethylamiloride (DEA) on mechanical function have been studied in the rabbit isolated, arterially perfused interventricular septum. At concentrations of 10(-6)-10(-5) M, DEA induced a significant, dose-dependent, negative inotropic effect (a 54% decrease from control values at the highest concentration), which was slow to develop. After a 45 min washout, recovery was almost complete (95 +/- 3.4%). At concentrations greater than 5 x 10(-5) M, DEA induced a rapid and marked decrease in developed tension, associated with a progressive decrease in excitability and incomplete recovery. Resting tension was not significantly modified at any of the concentrations tested. At greater than 10(-6) M DEA enhanced significantly the transient negative inotropic effect of the brief intracellular acidosis induced by removal of NH4Cl perfusion, both by decreasing the minimal value of developed tension and by increasing the time required to produce this effect. These effects suggest that the dose-dependent DEA negative inotropic effect could be mediated by a progressive intracellular acidosis produced by inhibition of the Na+/H+ exchange system.

Abstract 578: Urotensin-II Upregulation in Human Heart Failure: Direct Effects on Contractility and Sarcoplasmic Reticulum Calcium Load

While studies have reported that Urotensin II (UII) is upregulated in failing (F) hearts, the pathophysiologic implications of increased UII on cardiac function is incompletely understood. We investigated the direct effects of exogenous UII on in vitro contractile performance and sarcoplasmic reticulum (SR) Ca 2+ load in F and non failing (NF) human myocardium. Methods: Isometric force was measured in F (n=11) and NF (n=6) right ventricular trabeculae (0.5 Hz, 37°C) before and after bath application of UII (UII; 0.1, 1.0, 10, 100, 1000 nM). Single and paired rapid cooling contractures (RCCs) were employed to define SR Ca 2+ load and the balance between SERCA and the NCX. Results: UII induced increases in developed tension (DT) in NF trabeculae and dose-dependent decreases in DT in F trabeculae. UII did not alter resting tension in either group. RCC amplitude increased during graded increases in UII in NF trabeculae, suggesting increases in SR calcium load, but remained stable in F trabeculae. In NF trabeculae, UII did not alter the RCC2/RCC1 ratio at any concentration, but in F trabeculae the RCC2/RCC1 ratio increased at 0.1 nM UII then exhibited dose-dependent decreases at higher concentrations. Conclusion: In contrast to the positive inotropic response elicited in NF hearts, UII is a negative inotrope in ventricular trabeculae obtained from F human hearts. Based on the RCC data, UII has little effect on SR Ca 2+ load in F myocardium suggesting that negative inotropic responses reflect decreases in myofilament Ca 2+ sensitivity. The paired RCC data suggests that U-II also affects the functional competition between SERCA and NCX differently in F and NF human hearts.

Effects of High-K+, Na+-Deficient Solution on Contractility of the Smooth Muscles of the Bovine Trachea

A high-K+, Na+-deficient (I-154 K+) solution induced contraction followed by gradual relaxation of the smooth muscles of the bovine trachea, while hyperosmotic addition of 65 mM KCl induced a large sustained contraction. Exposure of the muscle to the I-154 K+ solution induced an increase in the ratio of cellular water content and a sustained increase in oxidized flavoprotein fluorescence or reduced pyridine nucleotide fluorescence. The I-154 K+ solution also induced a sustained increase in [Ca2+]i level. Decreases in developed tension and increases in cellular water content were both prevented by the addition of sucrose or NaCl but not pyruvate. Substitution of KI for KCl in the I-154 K+ solution produced a greater inhibition of contraction, while substitution with K-propionate produced no inhibition of contraction. Moreover, decreases in developed tension and increases in cellular water content were both prevented by addition of 100 microM 4,4'-diisothiocyanatostilbene-2,2'-disulphonic acid (DIDS), but not 10 microM bumetanide or 1 mM acetazolamide. In conclusion, I-154 K+ solution induced-relaxation in the bovine trachea may be due to swelling of smooth muscle cells and the mechanism of swelling is probably involved in DIDS-sensitive anion movement.

