Spontaneous Contractions Research Articles

Abstract 17931: Auxotonic MyoPod® Engineered Heart Tissues Enable Longitudinal Tracking of in vitro Development of Hypertrophic Cardiomyopathy

Introduction: Engineered heart tissues (EHTs) formed from human induced pluripotent stem cell (hiPSC)-derived cardiomyocytes carrying relevant mutations are a valuable tool for in vitro disease modeling and therapeutic testing. Tracking the emergence of a phenotype in vitro is key to targeting the optimal window to test therapeutic intervention. We hypothesized that auxotonic MyoPod® EHTs would enable longitudinal characterization with video analysis. Methods: Four auxotonic MyoPod® scaffold designs of decellularized porcine ventricular myocardium were seeded with wild type (WT) hiPSC-cardiomyocytes and human cardiac fibroblasts and cultured for 3 weeks. Mechanical testing was then performed on a MyoLab® instrument to obtain the contractile phenotype. One auxotonic MyoPod® design was then selected to form EHTs with WT or MYH7 R403Q +/- hiPSC-cardiomyocytes and fibroblasts. Videos of spontaneous contractions were obtained every 2-3 days. Results: All auxotonic MyoPod® scaffold designs tested (A-D) formed spontaneously contracting EHTs. In optimization studies, design C auxotonic MyoPod® EHTs were not significantly different from isometric control in any of the contractile metrics measured including twitch force (256.2 ± 25.8 μN vs 214.7 ± 78.1 μN) and time from peak force to 50% relaxation (103.8 ± 6.6 ms vs 111.2 ± 7.6 ms; control vs C, n=3 EHTs/group). When design C MyoPod® EHTs were seeded with WT and MYH7 R403Q +/- cardiomyocytes, video analysis showed slower relaxation time in MYH7 R403Q +/- EHTs that was significantly longer compared to WT control by one week of culture. Time to 90% relaxation (RT90) was lengthened by 38.8% ± 2.2% in MYH7 R403Q +/- (WT: n=7, MYH7 R403Q +/- : n=6; p<0.01). These video analysis results agree with MyoLab® testing data of isometric WT and MYH7 R403Q +/- EHTs where RT90 was slowed by 36.8% ± 0.1% in MYH7 R403Q +/- (WT: n=8, MYH7 R403Q +/- : n=5; p<0.01). Conclusion: In these studies, an auxotonic MyoPod® scaffold design was identified that does not alter the baseline contractile phenotype of EHTs, and it was used to track the development of relaxation deficit in MYH7 R403Q +/- EHTs over one week in culture. This technology can be used to track both the development of a disease phenotype and its response to therapies delivered in vitro.

Amyloid-β mediates intestinal dysfunction and enteric neurons loss in Alzheimer's disease transgenic mouse.

Alzheimer's disease (AD) is traditionally considered as a brain disorder featured by amyloid-β (Aβ) deposition. The current study on whether pathological changes of AD extend to the enteric nervous system (ENS) is still in its infancy. In this study, we found enteric Aβ deposition, intestinal dysfunction, and colonic inflammation in the young APP/PS1 mice. Moreover, these mice exhibited cholinergic and nitrergic signaling pathways damages and enteric neuronal loss. Our data show that Aβ42 treatment remarkably affected the gene expression of cultured myenteric neurons and the spontaneous contraction of intestinal smooth muscles. The intra-colon administration of Aβ42 induced ENS dysfunction, brain gliosis, and β-amyloidosis-like changes in the wild-type mice. Our results suggest that ENS mirrors the neuropathology observed in AD brains, and intestinal pathological changes may represent the prodromal events, which contribute to brain pathology in AD. In summary, our findings provide new opportunities for AD early diagnosis and prevention.

