Synthesis of guinea-pig cardiac myosin as measured by constant infusion.
Ontology highlight
ABSTRACT:
SUBMITTER: Wyborny LE
PROVIDER: S-EPMC1183879 | biostudies-other | 1978 Jan
REPOSITORIES: biostudies-other
Similar Datasets
Project description:BackgroundSeveral chronic conditions leading to skeletal muscle dysfunction are known to be associated with changes in the expression of myosin heavy chain (MHC) isoforms at both the mRNA and protein level. Many of these conditions are modelled, pre-clinically, in the guinea pig due to similar disease onset and progression to the human condition, and their generally well-characterised anatomy. MHC composition is amenable to determination by protein and mRNA based methodologies, the latter quantifying the expression of MHC isoform-specific gene transcripts allowing the detection of earlier, and more subtle changes. As such, the MHC mRNAs, and specific oligonucleotide primers of all common laboratory species have been available for some time. However, due to incomplete genomic annotation, assessment of guinea pig MHC mRNA expression has not been previously possible, precluding the full characterisation of early changes in skeletal muscle in response to disease and disease modulation.The purpose of this study was to characterise the multigenic structure of the sarcomeric MHC family in the guinea pig, and to design and validate specific oligonucleotide primers to enable the assessment of the predominant adult-muscle associated MHC mRNAs in relevant disease models.ResultsUsing a combination of ligase-mediated rapid amplification of 5' and 3' cDNA ends (RACE) and bioinformatics, mRNAs to the four main skeletal-muscle isoforms of MHC were determined. Specific oligonucleotide primers were designed, and following verification of their specificity, found to successfully determine the expression of each MHC mRNA independently.ConclusionsBecause of their utilisation in the in vivo modelling of disease, there is a requirement to develop molecular methods that accurately differentiate the different MHC mRNAs in the guinea pig to enable rapid profiling of muscle composition in appropriate disease models. The methods developed here are suitable for the characterisation of muscle MHC expression at the molecular level from animal tissue samples and biopsy material. The publication of these specific oligonucleotide primers for the guinea pig MHC variants will enable researchers to rapidly and accurately quantify acute changes in MHC mRNA expression in either developmental or in guinea pig disease models where a marker of altered skeletal muscle function is required.
| S-EPMC2911452 | biostudies-literature
Project description:Under high Ca(2+) load conditions, Ca(2+) concentrations in the extra-mitochondrial and mitochondrial compartments do not display reciprocal dynamics. This is due to a paradoxical increase in the mitochondrial Ca(2+) buffering power as the Ca(2+) load increases. Here we develop and characterize a mechanism of the mitochondrial Ca(2+) sequestration system using an experimental data set from isolated guinea pig cardiac mitochondria. The proposed mechanism elucidates this phenomenon and others in a mathematical framework and is integrated into a previously corroborated model of oxidative phosphorylation including the Na(+)/Ca(2+) cycle. The integrated model reproduces the Ca(2+) dynamics observed in both compartments of the isolated mitochondria respiring on pyruvate after a bolus of CaCl2 followed by ruthenium red and a bolus of NaCl. The model reveals why changes in mitochondrial Ca(2+) concentration of Ca(2+) loaded mitochondria appear significantly mitigated relative to the corresponding extra-mitochondrial Ca(2+) concentration changes after Ca(2+) efflux is initiated. The integrated model was corroborated by simulating the set-point phenomenon. The computational results support the conclusion that the Ca(2+) sequestration system is composed of at least two classes of Ca(2+) buffers. The first class represents prototypical Ca(2+) buffering, and the second class encompasses the complex binding events associated with the formation of amorphous calcium phosphate. With the Ca(2+) sequestration system in mitochondria more precisely defined, computer simulations can aid in the development of innovative therapeutics aimed at addressing the myriad of complications that arise due to mitochondrial Ca(2+) overload.
| S-EPMC3615037 | biostudies-literature
Project description:Otitis media (OM) is an inflammatory or infectious disease of the middle ear. Acute otitis media (AOM) and otitis media with effusion (OME) are the two major types of OM. However, the tympanic membrane (TM) motion differences induced by AOM and OME have not been quantified in animal models in the literature. In this study, the guinea pig AOM and OME models were created by transbullar injection of Streptococcus pneumoniae type 3 and lipopolysaccharide, respectively. To explore the effects of OM on the entire TM vibration, the measurements of full-field TM motions were performed in the AOM, OME and untreated control ears by using scanning laser Doppler vibrometry (SLDV). The results showed that both AOM and OME generally reduced the displacement peak and produced the traveling-wave-like motions at relatively low frequencies. Compared with the normal ear, OME resulted in a significant change of the TM displacement mainly in the inferior portion of the TM, and AOM significantly affected the surface motion across four quadrants. The SLDV measurements provide more insight into sound-induced TM vibration in diseased ears.