Cardioprotective effect of MCC-135 is associated with inhibition of Ca 2+ overload in ischemic/reperfused hearts

Calcium (Ca 2+) overload is an important pathophysiological factor in myocardial ischemic/reperfusion injury. We investigated the effects of a cardioprotective drug, MCC-135, 5-methyl-2-(1-piperazinyl) benzenesulfonic acid monohydrate, on (1) cardiac contractile dysfunction and Ca 2+ overload induced by ischemia and reperfusion, and (2) the Na +/Ca 2+ exchanger in Langendorff-perfused rat hearts. Low-flow 45-min ischemia and 30-min reperfusion decreased developed tension and increased ventricular Ca 2+ content, effects which were ameliorated by MCC-135 and amiloride given after reperfusion. Combination of intracellular Na + overload induced by monensin (Na + ionophore; 5 μM) and zero-flow 15-min ischemia followed by 30-min reperfusion resulted in a decrease in developed tension and in the intracellular Na +-dependent increase in ventricular Ca 2+ content. MCC-135 and the highest dose of amiloride given after reperfusion reduced the increase in ventricular Ca 2+ content, whereas developed tension was increased only with MCC-135. These results suggest that the cardioprotective effect of MCC-135 in ischemia/reperfusion is associated with suppression of Ca 2+ overload and is attributable to inhibition of intracellular Na +-dependent Ca 2+ influx via the Na +/Ca 2+ exchanger.

Interaction of blood flow and oxygen delivery affects peak VO2 and fatigue in canine muscle in situ.

This investigation was designed to examine the interactive effects of blood flow (Q) and oxygen delivery (QO2) on peak oxygen consumption (VO2peak) and fatigue in the gastrocnemius-plantaris dog muscle in situ. Arterial oxygen content (CaO2) was manipulated by varying the fraction of inspired O2 to create a high- and a low-QO2 treatment at each of three levels of Q (approximately/=800, 1100 and 1500 ml x kg(-1) x min(-1)). Isometric contraction was achieved by stimulation with one tetanic contraction per second (200 ms at 70 Hz) at supramaximal voltage for 10 min to achieve VO2peak under each condition. Arterial and venous blood were sampled, and Q and tension development (TD) were measured at 1, 5 and 10 min into the contractions. Fatigue was calculated as the percentage (%) change in TD from min 1 to min 10. While VO2peak was highly correlated to QO2 (R = 0.97) in both the moderate- and high-Q conditions, both low-Q conditions had a significantly (P < 0.05) higher VO2peak than predicted by the QO2-VO2 relationship. In addition, VO2 peak was significantly greater in the low-Q versus the high-Q condition. Fatigue (% decrease in TD) was significantly different between the two low-Q conditions [Low-QO2=38.4 (6.9); Norm QO2=27.9 (1.8)] and the two moderate-Q conditions [low-QO2=31.1 (7.3); high-QO2 = 20.9 (4.1)] but not between the two high-Q conditions [normal-QO2 = 10.4 (6.9); high-QO2 = 11.9 (4.5)]. In the two conditions of equal QO2, fatigue was significantly less in the high-Q condition (10.4%) compared to the low (27.9%). Thus, it seems that there is a unique interaction between Q and QO2 which determine the VO2peak and rate of fatigue in contracting muscle.

Anti-muscarinic actions of mitoxantrone in isolated heart muscles of guinea pigs

A hypotheses that mitoxantrone is a competitive antagonist at muscarinic cholinergic receptors was examined in guinea-pig hearts. In isolated left atrial muscle preparations, electrically paced at 2 Hz, the muscarinic agonist, carbachol, caused a concentration-dependent decrease in developed tension. Mitoxantrone caused a parallel right-ward shift of the concentration–response curve for carbachol. Schild plots for the effect of mitoxantrone on the carbachol concentration–response relationship were linear with a slope of 0.88 which was not significantly different from the unity. The right-ward shift of the carbachol concentration–response relationship by mitoxantrone significantly reversed after an additional incubation with a mitoxantrone-free solution, although the reversal was incomplete after a 2-h incubation in the mitoxantrone-free solution. Mitoxantrone caused a concentration-dependent displacement of specific [ 3H]quinuclidinyl benzilate binding to membrane preparations obtained from ventricular muscles of guinea-pig hearts. These results indicate that mitoxantrone acts as a competitive antagonist for the muscarinic receptors.