Polystyrene nanoplastics induce developmental impairments and vasotoxicity in zebrafish (Danio rerio)

The exponential use of plastics has significantly increased environmental pollution by nanoplastics (NPs). In the aquatic environment, NPs interact and bioaccumulate in the biota, posing a potential ecotoxicological risk. The present study investigated the developmental toxicity, vasotoxicity, cytotoxicity, ROS induction, and behavioral impairments in zebrafish (Danio rerio) exposed to environmentally relevant polystyrene NPs (PS-NPs) concentrations (0.04, 34 ng L-1, and 34 μg L-1) for 144 h through multiple biomarkers response (mortality, frequency of spontaneous contractions, heart rate, and morphological changes). Furthermore, vasotoxicity (head, yolk sac, tail, and branchial vessels) was evaluated using the transgenic zebrafish tg(Fli1:eGFP). Results showed that PS-NPs interacted mainly with zebrafish chorion, gills, tail, and larvae head. PS-NPs at 34 ng L-1 and 34 μg L-1 induced neurotoxicity (decreased frequency of spontaneous contractions), cardiotoxicity (bradycardia), and morphological changes in the eyes and head, indicating that PS-NPs induce developmental impairments in zebrafish. In addition, cytotoxicity in the caudal region (34 ng L-1), ROS production, decreased mean swimming speed, and distance covered were observed in all tested concentrations. PS-NPs also induced vasotoxicity (yolk sac region) in transgenic zebrafish. Overall, the present study demonstrates the harmful effects of PS-NPs on the early developmental stages of freshwater fish, indicating their environmental risk.

The Effects of Age on Prostatic Responses to Oxytocin and the Effects of Antagonists.

Benign prostatic hyperplasia (BPH) is an age-related enlargement of the prostate with urethral obstruction that predominantly affects the middle-aged and older male population, resulting in disruptive lower urinary tract symptoms (LUTS), thus creating a profound impact on an individual's quality of life. The development of LUTS may be linked to overexpression of oxytocin receptors (OXTR), resulting in increased baseline myogenic tone within the prostate. Thus, it is hypothesised that targeting OXTR using oxytocin receptor antagonists (atosiban, cligosiban, and β-Mercapto-β,β-cyclopentamethylenepropionyl1, O-Me-Tyr2, Orn8]-Oxytocin (ßMßßC)), may attenuate myogenic tone within the prostate. Organ bath and immunohistochemistry techniques were conducted on prostate tissue from young and older rats. Our contractility studies demonstrated that atosiban significantly decreased the frequency of spontaneous contractions within the prostate of young rats (**** p < 0.0001), and cligosiban (* p < 0.05), and ßMßßC (**** p < 0.0001) in older rats. Additionally, immunohistochemistry findings revealed that nuclear-specific OXTR was predominantly expressed within the epithelium of the prostate of both young (*** p < 0.001) and older rats (**** p < 0.0001). In conclusion, our findings indicate that oxytocin is a key modulator of prostate contractility, and targeting OXTR is a promising avenue in the development of novel BPH drugs.

Electroacupuncture inhibits PDK1/Akt/HCN4 pathway to improve neurogenic urinary retention in rats.

To observe the therapeutic effect of electroacupuncture (EA) on neurogenic urinary retention rats, so as to explore the underlying mechanism of EA in treating neurogenic urinary retention by focusing on 3-phosphoinositide-dependent protein kinase 1 (PDK1)/protein kinase B (Akt)/hyperpolarization activated cyclic nucleotide-gated cation channel 4 (HCN4) pathway. Female SD rats were randomly divided into sham operation, model, EA, PDK1 inhibitor, HCN4 blocker and EA + HCN4 blocker groups, with 20 rats in each group. The model of sacral spinal cord injury was established by modified Hassan Shaker spinal cord transection method. EA (2 Hz/15 Hz, 0.5 mA) was applied to "Zhongji" (CV3) and "Zhongliao" (BL33) for 20 min, once daily for 10 days. Rats of the PDK1 inhibitor group received intraperitoneal injection of OSU-03012 (20 mg/kg), and rats of the HCN4 blocker group received intraperitoneal injection of ivabradine (10 mg/kg), both once every other day for 10 days. The urodynamic indexes of rats were detected by multi-channel physiological recorder；muscle strip test was used to detect detrusor excitability；the morphological changes of bladder were observed by HE staining. Immunofluorescence double staining was used to detect the co-expression of HCN4 and C-Kit, a specific marker of interstitial cells of Cajal in bladder. Western blot was used to detect the expression of PDK1/Akt/HCN4 pathway proteins in bladder tissue and heat shock protein 27 (HSP27), a protein related to bladder contraction function. Compared with the sham operation group, the rats in the model group showed urinary dysfunction, decreased leak point pressure, isolated detrusor spontaneous contraction frequency, fluorescence intensity of C-Kit positive cells, HCN4+/C-Kit+ co-expression, HCN4 and p-HSP27/HSP27 protein expression in bladder tissue (P<0.05), and increased maximum bladder capacity and comp-liance, minimum tension during contraction of isolated detrusor, PDK1 and p-Akt/Akt protein expression in bladder tissue (P<0.05). Meanwhile, the above index were all reversed after EA and PDK1 inhibitor intervention (P<0.05). In comparison with the EA group, the rats had severe urinary dysfunction, the urine leakage point pressure, spontaneous contraction frequency, fluorescence intensity of C-Kit positive cells, the co-expression of HCN4+/C-Kit+, and the protein expression of HCN4 and p-HSP27/HSP27 were decreased (P<0.05), the maximum bladder capacity and compliance, the minimum tension during contraction of isolated detrusor, and the protein expression of PDK1 and p-Akt/Akt in bladder tissue were increased (P<0.05) in both HCN4 blocker and EA+HCN4 blocker groups. HE staining showed exfoliated bladder epithelium and disordered layers, vacuolization of bladder wall cells, with infiltration of neutrophils in mucosal and muscular layers in the model group, which were relatively milder in the EA and PDK1 inhibitor groups, but worse in the HCN4 blocker and EA + HCN4 blocker groups. EA can improve the urinary dysfunction in rats with neurogenic urinary retention, which may be related to its effect in inhibiting the activation of PDK1/Akt pathway, promo-ting HCN4-mediated detrusor excitatory contraction and urinary electrical signal activation.

