Mechano-electric feedback means that muscle stretching causes depolarization of membrane potential. We investigated whether muscle stretching induces action potential and twitch contraction with a ...threshold of sarcomere length (SL) and what roles stretch-activated channels (SACs) and stretch-activated NADPH oxidase (X-ROS signaling) play in the induction. Trabeculae were obtained from the right ventricles of rat hearts. Force, SL, and Ca
2+
i
were measured. Various degrees of stretching from the SL of 2.0 μm were applied 0.5 s after the last stimulus of the electrical train with 0.4-s intervals for 7.5 s. The SL
twitch
was defined as the minimal SL at which twitch contraction was induced by the stretching. Muscle stretching induced twitch contraction with a threshold of SL at 0.4-s stimulus intervals (Ca
2+
o
= 0.7 mmol/L). The SL
twitch
was not changed by increasing the stimulus intervals and Ca
2+
o
and by adding 1 μmol/L isoproterenol. The SL
twitch
was not changed by adding 10 μmol/L Gd
3+
, 100 μmol/L or 200 μmol/L streptomycin, and 5 μmol/L GsMTx4. The SL
twitch
was not changed by adding 1 μmol/L ryanodine and 3 μmol/L diphenyleneiodonium chloride. In contrast, the SL
twitch
was increased by elevating extracellular K
+
from 5 to 10 mmol/L and by adding the stretching during the refractory period of membrane potential. The addition of the stretching-induced twitch contraction more frequently induced arrhythmias. These results suggest that muscle stretching can induce twitch contraction with a threshold of SL and concern the occurrence of arrhythmias and that SACs and X-ROS signaling play no roles in the induction.
In non-diabetic patients with severe disease, such as acute myocardial infarction or acute heart failure, admission blood glucose level is associated with their short-term and long-term mortality. We ...examined whether transient elevation of glucose affects contractile properties in non-diabetic hearts. Force, intracellular Ca
2+
(Ca
2+
i
), and sarcomere length were measured in trabeculae from rat hearts. To assess contractile properties, maximum velocity of contraction (Max
dF
/
dt
) and minimum velocity of relaxation (Min
dF
/
dt
) were calculated. The ratio of phosphorylated troponin I (P-TnI) to troponin I (TnI) was measured. One hour after elevation of glucose from 150 to 400 mg/dL, developed force, Max
dF
/
dt
, and Min
dF
/
dt
were reduced without changes in Ca
2+
i
transients at 2.5 Hz stimulation and 2.0 mM Ca
2+
o
, while developed force and Ca
2+
i
transients showed no changes at 0.5 Hz stimulation and 0.7 mM Ca
2+
o
. In the presence of 1 μM KN-93, a Ca
2+
/calmodulin-dependent protein kinaseII (CaMKII) inhibitor, or 50 μM diazo-5-oxonorleucine, a
l
-glutamine-
d
-fructose-6-phosphate amidotransferase inhibitor, the reduction of contractile properties after elevation of glucose was suppressed. Furthermore, 1 h after elevation of glucose to 400 mg/dL at 2.0 mM Ca
2+
o
, the ratio of P-TnI to TnI was increased. These results suggest that in non-diabetic hearts under higher Ca
2+
-load, transient elevation of glucose for 1 h reduces contractile properties probably by activating CaMKII through O-GlcNAcylation. Thus, in the patients with severe disease, transient elevation of blood glucose, such as due to stress, may worsen cardiac function and thereby affect their mortality without known diabetes.