| S-EPMC5018450 | biostudies-literature
Project description:We previously reported in guinea pig tracheal smooth muscle that maximal shortening velocity decreases from 3 weeks of age to adulthood. It is not known whether myosin light chain kinase (MLCK), a key enzyme determining the velocity of smooth muscle contraction, undergoes maturational changes. In the present work, we investigated MLCK protein content and mRNA expression in 1-week-old, 3-week-old, and adult guinea pigs. We extracted either proteins or RNA from isolated tracheal smooth muscle. The content of MLCK was assessed by Western immunoblots. MLCK mRNA was evaluated by Northern analysis and by quantitative real time reverse transcriptase-polymerase chain reaction (RT-PCR). The content of MLCK increased 3-fold at 3 weeks of age and then decreased in adults, being 0.116 +/- 0.042, 0.330 +/- 0.125 (P < 0.05), and 0.153 +/- 0.054 microg/mg of total protein, respectively, in 1-week, 3-week, and adult animals. Quantitative RT-PCR revealed that MLCK mRNA increased with age to 135 +/- 35% and 177 +/- 23% (P < 0.01) in 3-week and adult animals, respectively, compared to 1-week animals. The transient increase of MLCK content in juvenile guinea pig tracheal smooth muscle may contribute to the increased shortening velocity at this age. We suggest that this increased content of MLCK is one of the mechanisms leading to maturation of airway smooth muscle contractility, which in turn contributes to the airway hyperresponsiveness reported in children and young animals.
| S-EPMC2527445 | biostudies-literature
Project description:1. The synthesis of phosphoenolpyruvate and the O(2) consumption from the tricarboxylic acid-cycle intermediates citrate, alpha-oxoglutarate, malate and succinate by guinea-pig mitochondria were compared. Malate was the most effective of these precursors; there was no synthesis of phosphoenolpyruvate from succinate. 2. The addition of palmitate, acetoacetate and ATP enhanced the synthesis of phosphoenolpyruvate from citrate and alpha-oxoglutarate. Palmitate and ATP increased the O(2) consumption, whereas acetoacetate had no effect on this parameter. 3. Octanoate depressed the synthesis of phosphoenolpyruvate from citrate, alpha-oxoglutarate and malate and increased the O(2) consumption. Pentenoic acid had no effect on phosphoenolpyruvate synthesis from any of the substrates used, although it increased the uptake of O(2). These findings might be relevant to the control of gluconeogenesis in vivo.
| S-EPMC1177620 | biostudies-other
Project description:Here, we examine key regulatory pathways underlying the transition from compensated hypertrophy (HYP) to decompensated heart failure (HF) and sudden cardiac death (SCD) in a guinea pig pressure-overload model by integrated multiome analysis. Relative protein abundances from sham-operated HYP and HF hearts were assessed by iTRAQ LC-MS/MS. Metabolites were quantified by LC-MS/MS or GC-MS. Transcriptome profiles were obtained using mRNA microarrays. The guinea pig HF proteome exhibited classic biosignatures of cardiac HYP, left ventricular dysfunction, fibrosis, inflammation, and extravasation. Fatty acid metabolism, mitochondrial transcription/translation factors, antioxidant enzymes, and other mitochondrial procsses, were downregulated in HF but not HYP. Proteins upregulated in HF implicate extracellular matrix remodeling, cytoskeletal remodeling, and acute phase inflammation markers. Among metabolites, acylcarnitines were downregulated in HYP and fatty acids accumulated in HF. The correlation of transcript and protein changes in HF was weak (R(2) = 0.23), suggesting post-transcriptional gene regulation in HF. Proteome/metabolome integration indicated metabolic bottlenecks in fatty acyl-CoA processing by carnitine palmitoyl transferase (CPT1B) as well as TCA cycle inhibition. On the basis of these findings, we present a model of cardiac decompensation involving impaired nuclear integration of Ca(2+) and cyclic nucleotide signals that are coupled to mitochondrial metabolic and antioxidant defects through the CREB/PGC1? transcriptional axis.
| S-EPMC5779628 | biostudies-literature
Project description:Cardiac output is regulated by the coordinate interactions of stimulatory sympathetic and inhibitory parasympathetic signals. Intracardiac parasympathetic ganglia are integrative centers of cardiac regulation, and modulation of the parasympathetic drive on the heart is accomplished by altering intrinsic cardiac ganglion neuron excitability. The pituitary adenylate cyclase-activating polypeptide (PACAP)/vasoactive intestinal peptide (VIP) family of peptides modulates cardiac function, and in guinea pig heart, PACAP appears to act directly on intrinsic parasympathetic cardiac ganglia neurons through PACAP-selective receptors. A multidisciplinary project tested whether cardiac PACAP peptides act through PACAP-selective receptors as excitatory neuromodulators amplifying the parasympathetic inhibition from guinea pig cardiac ganglia. The in vivo sources of regulatory PACAP peptides were localized immunocytochemically to neuronal fibers and a subpopulation of intrinsic postganglionic cardiac neurons. RT-PCR confirmed that cardiac ganglia expressed proPACAP transcripts and have PACAP peptide biosynthetic capabilities. Messenger RNA encoding PACAP-selective PAC1 receptor isoforms were also present in cardiac ganglia. Alternative splicing of PAC1 receptor transcripts produced predominant expression of the very short variant with neither HIP nor HOP cassettes; lower levels of the PAC1HOP2 receptor mRNA were present. Almost all of the parasympathetic neurons expressed membrane-associated PAC1 receptor proteins, localized immunocytochemically, which correlated with the population of cells that responded physiologically to PACAP peptides. PACAP depolarized cardiac ganglia neurons and increased neuronal membrane excitability. The rank order of peptide potency on membrane excitability in response to depolarizing currents was PACAP27>PACAP38>VIP. The PACAP-induced increase in excitability was not a function of membrane depolarization nor was it caused by alterations in action potential configuration. These results support roles for PACAP peptides as integrative modulators amplifying, through PACAP-selective receptors, the parasympathetic cardiac ganglia inhibition of cardiac output.