Morphology, organo- and histometric features of the heart and lungs of a sexually mature domestic dog (Canis Lupus Familiaris L., 1758)

The cardiovascular system and respiratory organs in animals are interconnected, they perform extremely important functions for the vital activity of the organism, the main of which is gas exchange. Therefore, the study of the cardiovascular system and respiratory organs is an urgent issue of the present. The aim of the study was to morphologically evaluate the macro- and histological structures of the heart and lungs of the domestic dog. Comprehensive morphological methods of research were used: histological, anatomical, organ, histo-, cytometric, and statistical, which provided new data on the peculiarities of macro-, histo-, and cytomorphometric characteristics of the morphological structures of the heart and lungs. The dog heart has a rounded shape, its absolute weight is 167.58±9.46 g (without epicardial fat – 154.22±8.04 g), relative weight – 0.72±0.005%. It was found that cardiomyocytes of the left ventricle had the largest volume, the right ventricle – the smallest, and atrial cardiomyocytes – the smallest. At the same time, the nuclearcytoplasmic ratio of cardiomyocytes of the left ventricle is 0.0224±0.0076, the cardiomyocytes of the right ventricle have a greater value – 0.0275±0.0081 and the atrial cardiomyocytes have the highest value – 0.0367±0.0105. Such ambiguous cytometric parameters of cardiomyocytes are associated with the morphological and functional activity of the myocardial ventricular muscle tissue and its functional features inherent in spontaneous and rhythmic contractions, which result in blood flow through a closed system of vessels. The absolute weight of the domestic dog’s lungs is 201.3±18.4 g, the relative weight is 1.21±0.14%, the ratio of the absolute weight of the left to the right lung is 1:1.33. According to the asymmetry coefficient (1.37:1), the lungs of dogs are of the narrowed-elongated type. The connective tissue stroma of the lungs occupies 59.62±3.4%, the respiratory part – 40.38±2.6%. The data on the morphology of the heart and lungs of the domestic dog, including the results of the study of the macro- and microscopic structure of the organs under investigation, which are presented in the publication, are of great importance for histology and comparative anatomy, and also make a significant contribution to clinical veterinary medicine

