A combined Langendorff-injection technique for simultaneous isolation of single cardiomyocytes from atria and ventricles of the rat heart

Butova, Xenia; Myachina, Tatiana; Khokhlova, Anastasia

doi:10.1016/j.mex.2020.101189

Cited by 12 publications

(9 citation statements)

References 17 publications

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“…Single cardiomyocytes from LA and RA were isolated using a combined technique of Langedorff perfusion and intra-chamber injections as described in detail elsewhere [ 22 ]. Briefly, the isolated heart was cannulated via the aorta to the Langendorff apparatus and perfused at a rate of 3.0–3.5 mL/min with a sequence of three solutions equilibrated with 100% O 2 .…”

Section: Methodsmentioning

confidence: 99%

Peculiarities of the Acetylcholine Action on the Contractile Function of Cardiomyocytes from the Left and Right Atria in Rats

Butova

Myachina

Simonova

et al. 2022

Cells

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Acetylcholine (ACh) is the neurotransmitter of the parasympathetic nervous system that modulates cardiac function, and its high concentrations may induce atrial fibrillation. We compared the ACh action on the mechanical function of single cardiomyocytes from the left atria (LA) and the right atria (RA). We exposed single rat LA and RA cardiomyocytes to 1, 10, and 100 µM ACh for 10–15 min and measured the parameters of sarcomere shortening–relengthening and cytosolic calcium ([Ca2+]i) transients during cell contractions. We also studied the effects of ACh on cardiac myosin function using an in vitro motility assay and analyzed the phosphorylation level of sarcomeric proteins. In LA cardiomyocytes, ACh decreased the time to peak sarcomere shortening, time to 50% relengthening, and time to peak [Ca2+]i transients. In RA cardiomyocytes, ACh affected the time of shortening and relengthening only at 10 µM. In the in vitro motility assay, ACh reduced to a greater extent the sliding velocity of F-actin over myosin from LA cardiomyocytes, which was accompanied by a more pronounced decrease in phosphorylation of the myosin regulatory light chain (RLC) in LA cardiomyocytes than in RA cardiomyocytes. Our findings indicate that ACh plays an important role in modulating the contractile function of LA and RA, provoking more pronounced changes in the time course of sarcomere shortening–relengthening and the kinetics of actin–myosin interaction in LA cardiomyocytes.

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Section: Methodsmentioning

confidence: 99%

Peculiarities of the Acetylcholine Action on the Contractile Function of Cardiomyocytes from the Left and Right Atria in Rats

Butova

Myachina

Simonova

et al. 2022

Cells

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“…After the appearance of viscous drops at the heart apex, the heart was transferred to a Petri dish with enzyme solution. The LV was gently digested by the Langendorff-free injection technique ( Butova et al, 2021 ) at a rate of 6.0–7.0 ml/min and 36°C. After complete digestion, thin ENDO and EPI layers (<1/3 width of the LV free wall) were obtained using fine scissors and forceps.…”

Section: Methodsmentioning

confidence: 99%

Contractile State Dependent Sarcomere Length Variability in Isolated Guinea-Pig Cardiomyocytes

et al. 2022

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Cardiomyocytes contract keeping their sarcomere length (SL) close to optimal values for force generation. Transmural heterogeneity in SL across the ventricular wall coordinates the contractility of the whole-ventricle. SL heterogeneity (variability) exists not only at the tissue (macroscale) level, but also presents at the level of a single cardiomyocyte (microscale level). However, transmural differences in intracellular SL variability and its possible dependence on the state of contraction (e.g. end-diastole or end-systole) have not been previously reported. In the present study, we studied three aspects of sarcomere-to-sarcomere variability in intact cardiomyocytes isolated from the left ventricle of healthy guinea-pig: 1) transmural differences in SL distribution between subepi- (EPI) and subendocardial (ENDO) cardiomyocytes; 2) the dependence of intracellular variability in SL upon the state of contraction; 3) local differences in SL variability, comparing SL distributions between central and peripheral regions within the cardiomyocyte. To characterize the intracellular variability of SL, we used different normality tests for the assessment of SL distributions, as well as nonparametric coefficients to quantify the variability. We found that individual SL values in the end-systolic state of contraction followed a normal distribution to a lesser extent as compared to the end-diastolic state of contraction (∼1.3-fold and ∼1.6-fold in ENDO and EPI, respectively). The relative and absolute coefficients of sarcomere-to-sarcomere variability in end-systolic SL were significantly greater (∼1.3-fold) as compared to end-diastolic SL. This was independent of both the transmural region across the left ventricle and the intracellular region within the cardiomyocyte. We conclude that the intracellular variability in SL, which exists in normal intact guinea-pig cardiomyocytes, is affected by the contractile state of the myocyte. This phenomenon may play a role in inter-sarcomere communication in the beating heart.