The effects of etomidate on the contractility of failing and nonfailing human heart muscle.

We measured the effects of etomidate on contractility of human cardiac muscle. Muscles were obtained from the left ventricle and right atrium of 12 patients undergoing cardiac transplantation, and from the right atrium of 12 patients undergoing coronary artery bypass surgery. Muscles were studied at 37 degrees C and 1.0 Hz. Variables of isometric contraction were recorded before and after etomidate (0.04-80 microM) or its solvent, propylene glycol. The ability of beta-adrenergic stimulation to cause an inotropic effect after etomidate was also assessed. Etomidate caused a dose-dependent decrease in developed tension, which was statistically significant only at concentrations exceeding clinical doses (> or =20 microM; P < 0. 05). Decreases in maximum rates of contraction and relaxation paralleled changes in developed tension. beta-Adrenergic stimulation reversed the etomidate-induced decreases in developed tension and rates of contraction and relaxation to baseline (P > 0.05 compared with baseline). Thus, in human myocardium, etomidate exerts a dose-dependent negative inotropic effect, which is reversible with beta-adrenergic stimulation. Concentrations required to produce these negative inotropic effects are, however, in excess of those reached during clinical use. Therefore, etomidate-induced negative inotropy is unlikely to be a problem clinically, even in patients with cardiac dysfunction. Etomidate produced a similar dose-dependent negative inotropic effect in both failing and nonfailing human myocardium. This effect was present only at concentrations exceeding those attained clinically and was reversible with beta-adrenergic stimulation.

Differential sensitivity to hydroxyl radicals of pre- and postjunctional neurovascular transmission in the isolated canine mesenteric vein

In some pathophysiological conditions, the first target of reactive oxygen intermediates is the vascular system. Superoxide anions, when generated in the vascular circulation, may then escape into the extracellular space via an anion channel and, following dismutation to hydrogen peroxide (H 2O 2), form hydroxyl radicals (HO •). In an attempt to understand the role of HO • in the regulation of transmission at the sympathetic neurovascular junction, the effect of HO • at nerve terminals was examined by measuring the amount of noradrenaline (NA) released from isolated, spirally cut, superfused canine mesenteric vein during basal and electrical stimulation (ES; 5 Hz, 2 ms, 9 V); tension development evoked by ES was also recorded simultaneously. HO • was generated from Fenton's reagent (1.5×10 −4 M H 2O 2 plus 10 −4 M FeSO 4); generation of HO • from H 2O 2/FeSO 4 in the superfusate was monitored by electron spin resonance spectroscopy using the spin-trap 5,5-dimethyl-1-pyrroline- N-oxide throughout the experimental time course. Exposure to HO • of the helical strips produced an irreversible decrease in tension development evoked by ES with no effect on NA release, suggesting that the observed effect is elicited postjunctionally. The susceptibility of the processes of NA-mediated contraction to HO • may differ greatly from that of the NA release mechanism at the prejunctional site. Exposure of the strip preparation to HO • leads to a substantial stimulation of basal release of NA without affecting ES-evoked NA release, possibly due to enhanced non-exocytotic Ca 2+-independent release elicited by HO •. A direct demonstration of this concept was obtained by showing a significant increase in the basal response of NA release in Ca 2+-free solution. The major conclusion of the present study is that HO • can damage NA-mediated contraction of the vascular preparations at the postjunctional site, and may selectively induce a non-exocytotic release of NA from the prejunctional site of sympathetic neurotransmission.

Protective effect of TA-993, a novel therapeutic agent for peripheral circulatory insufficiency, on skeletal muscle fatigue in a rat model of hindlimb ischemia.