Pico145 inhibits TRPC4-mediated mICAT and postprandial small intestinal motility

In intestinal smooth muscle cells, receptor-operated TRPC4 are responsible for the majority of muscarinic receptor cation current (mICAT), which initiates cholinergic excitation-contraction coupling. Our aim was to examine the effects of the TRPC4 inhibitor Pico145 on mICAT and Ca2+ signalling in mouse ileal myocytes, and on intestinal motility. Ileal myocytes freshly isolated from two month-old male BALB/c mice were used for patch-clamp recordings of whole-cell currents and for intracellular Ca2+ imaging using Fura-2. Functional assessment of Pico145’s effects was carried out by standard in vitro tensiometry, ex vivo video recordings and in vivo postprandial intestinal transit measurements using carmine red. Carbachol (50 µM)-induced mICAT was strongly inhibited by Pico145 starting from 1 pM. The IC50 value for the inhibitory effect of Pico145 on this current evoked by intracellularly applied GTPγS (200 µM), and thus lacking desensitisation, was found to be 3.1 pM, while carbachol-induced intracellular Ca2+ rises were inhibited with IC50 of 2.7 pM. In contrast, the current activated by direct TRPC4 agonist (-)-englerin A was less sensitive to the action of Pico145 that caused only ∼43 % current inhibition at 100 pM. The inhibitory effect developed rather slowly and it was potentiated by membrane depolarisation. In functional assays, Pico145 produced concentration-dependent suppression of both spontaneous and carbachol-evoked intestinal smooth muscle contractions and delayed postprandial intestinal transit. Thus, Pico145 is a potent GI-active small-molecule which completely inhibits mICAT at picomolar concentrations and which is as effective as trpc4 gene deficiency in in vivo intestinal motility tests.

Role of nitric oxide and calcium ions in the effects of hydrogen sulfide on contractile activity of rat jejunum

This study was performed to explore the role of nitric oxide, intracellular and extracellular calcium in the effects of hydrogen sulfide on spontaneous and carbachol-induced contractions of a rat jejunum preparation during a isometric contraction. Application of H2S donor, sodium hydrosulfide, led to a decrease in tonic tension, the amplitude and frequency of spontaneous contractions, as well as in the amplitude induced by carbachol, a nonspecific acetylcholine receptor agonist. Inhibiting the production of endogenous NO synthesis by with L-NAME, the effect of H2S donor remained unchanged, while in the presence of SNAP, a NO donor, the effects of NaHS on the amplitude of spontaneous and carbachol-induced contractions were less pronounced. Dantrolene, a ryanodine receptor inhibitor was used to stop a decrease in tonic tension in the presence of NaHS. The calcium-free solution reduced the inhibitory effect of NaHS on carbachol-induced contractions. This suggests that the inhibitory effect of H2S is associated with the dynamics of the intracellular concentration of calcium ions, and the interaction between NO and H2S occurs at the level of common targets of two gases.

Antispasmodic Effect of Alstonia boonei De Wild. and Its Constituents: Ex Vivo and In Silico Approaches.

Alstonia boonei, belonging to the family Apocynaceae, is one of the best-known medicinal plants in Africa and Asia. Stem back preparations are traditionally used as muscle relaxants. This study investigated the antispasmodic properties of Alstonia boonei Stem back and its constituents. The freeze-dried aqueous Stem back extract of A. boonei, as well as dichloromethane (DCM), ethyl acetate, and aqueous fractions, were evaluated for their antispasmodic effect via the ex vivo method. Two compounds were isolated from the DCM fraction using chromatographic techniques, and their antispasmodic activity was evaluated. An in silico study was conducted by evaluating the interaction of isolated compounds with human PPARgamma-LBD and human carbonic anhydrase isozyme. The Stem back crude extract, DCM, ethyl acetate, and aqueous fractions showed antispasmodic activity on high-potassium-induced (K+ 80 mM) contractions on isolated rat ileum with IC50 values of 0.03 ± 0.20, 0.02 ± 0.05, 0.03 ± 0.14, and 0.90 ± 0.06 mg/mL, respectively. The isolated compounds from the DCM fraction were β-amyrin and boonein, with only boonein exhibiting antispasmodic activity on both high-potassium-induced (IC50 = 0.09 ± 0.01 µg/mL) and spontaneous (0.29 ± 0.05 µg/mL) contractions. However, β-amyrin had a stronger interaction with the two proteins during the simulation. The isolated compounds boonein and β-amyrin could serve as starting materials for the development of antispasmodic drugs.

Prednisolone Accelerates Embryonic Development of Zebrafish via Glucocorticoid Receptor Signaling at Low Concentrations.