Background:In non-diabetic patients with acute coronary syndrome, stress hyperglycemia occasionally occurs and is related to their mortality. Whether transient elevation of glucose affects arrhythmia ...susceptibility in non-diabetic hearts with non-uniform contraction was examined.Methods and Results:Force, intracellular Ca2+(Ca2+i), and membrane potential were measured in trabeculae from rat hearts. Non-uniform contraction was produced by a jet of paralyzing solution. Ca2+waves and arrhythmias were induced by electrical stimulation (2.0 mmol/L Ca2+o). The activity of Ca2+/calmodulin-dependent protein kinaseII (CaMKII) was measured. An elevation of glucose from 150 to 400 mg/dL increased the velocity of Ca2+waves and the number of spontaneous action potentials triggered by electrical stimulation. Besides, the elevation of glucose increased the CaMKII activity. In the presence of 1 μmol/L KN-93, the elevation of glucose did not increase the velocity of Ca2+waves and the number of triggered action potentials. In addition, in the presence of 1 μmol/L autocamtide-2 related inhibitory peptide or 50 μmol/L diazo-5-oxonorleucine, the elevation of glucose did not increase the number of triggered action potentials. Furthermore, the elevation of glucose by adding L-glucose did not increase their number.Conclusions:In non-diabetic hearts with non-uniform contraction, transient elevation of glucose increases the velocity of Ca2+waves by activating CaMKII,probably through glycosylation with O-linked β-N-acetylglucosamine, thereby increasing arrhythmia susceptibility.
The propagation velocity of Ca2+ waves determines delayed afterdepolarization and affects the occurrence of triggered arrhythmias in cardiac muscle. We focused on myofilament Ca2+ sensitivity, ...investigating how the velocity of Ca2+ waves responds to its increased sensitivity resulting from muscle stretch or the addition of a myofilament Ca2+ sensitizer, SCH00013. We further investigated whether production of reactive oxygen species (ROS) may be involved in the change in velocity.
Trabeculae were obtained from rat hearts. Force, sarcomere length, and Ca2+i were measured. ROS production was estimated from 2′,7′-dichlorofluorescein (DCF) fluorescence. Trabeculae were exposed to a 10mM Ca2+ jet for the induction of Ca2+ leak from the sarcoplasmic reticulum in its exposed region. Ca2+ waves were induced by 2.5-Hz stimulus trains for 7.5s (24°C, 2.0mM Ca2+o). Muscle stretch of 5, 10, and 15% was applied 300ms after the last stimulus of the train.
Muscle stretch increased the DCF fluorescence, the amplitude of aftercontractions, and the velocity of Ca2+ waves depending on the degree of stretch. After preincubation with 3μM diphenyleneiodonium (DPI), muscle stretch increased only the amplitude of aftercontractions but not the DCF fluorescence nor the velocity of Ca2+ waves. SCH00013 (30μM) increased the DCF fluorescence, the amplitude of aftercontractions, and the velocity of Ca2+ waves. DPI suppressed these increases.
Muscle stretch increases the velocity of Ca2+ waves by increasing ROS production, not by increasing myofilament Ca2+ sensitivity. In the case of SCH00013, ROS production increases myofilament Ca2+ sensitivity and the velocity of Ca2+ waves. These results suggest that ROS rather than myofilament Ca2+ sensitivity plays an important role in the determination of the velocity of Ca2+ waves, that is, arrhythmogenesis.
•Muscle stretch increases ROS production depending on its degree.•Muscle stretch increases the velocity of Ca2+ waves by increasing ROS production.•SCH00013, a myofilament Ca2+ sensitizer, increases ROS production.•ROS production by SCH00013 increases the velocity of Ca2+ waves.
The Kidney Disease: Improving Global Outcomes chronic kidney disease (CKD) guidelines recommend that CKD be classified based on the etiology, glomerular filtration rate (GFR) and degree of ...albuminuria. The present study aimed to establish a method that predicts the presence of microalbuminuria by measuring the total urine protein-to-creatinine ratio (TPCR) in patients with cardiovascular disease (CVD) risk factors.
We obtained urine samples from 1,033 patients who visited the cardiovascular clinic at St. Luke's International Hospital from February 2012 to August 2012. We measured the TPCR and the urine albumin-to-creatinine ratio (ACR) from random spot urine samples. We performed correlation, receiver operating characteristic (ROC) curve, sensitivity, and subgroup analyses. There was a strong positive correlation between the TPCR and ACR (R2 = 0.861, p<0.001). A ROC curve analysis for the TPCR revealed a sensitivity of 94.4%, a specificity of 86.1%, and an area under the curve of 0.903 for detecting microalbuminuria for a TPCR cut-off value of 84 mg/g of creatinine. The subgroup analysis indicated that the cut-off value could be used for patients with CVD risk factors.
These results suggest that the TPCR with an appropriate cut-off value could be used to screen for the presence of microalbuminuria in patients with CVD risk factors. This simple, inexpensive measurement has broader applications, leading to earlier intervention and public benefit.