TA-993 (cis-(-)-2-(4-methylphenyl)-3-acetoxy-2,3-dihydro-5-(2-dimethylaminoethy l)-8-methyl1,5-benzothiazepine-4(5H)-one maleate), a new 1,5-benzothiazepine derivative, has a selective increasing action on limb blood flow in addition to an antiplatelet action. In this report we studied the effect of TA-993 on a time dependent decrease in developed tension of electrically-induced contraction of tibialis anterior muscle in a rat model of peripheral circulatory insufficiency induced by occlusion of abdominal aorta. In our preparation, the developed tension decreased by 20-30% in a sham-operated group and 30-40% in an abdominal aorta-occluded group at the end of the experimental period of 60 min. Intraduodenal administration (i.d.) of TA-993 (10 mg/kg) to the abdominal aorta-occluded rats ameliorated the decrease in developed tension to the level of the sham-operated group. Moreover, TA-993 at 10 mg/kg, i.d. significantly increased femoral arterial blood flow supplied through collateral circulation and decreased the whole blood viscosity in this model. These results suggest that TA-993 improves dysfunction of skeletal muscle contraction due to peripheral circulating insufficiency through an increase in collateral blood flow and an improvement of red blood cell deformability.

Protective Effect of TA-993, a Novel Therapeutic Agent for Peripheral Circulatory Insufficiency, on Skeletal Muscle Fatigue in a Rat Model of Hindlimb Ischemia

TA-993 (cis-(–)-2-(4-methylphenyl)-3-acetoxy-2,3-dihydro-5-(2-dimethylaminoethyl)-8-methyll,5-benzothiazepine-4(5H)-one maleate), a new 1,5-benzothiazepine derivative, has a selective increasing action on limb blood flow in addition to an antiplatelet action. In this report we studied the effect of TA-993 on a time dependent decrease in developed tension of electrically-induced contraction of tibialis anterior muscle in a rat model of peripheral circulatory insufficiency induced by occlusion of abdominal aorta. In our preparation, the developed tension decreased by 20–30% in a sham-operated group and 30–40% in an abdominal aorta-occluded group at the end of the experimental period of 60 min. Intraduodenal administration (i.d.) of TA-993 (10 mg/kg) to the abdominal aorta-occluded rats ameliorated the decrease in developed tension to the level of the sham-operated group. Moreover, TA-993 at 10 mg/kg, i.d. significantly increased femoral arterial blood flow supplied through collateral circulation and decreased the whole blood viscosity in this model. These results suggest that TA-993 improves dysfunction of skeletal muscle contraction due to peripheral circulating insufficiency through an increase in collateral blood flow and an improvement of red blood cell deformability.

Positive and negative inotropic effects of muscarinic receptor stimulation in mouse left atria

In isolated mouse left atria, acetylcholine (ACh) produced a biphasic inotropic response; a transient decrease in developed tension was followed by an increase. Both negative and positive responses were concentration dependent and were inhibited by atropine. The negative and positive inotropic responses were also observed with a nonselective muscarinic stimulant, oxotremorine-M, but not with an M1-receptor selective stimulant, McN-A343. Pirenzepine, an M1-receptor antagonist, inhibited both negative and positive inotropic responses at high concentrations. Gallamine, an M2-receptor antagonist, inhibited the negative response. Hexahydro-siladifenidol hydrochloride, p-fluoro analog (p-F-HHSiD), an M3-receptor antagonist, inhibited the positive response with no effect on the negative phase. In pertussis toxin (PTX) treated preparations, negative inotropic response to ACh was not observed. These results suggest that the negative and positive inotropic responses to acetylcholine in mouse atria are mediated by M2 and M3 receptors, respectively. The negative phase, but not the positive phase, was mediated by a PTX-sensitive G protein.

Felodipine protects human atrial muscle from hypoxia–reoxygenation dysfunction: a force–frequency relationship study in an in vitro model of stunning