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“…Adult male Wistar rats at 10 weeks of age (250–300 g) were housed in the institutional vivarium with free access to food (Delta Feeds LbK 120 S-19, BioPro, Novosibirsk, Russia) and water. Single atrial and ventricular myocytes were isolated as previously described [ 56 ]. Briefly, rats were deeply anesthetized with an intramuscular injection of 2% Xylazine (1 mL/kg body weight, Alfasan, Woerden, Netherlands) and Zoletil-100 (0.3 mL/kg body weight, Virbac, Carros, France), heparinized (5000 IU/kg, Ellara, Pokrov, Russia), and euthanized by exsanguination.…”

Section: Methodsmentioning

confidence: 99%

The Acute Effects of Leptin on the Contractility of Isolated Rat Atrial and Ventricular Cardiomyocytes

Khokhlova

Myachina

Butova

et al. 2022

IJMS

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Leptin is a pleiotropic peptide playing an important role in the regulation of cardiac functions. It is not clear whether leptin directly modulates the mechanical function of atrial cardiomyocytes. We compared the acute effects of leptin on the characteristics of mechanically non-loaded sarcomere shortening and cytosolic Ca2+ concentration ([Ca2+]i) transients in single rat atrial and ventricular cardiomyocytes. We also studied the functional properties of myosin obtained from cardiomyocytes using an in vitro motility assay and assessed the sarcomeric protein phosphorylation. Single cardiomyocytes were exposed to 5, 20, and 60 nM leptin for 60 min. In ventricular cardiomyocytes, 60 nM leptin depressed sarcomere shortening amplitude and decreased the rates of shortening and relaxation. These effects were accompanied by a decrease in the phosphorylation of cMyBP-C, an increase in Tpm phosphorylation, and a slowdown of the sliding velocity of thin filaments over myosin in the in vitro motility assay. In contrast, in atrial cardiomyocytes, the phosphorylation of cMyBP-C and TnI increased, and the characteristics of sarcomere shortening did not change. Leptin had no effect on the characteristics of [Ca2+]i transients in ventricular cardiomyocytes, while 5 nM leptin prolonged [Ca2+]i transients in atrial cardiomyocytes. Thus, leptin-induced changes in contractility of ventricular cardiomyocytes may be attributed to the direct effects of leptin on cross-bridge kinetics and sarcomeric protein properties rather than changes in [Ca2+]i. We also suggest that the observed differences between atrial and ventricular cardiomyocytes may be associated with the peculiarities of the expression of leptin receptors, as well as signaling pathways in the atrial and ventricular myocardium.

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A combined Langendorff-injection technique for simultaneous isolation of single cardiomyocytes from atria and ventricles of the rat heart

Cited by 12 publications

References 17 publications

Peculiarities of the Acetylcholine Action on the Contractile Function of Cardiomyocytes from the Left and Right Atria in Rats

Peculiarities of the Acetylcholine Action on the Contractile Function of Cardiomyocytes from the Left and Right Atria in Rats

Contractile State Dependent Sarcomere Length Variability in Isolated Guinea-Pig Cardiomyocytes

The Acute Effects of Leptin on the Contractility of Isolated Rat Atrial and Ventricular Cardiomyocytes

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