Synthetic glucocorticoids have been widely detected in aquatic ecosystems and may pose a toxicological risk to fish. In the present study, we described multiple end point responses of zebrafish to a commonly prescribed glucocorticoid, prednisolone (PREL), at concentrations between 0.001 and 9.26 μg/L. Of 23 end points monitored, 7 were affected significantly. Significant increases in the frequency of yolk extension formation, spontaneous contraction, heart rate, and ocular melanin density and significant decreases of ear-eye distance at PREL concentrations of 0.001 μg/L and above clearly pointed to the acceleration of embryonic development of zebrafish by PREL. Further confirmation came from the alterations in somite numbers, head-trunk angle, and yolk sac size, as well as outcomes obtained via RNA sequencing, in which signaling pathways involved in tissue/organ growth and development were highly enriched in embryos upon PREL exposure. In addition, the crucial role of glucocorticoid receptor (GR) for PREL-induced effects was confirmed by both, the coexposure to antagonist mifepristone (RU486) and GR-/- mutant zebrafish experiments. We further demonstrated similar accelerations of embryonic development of zebrafish upon exposure to 11 additional glucocorticoids, indicating generic adverse effect characteristics. Overall, our results revealed developmental alterations of PREL in fish embryos at low concentrations and thus provided novel insights into the understanding of the potential environmental risks of glucocorticoids.

H2O2 enhances the spontaneous phasic contractions of isolated human-bladder strips via activation of TRPA1 channels on sensory nerves and the release of substance P and PGE2

Several studies have indicated that reactive oxygen species (ROS) can lead to detrusor overactivity (DO), but the underlying mechanisms are not known. Hydrogen dioxide (H2O2) is used commonly to investigate the effects of ROS. In present study, we investigated the effects of H2O2 on phasic spontaneous bladder contractions (SBCs) of isolated human-bladder strips (iHBSs) and the underlying mechanisms. Samples of bladder tissue were obtained from 26 patients undergoing cystectomy owing to bladder cancer. SBCs of iHBSs were recorded in organ-bath experiments. H2O2 (1μM–10mM) concentration-dependently increased the SBCs of iHBSs. These enhancing effects could be mimicked by an agonist of transient receptor potential (TRP)A1 channels (allyl isothiocyanate) and blocked with an antagonist of TRPA1 channels (HC030031; 10 μM). H2O2 induced enhancing effects also could be attenuated by desensitizing sensory afferents with capsaicin (10 μM), blocking nerve firing with TTX (1 μM), blocking neurokinin effects with NK2 receptor antagonist (SR48968, 10 μM), and blocking PGE2 synthesis with indomethacin (10 μM), respectively. Our study: (i) suggests activation of TRPA1 channels on bladder sensory afferents, and then release of substance P or PGE2 from sensory nerve terminals, contribute to the H2O2-induced enhancing effects on SBCs of iHBSs; (ii) provides insights for the mechanisms underlying ROS leading to DO; (iii) indicates that targeting TRPA1 channels might be the promising strategy against overactive bladder in conditions associated with excessive production of ROS.

Prebiotic prevents the development of gastrointestinal motility disorders caused by omeprazole

The synthetic disaccharide lactulose, consisting of fructose and galactose, after oral administration in an unchanged form reaches the lower part of the gastrointestinal tract, where under the action of normal flora it is broken down into short-chain fatty acids that stimulate colon motility. The effect of lactulose on gastrointestinal tract motility in the conditions of its long-term suppression by omeprazole has not been investigated. We studied the influence of lactulose on spontaneous and carbachol-stimulated gastric and colonic motility in rats treated with omeprazole for 28 days. The animals were divided into 3 groups. The first group of animals served as a control. The animals in the second group were administered omeprazole intraperitoneally at a dose of 14 mg/kg orally once a day for 28 days. The animals in the third group were simultaneously injected intraperitoneally with omeprazole and prebiotic lactulose at a dose of 0.2 g/kg orally once a day for 28 days. On the day after the last injections of drugs, we investigated the spontaneous and carbachol-stimulated contractions in the stomach and colon by the balloon graphic method. It was found that the frequency of spontaneous and stimulated contractions in the stomach and colon did not change significantly after 28 days of omeprazole treatment. The amplitude and index of spontaneous and carbachol-stimulated contractions in the stomach and colon were significantly weaker compared to the control. One day after the 28-day simultaneous administration of omeprazole and lactulose the amplitude and index of spontaneous and stimulated contractions in the stomach and colon increased compared with the group of rats treated with omeprazole alone. We concluded that the positive effect of lactulose on gastric and colon motility is a result of the prebiotic properties of lactulose which leads to the normalization of the microbiocenosis in the gastrointestinal tract and the elimination of the inflammatory process in it.