Aims: We aimed at investigating contractile changes after hypoxia–reoxygenation and dobutamine challenge in superfused human atrial pectinate muscle to see whether high versus low stimulation rate during hypoxia might account for outcome differences compatible with the definition of an in vitro model of myocardial stunning and whether pretreatment with the dihydropyridine Ca2+ entry blocker felodipine might afford protection. Methods: Human right atrial trabeculae obtained from adult patients were superfused in an organ bath with oxygenated (O2 content 16 ml/l) and modified (NaHCO3 25.7 mmol/l) Tyrode's solution at 37°C. Dobutamine (1 nmol/l to 10 μmol/l) was superfused in 10 oxygenated preparations to select the optimal drug concentration to be used in another 22 which were randomized. Group (A) consisted of time-related controls (Tyrodes's solution for 225 min at cycle length (CL) 1600 ms and no dobutamine). There were two test groups, respectively: (B) low (1600 ms CL) and (C) high (400 ms CL) stimulation rate. After 60 min of stabilization, in groups B and C, hypoxic superfusion (O2 content 5 ml/l) lasted 60 min, then reoxygenation (60 min) and dobutamine challenge (1 μmol/l, 15 min) were performed. Analysis of variance for repeated measures with the Greenhouse–Geisser correction, and a repeated measures model with structured covariance (preparation mass, length, width and time-varying time to peak tension) matrices were used whereby grouping (G), time (T) and G·T interaction were weighted. Force–frequency relationship and post-pausal potentiation were studied after each phase. Electrophysiology, histomorphometry and electron microscopy were carried out (n=6). Felodipine (0.1 μmol/l, n=5) pretreatment (15 min before hypoxia) was given in parallel experiments. Results: Time-related controls showed =̃ 10% per hour decrease of developed tension and the Paradise test provided ≅80% of control values. In test groups (as compared to baseline values) contractility was decreased ≅65% after hypoxia–reoxygenation and it increased ≅25% after dobutamine (G, 0.0155<P<0.0065; T, P=0.00005; G·T, P=0.00005). High stimulation rate during hypoxia worsened hypoxia–reoxygenation contractile changes, whereas reversibility after dobutamine was less. In both B and C groups during hypoxia, contractility decreased quite rapidly, although by 10 min or so a plateau (≅50%) was reached in group B, whereas in group C contractility decreased to <20%. None of the covariates contributed significantly to predict the dependent variables investigated. Force–frequency relationship and post-pausal potentiation were repeatable, paralleled overall changes due to hypoxia, reoxygenation and dobutamine challenge and were useful to discriminate Ca2+-related diastolic processes thus helping index myocardial contractile reserve. Force–frequency relationship was negative at high stimulation rates, concomitant to an abrupt change of shape and duration of action potential with little time for Ca2+-related Ca2+ release and ensuing systolic processes. Felodipine pretreatment enabled an unblunted response to dobutamine. Histomorphometry showed an unexpected `fibrotic core'. At electron microscopy, subendocardial and deep part of the same pectinate muscles showed identical degrees of degenerative lesions. Superfused samples showed, unexpectedly, less anoxic lesions than preparations fixed within 15 min from surgical explant, although lesions were higher than in samples fixed immediately after explant. Conclusions: This might be a relevant model, whereby pharmacological or physical interventions are tested. Native human atrial trabeculae might be used without dissection and/or preservatives. If high stimulation rate during hypoxia is used the power of hypothesis testing is maximized. Future studies with this material will be easier and comparatively smaller series might be investigated. Felodipine at low concentration was protective in this model.

Effect of acidosis on contraction, intracellular pH, and calcium in the newborn and adult rabbit aorta.

This study investigated the effect of acidosis on intracellular pH (pHi), intracellular calcium concentration ([Ca]i), and vascular contraction in the aorta of the newborn and adult rabbit. Isometric tension, pHi, and [Ca]i were measured in an isolated ring preparation. After the vascular contraction was induced with 50mM KC1, the effect of respiratory acidosis produced by elevation of PCO2 was studied. Respiratory acidosis caused a transient depression followed by a recovery of contractile tension. The decrease in developed tension was greater in the newborn than in the adult. The decrease in pHi during acidosis was similar in the two age groups. [Ca]i increased during acidosis and the increase was greater in the newborn than in the adult. These data show that the vasorelaxant effect of acidosis in the newborn aorta is greater than that in the adult aorta. The greater vasodilation in the newborn cannot be explained by the difference in pHi or [Ca]i.

Endothelin-1 partially inhibits ATP-sensitive K+ current in guinea pig ventricular cells.