Background:Connexin43 (Cx43) is a major connexin that forms gap junction (GJ) channels in the heart and is also present in the cell membrane as unopposed/non-junctional hemichannels and in the inner ...mitochondrial membrane. By using carbenoxolone (CBX), a blocker of Cx43, the effect of the blockade of Cx43 on Ca2+waves and triggered arrhythmias in the myocardium with non-uniform contraction was examined.Methods and Results:Trabeculae were obtained from rat hearts. Force, Ca2+i, and the diffusion coefficient were measured. Non-uniform contraction was produced with a 2,3-butanedione monoxime jet. Ca2+waves were induced by electrical stimulation. Inducibility of arrhythmias was estimated based on the minimal Ca2+oat which arrhythmias were induced. The Ca2+spark rate was measured in isolated single rat ventricular myocytes. CBX reduced the GJ permeability, whereas it did not change force and Ca2+itransients. CBX increased the Ca2+leak from the sarcoplasmic reticulum in trabeculae and increased the Ca2+spark rate in isolated single myocytes. CBX increased the velocity of Ca2+waves and further increased the inducibility of arrhythmias. Modulation of mitochondrial KATPchannels by diazoxide, cromakalim and 5-hydroxydecanoic acid affected the inducibility of arrhythmias increased by CBX.Conclusions:These results suggest that in diseased hearts, Cx43 plays an important role in the occurrence of triggered arrhythmias, probably under the modulation of mitochondrial KATPchannels. (Circ J 2016; 80: 76–84)
Arrhythmias are benign or lethal, depending on their sustainability and frequency. To determine why lethal arrhythmias are prone to occur in diseased hearts, usually characterized by nonuniform ...muscle contraction, we investigated the effect of nonuniformity on sustainability and frequency of triggered arrhythmias.
Force, membrane potential, and intracellular Ca(2+) concentration (Ca(2+)(i)) were measured in 51 rat ventricular trabeculae. Nonuniform contraction was produced by exposing a restricted region of muscle to a jet of 20 mmol/L 2,3-butanedione monoxime (BDM) or 20 mumol/L blebbistatin. Sustained arrhythmias (>10 seconds) could be induced by stimulus trains for 7.5 seconds only with the BDM or blebbistatin jet (100 nmol/L isoproterenol, 1.0 mmol/L Ca(2+)(o), 24 degrees C). During sustained arrhythmias, Ca(2+) surges preceded synchronous increases in Ca(2+)(i), whereas the stoppage of the BDM jet made the Ca(2+) surges unclear and arrested sustained arrhythmias (n=6). With 200 nmol/L isoproterenol, 2.5 mmol/L Ca(2+)(o), and the BDM jet, lengthening or shortening of the muscle during sustained arrhythmias accelerated or decelerated their cycle in both the absence (n=10) and presence (n=10) of 100 mumol/L streptomycin, a stretch-activated channel blocker, respectively. The maximum rate of force relaxation correlated inversely with the change in cycle lengths (n=14; P<0.01). Sustained arrhythmias with the BDM jet were significantly accelerated by 30 mumol/L SCH00013, a Ca(2+) sensitizer of myofilaments (n=10).
These results suggest that nonuniformity of muscle contraction is an important determinant of the sustainability and frequency of triggered arrhythmias caused by the surge of Ca(2+) dissociated from myofilaments in cardiac muscle.