| S-EPMC6793321 | biostudies-literature
Project description:Ischemia-induced shortening of the cardiac action potential and its heterogeneous recovery upon reperfusion are thought to set the stage for reentrant arrhythmias and sudden cardiac death. We have recently reported that the collapse of mitochondrial membrane potential (DeltaPsi(m)) through a mechanism triggered by reactive oxygen species (ROS), coupled to the opening of sarcolemmal ATP-sensitive potassium (K(ATP)) channels, contributes to electrical dysfunction during ischemia-reperfusion. Here we present a computational model of excitation-contraction coupling linked to mitochondrial bioenergetics that incorporates mitochondrial ROS-induced ROS release with coupling between the mitochondrial energy state and electrical excitability mediated by the sarcolemmal K(ATP) current (I(K,ATP)). Whole-cell model simulations demonstrate that increasing the fraction of oxygen diverted from the respiratory chain to ROS production triggers limit-cycle oscillations of DeltaPsi(m), redox potential, and mitochondrial respiration through the activation of a ROS-sensitive inner membrane anion channel. The periods of transient mitochondrial uncoupling decrease the cytosolic ATP/ADP ratio and activate I(K,ATP), consequently shortening the cellular action potential duration and ultimately suppressing electrical excitability. The model simulates emergent behavior observed in cardiomyocytes subjected to metabolic stress and provides a new tool for examining how alterations in mitochondrial oxidative phosphorylation will impact the electrophysiological, contractile, and Ca(2+) handling properties of the cardiac cell. Moreover, the model is an important step toward building multiscale models that will permit investigation of the role of spatiotemporal heterogeneity of mitochondrial metabolism in the mechanisms of arrhythmogenesis and contractile dysfunction in cardiac muscle.
| S-EPMC2756351 | biostudies-literature
Project description:Omecamtiv mecarbil (OM) is a pharmacological agent that augments cardiac contractile function by enhancing myofilament Ca2+ sensitivity. Given that interventions that increase myofilament Ca2+ sensitivity have the potential to alter length-dependent activation (LDA) of cardiac myofilaments, we tested the influence of OM on this fundamental property of the heart. This is significant not only because LDA is prominent in cardiac muscle but also because it contributes to the Frank-Starling law, a mechanism by which the heart increases stroke volume in response to an increase in venous return. We measured steady-state and dynamic contractile indices in detergent-skinned guinea pig (Cavia porcellus) cardiac muscle fibers in the absence and presence of 0.3 and 3.0 ?M OM at two different sarcomere lengths (SLs), short SL (1.9 ?m) and long SL (2.3 ?m). Myofilament Ca2+ sensitivity, as measured by pCa50 (-log of [Ca2+]free concentration required for half-maximal activation), increased significantly at both short and long SLs in OM-treated fibers when compared to untreated fibers; however, the magnitude of increase in pCa50 was twofold greater at short SL than at long SL. A consequence of this greater increase in pCa50 at short SL was that pCa50 did not increase any further at long SL, suggesting that OM abolished the SL dependency of pCa50. Furthermore, the SL dependency of rate constants of cross-bridge distortion dynamics (c) and force redevelopment (ktr) was abolished in 0.3-?M-OM-treated fibers. The negative impact of OM on the SL dependency of pCa50, c, and ktr was also observed in 3.0-?M-OM-treated fibers, indicating that cooperative mechanisms linked to LDA were altered by the OM-mediated effects on cardiac myofilaments.
| S-EPMC5567428 | biostudies-literature
Project description:1. Acetate-CoA ligase, acetyl-CoA-carbon dioxide ligase and fatty acid synthetase were shown to be present in particle-free fractions of guinea-pig intestinal mucosa. 2. Each of these enzymes was partially purified by ammonium sulphate precipitation from the particle-free supernatant. 3. The incorporation of acetate and citrate into fatty acid was measured. 4. Gas-liquid radiochromatography was used to investigate the pattern of fatty acids synthesized. 5. The rate-limiting step in fatty acid synthesis was shown to be acetyl-CoA-carbon dioxide ligase.
| S-EPMC1198368 | biostudies-other