To clarify the pathophysiological significance of endothelin (ET) in the ischemic myocardium, we examined the effect of endothelin-1 (ET-1) on the ATP-sensitive K+ current (IK.ATP) and compared it with that of ET-3 in guinea pig ventricular cells using conventional microelectrode and patch clamp techniques. In isolated guinea pig papillary muscles, ET-1 (30 nM) markedly increased developed tension (DT), with little influence on action potential duration (APD), whereas ET-3 at the same concentration failed to affect DT or APD. Both nicorandil (1 mM) and cromakalim (30 microM) markedly shortened APD and decreased DT in papillary muscles. ET-1, but not ET-3, partially reversed the nicorandil-induced decreases in APD and DT in a concentration-dependent manner. ET-1 also attenuated the cromakalim-induced decreases in APD and DT. In single ventricular myocytes, both nicorandil and cromakalim increased a steady-state outward current, which was sensitive to 1 microM glibenclamide, suggesting that these drugs activate IK.ATP. ET-1 (30 nM) significantly inhibited the IK.ATP, whereas ET-3 failed to affect it. The ET-1 induced inhibition of IK.ATP was abolished by BQ-485 (100 nM), an ETA receptor-selective antagonist. Neither the protein kinase C (PKC) inhibitor staurosporine (20 nM) nor the calmodulin antagonist W-7 (50 microM) affected the inhibitory action of ET-1 on the nicorandil-induced IK.ATP. In pertussis toxin (PTX)-treated cells, the inhibitory action of ET-1 on IK.ATP was augmented rather than attenuated. These results suggest that ET-1 partially inhibits the IK.ATP through the activation of ETA receptors, although the precise intracellular mechanism remains to be clarified. Because activation of the ATP-sensitive K+ channels is considered to protect the ischemic myocardium, the partial inhibition of IK.ATP by ET-1 may lead to the aggravation of myocardial injury, potentially due to an increase in transmembrane Ca2+ influx.

Induction of expression of the sodium-hydrogen exchanger in rat myocardium

The aim was to examine the regulation of the cardiac Na+/H+ exchanger NHE-1 isoform mRNA in response to ischaemia and acidosis in the mammalian myocardium. Male Sprague Dawley rat hearts were perfused in a non-circulated retrograde fashion according to the Langendorff method. Hearts were perfused for 3 h at flow rates of either 10 ml.min-1 (control), or 3, 1, or 0 ml.min-1 (ischaemia) followed by 5 min of reperfusion. Hearts were immediately frozen in liquid N2, and stored at -80 degrees C until ready for RNA isolation. Northern blot analysis was used to examine expression of the NHE-1 isoform of the Na+/H+ exchanger message in these isolated perfused hearts. Activity of the Na+/H+ exchanger was assessed in primary cultures of neonatal rat myocytes under either control conditions or after treatment with chronic, low external pH. A decrease in developed tension and an increase in resting tension was observed which was dependent upon the severity of the ischaemic episode. Low flow ischaemia of 3 ml.min-1 caused increased Na+/H+ exchanger message levels, while perfusion at more reduced flow rates eliminated the increase. Treatment of primary cultures of isolated myocytes with low external pH resulted in increased ability to recover from an acute acid load. Low flow ischaemia can increase the Na+/H+ exchanger message in the intact mammalian myocardium. More severe ischaemia prevents the increase, suggesting that severely damaged tissue may not be capable of the ischaemic response. Primary cultures of isolated myocytes can respond to chronic low external pH by increasing Na+/H+ exchanger activity.

Cellular Ca 2+ loading and inotropic effects of doxorubicin in atrial muscle preparations isolated from rat or guinea-pig hearts

The inotropic effects of doxorubicin were examined under conditions that alter the Ca 2+ loading of myocardial cells. Atrial muscle preparations isolated from rat or guinea-pig hearts were used. Cellular Ca 2+ loading was altered by changing the temperature, extracellular Ca 2+ concentrations or the frequency of electrical stimulation. In guinea-pig heart muscle preparations stimulated at 2 Hz at 30°C in the presence of 1.2 mM CaCl 2, 30 μM doxorubicin caused biphasic (early and late phase) positive inotropic effects. A higher concentration (200 μM) of doxorubicin caused a transient positive inotropic effect followed by a gradual decrease in developed tension. An increase in CaCl 2 concentration from 1.2 to 2.4 mM decreased the positive inotropic effect of 30 μM doxorubicin and changed the inotropic effect of 200 μM doxorubicin from positive to negative after a transient increase in developed tension. At 0.5-Hz stimulation or 36°C incubation, 30 or 200 μM doxorubicin produced remarkable late phase positive inotropic effects. In rat heart muscle preparations, the patterns of the inotropic effects of doxorubicin were similar to those observed with guinea-pig hearts; however, the negative inotropic effect observed at 200 μM doxorubicin was greater. These results indicate that the early and the late phase positive inotropic effects of doxorubicin have different mechanisms. The pattern of inotropic effects of doxorubicin is affected by conditions that alter cellular Ca 2+ loading or the concentration of doxorubicin.