Ca2+ waves are initiated not only by Ca2+ leak from the sarcoplasmic reticulum (SR), but also by Ca2+ dissociation from the myofilaments in the myocardium with nonuniform contraction. We investigated ...whether contractile properties and the production of reactive oxygen species (ROS) affect Ca2+ wave propagation. Trabeculae were obtained from 76 rat hearts. Force was measured with a strain gauge, sarcomere length with a laser diffraction technique, and Ca2+i with fura-2 and a CCD camera (24°C, 2.0mmol/L Ca2+o). ROS production was estimated from 2′,7′-dichlorofluorescein (DCF) fluorescence. Trabeculae were regionally exposed to a jet of solution containing 1) 10mmol/L Ca2+ to initiate Ca2+ waves by SR Ca2+ leak due to Ca2+ overload within the jet-exposed region, and 2) 0.2mmol/L Ca2+ or 5mmol/L caffeine to initiate such waves by Ca2+ dissociation from the myofilaments due to nonuniform contraction. Ca2+ waves were induced by stimulus trains for 7.5s. Ten-percent muscle stretch increased DCF fluorescence and accelerated Ca2+ waves initiated due to both Ca2+ overload and nonuniform contraction. Preincubation with 3μmol/L diphenyleneiodonium or 10μmol/L colchicine suppressed the increase in DCF fluorescence but suppressed acceleration of Ca2+ waves initiated only due to Ca2+ overload. Irrespective of preincubation with colchicine, reduction of force after the addition of 10μmol/L blebbistatin did not decelerate Ca2+ waves initiated due to Ca2+ overload, while it did decelerate waves initiated due to nonuniform contraction. These results suggest that Ca2+ wave propagation is modulated by ROS production through an intact microtubule network only during stretch and may be additionally modulated by Ca2+ dissociated from the myofilaments in the case of nonuniform contraction.
► Ca2+ waves due to Ca2+ overload and nonuniform contraction were induced. ► Muscle stretch and 10μmol/L blebbistatin were applied. ► ROS production during stretch modulates Ca2+ waves. ► Ca2+ from the myofilaments modulates Ca2+ waves only due to nonuniform contraction.
Chronic kidney disease (CKD) is a global public health issue, and strategies for its early detection and intervention are imperative. The latest Japanese CKD guideline recommends that patients ...without diabetes should be classified using the urine protein-to-creatinine ratio (PCR) instead of the urine albumin-to-creatinine ratio (ACR); however, no validation studies are available. This study aimed to validate the PCR-based CKD risk classification compared with the ACR-based classification and to explore more accurate classification methods. We analyzed two previously reported datasets that included diabetic and/or cardiovascular patients who were classified into early CKD stages. In total, 860 patients (131 diabetic patients and 729 cardiovascular patients, including 193 diabetic patients) were enrolled. We assessed the CKD risk classification of each patient according to the estimated glomerular filtration rate and the ACR-based or PCR-based classification. The use of the cut-off value recommended in the current guideline (PCR 0.15 g/g creatinine) resulted in risk misclassification rates of 26.0% and 16.6% for the two datasets. The misclassification was primarily caused by underestimation. Moderate to substantial agreement between each classification was achieved: Cohen’s kappa, 0.56 (95% confidence interval, 0.45-0.69) and 0.72 (0.67-0.76) in each dataset, respectively. To improve the accuracy, we tested various candidate PCR cut-off values, showing that a PCR cut-off value of 0.08-0.10 g/g creatinine resulted in improvement in the misclassification rates and kappa values. Modification of the PCR cut-off value would improve its efficacy to identify high-risk populations who will benefit from early intervention.
Although budesonide/formoterol (BUD/FORM) is used clinically as a steroid/2-agonist single inhaler, it has not yet been clarified whether BUD/FORM has inotropic effects on diaphragm muscles after ...inhalation.
We examined the effects of BUD/FORM inhalation, endotoxin injection, and BUD/FORM inhalation plus endotoxin injection on diaphragm contractile properties and nitric oxide (NO) production. After these three treatments, the diaphragm muscle was dissected, and its contractile properties were measured. Histochemistry for the reduced form of nicotinamide adenine dinucleotide phosphate diaphorase was performed for each muscle to assess NO production.
The force-frequency curves showed an upward shift 1 h after inhalation (p<0.05) in the BUD/FORM inhalation only group. The force-frequency curves showed a downward shift 4 h after injection (p<0.001) in the endotoxin injection groups. In the BUD/FORM inhalation plus endotoxin injection groups, a downward shift in the force-frequency curves at 4 h after endotoxin injection was prevented. NO production was inhibited in the BUD/FORM inhalation plus endotoxin injection group compared with that of the endotoxin injection only groups.
BUD/FORM inhalation has an inotropic effect on diaphragm muscle, protects diaphragm muscle deterioration after endotoxin injection, and inhibits NO production. Increments in muscle contractility with BUD/FORM inhalation are induced through a synergistic effect of an anti-inflammatory agent and 02-agonist.
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