Length-dependent modulation of ANG II inotropism in rat myocardium: effects of myocardial infarction.

To determine whether changes in sarcomere length affect the inotropic response of the heart to angiotensin II (ANG II) differently in dilated and failing myocardium, papillary muscles were removed 2 days after infarction, and the effects of ANG II were studied at various muscle lengths. Myocardial infarction, which averaged 52% of the left ventricle inclusive of the interventricular septum, was characterized hemodynamically by left ventricular failure and right ventricular dysfunction. ANG II administration at 100% the muscle length where force development is maximal (Lmax) produced a 12% depression of developed tension in papillary muscles from noninfarcted ventricles and a 37% decrease in developed tension in the viable myocardium of infarcted rats. In contrast, at 85 and 92.5% Lmax and in the presence of ANG II, control muscles increased active tension by 16 and 1.0%, whereas muscles from coronary occluded hearts augmented developed tension by 13 and 22%, respectively. In conclusion, ANG II exerted a positive inotropic effect on rat myocardium at muscle lengths on the ascending limb of the Starling curve but a negative inotropic action at the muscle length normally associated with maximum force development. This phenomenon emphasizes that hormonal influences on the contractile state of the diseased heart may be modulated by the interaction of end-diastolic pressure, sarcomere length, and ventricular size and shape.

The acute effects of the creatine analogue, β-guanidinopropionic acid, on cardiac energy metabolism and function

1. Perfusion of isolated rat hearts with 150 mM βGPA led to the linear accumulation of intracellular PβGPA (approx. 150 nmol/min per g (dry wt.)) after an initial lag period of 20 min. 2. This accumulation of intracellular PβGPA was accompanied by a decrease in PCr (30%) and an increase in total phosphagen content (20%). These results show that PCr was not equally replaced by PβGPA, but was degraded at the expense of βGPA phosphorylation to produce a net increase in cardiac phosphagen content. Correspondingly, total phosphate (the sum of PCr, PβGPA, P i and ATP) was increased, indicating that there was no cellular necrosis and that the sarcolemma remained intact throughout the perfusion. 3. An increase in P i and decrease in ATP also occurred concomitantly with PβGPA accumulation, indicating that ATP synthesis was not keeping up with demand. This may be due to the gradual replacement of PCr by the less efficient phosphagen, PβGPA, resulting in inadequate transduction of energy and hence an imbalance between energy demand and supply. However, the increased hyperosmolarity of the perfusate may be partly responsible for these effects on cardiac energy metabolism, as perfusion with 150 mM mannitol produced a similar decrease in ATP, but a smaller rise in P i. 4. Perfusion with either 150 mM βGPA or mannitol led to a significant intracellular alkalosis (max. pH i 7.3), which was reversed on returning to normal perfusate. In addition, both hyperosmolar perfusions led to a significant reduction in cardiac frequency (40 and 15%, respectively). However, only βGPA caused significant negative inotropism. The time-courses for the changes in cardiac frequency and pH i did parallel the increase in PβGPA. This suggests that both hyperosmolarity and the production of PβGPA during βGPA perfusions determine the degree of negative chronotropism, but that hyperosmolarity alone causes alkalosis and βGPA phosphorylation, a decrease in developed tension. 5. When hearts, acutely loaded with PβGPA were perfused with control medium, the levels of ATP, PCr and PβGPA stabilised to produce a new steady state. There was no decrease in PβGPA concentration during this procedure, implying that βGPA efflux was negligible.