L-Fucose promotes enteric nervous system regeneration in type 1 diabetic mice by inhibiting SMAD2 signaling pathway in enteric neural precursor cells

BackgroundDiabetes can lead to extensive damage to the enteric nervous system (ENS), causing gastrointestinal motility disorders. However, there is currently a lack of effective treatments for diabetes-induced ENS damage. Enteric neural precursor cells (ENPCs) closely regulate the structural and functional integrity of the ENS. L-Fucose, is a dietary sugar that has been showed to effectively ameliorate central nervous system injuries, but its potential for ameliorating ENS damage and the involvement of ENPCs in this process remains uncertain.MethodsGenetically engineered mice were generated for lineage tracing of ENPCs in vivo. Using diabetic mice in vivo and high glucose-treated primary ENPCs in vitro, the effects of L-Fucose on the injured ENS and ENPCs was evaluated by assessing gastrointestinal motility, ENS structure, and the differentiation of ENPCs. The key signaling pathways in regulating neurogenesis and neural precursor cells properties, transforming growth factor-β (TGF-β) and its downstream signaling pathways were further examined to clarify the potential mechanism of L-Fucose on the injured ENS and ENPCs.ResultsL-Fucose improved gastrointestinal motility in diabetic mice, including increased defecation frequency (p < 0.05), reduced total gastrointestinal transmission time (p < 0.001) and bead expulsion time (p < 0.05), as well as enhanced spontaneous contractility and electric field stimulation-induced contraction response in isolated colonic muscle strips (p < 0.001). The decrease in the number of neurons and glial cells in the ENS of diabetic mice were reversed by L-Fucose treatment. More importantly, L-Fucose treatment significantly promoted the proportion of ENPCs differentiated into neurons and glial cells both in vitro and in vivo, accompanied by inhibiting SMAD2 phosphorylation.ConclusionsL-Fucose could promote neurogenesis and gliogenesis derived from ENPCs by inhibiting the SMAD2 signaling, thus facilitating ENS regeneration and gastrointestinal motility recovery in type 1 diabetic mice.7Zzihak_jncvcnfCNwCPXsVideo

THU264 Enteric Activity Enhanced Ileal GLP-1 Secretion

Abstract Disclosure: Y. Chen: None. G. Jiao: None. Y. Lei: None. Y. Li: None. L. Sha: None. GLP-1 is a multifunctional endocrine hormone secreted by intestinal L cells in gastrointestinal tract contributing to the maintenance of glucose homeostasis. The distal part of small intestine such as ileum has a great density of L cells. In this study, we provided a direct evidence that enteric activity modulated GLP-1 secretion in rat ileum. Methods: A segment of rat ileum was dissected. The proximal section of the ileum segment was used to pace the enteric activity with an intestinal electrical stimulation(IES) designed from our previous intestinal myoelectric activity analyzing mimicking the ileum activity in feeding and the adjacent distal section was perfused vascularly to collect and measure GLP-1 secretion. Ileal motility in the distal section of the ileum segment was also recorded to monitor the enteric activity. The lumen of the ileum segment was filled with saline. Results: The frequency and the amplitude of spontaneous contraction in the distal ileum section was 14.14±2.82 cycles/min and 1.58±0.46 g (n=5). The area under curve (AUC) was 180.40±61.68 g·s. The GLP-1 secretion level in the distal ileum section was 15.47±0.46 pg/mL. With IES, The frequency was increased to 18.27±2.23 cycles/min (P&amp;lt;0.05, n = 5) and the AUC was increased to 303.64±79.75 (p&amp;lt;0.05). The amplitude had no significant change (P&amp;gt;0.05). The GLP-1 secretion was significantly increased to 24.71±2.47 pg/mL(p&amp;lt;0.05) with enhanced enteric activity. we provided a direct evidence to show the enhancement of GLP-1 secretion with enteric activity. Our result indicated that with feeding-simulated enteric activity, GLP-1 secretion could be elevated by around 60 percent in rat ileum. (supported by the National Natural Science Foundation of China, Grant No.81670492) Presentation: Thursday, June 15, 2023

Inhibitory effect and underlying mechanism of essential oil of Prangos ferulacea Lindl (L.) on spontaneous and induced uterine contractions in non-pregnant rats

Evidence suggests the use of natural compounds as support in the management of uterine contractility disorders. We recently demonstrated that the essential oil of Apiacea Prangos ferulacea (L.) (Prangoil) modulates intestinal smooth muscle contractility. Thus, we aimed to evaluate if Prangoil could also affect the contractility of uterine muscle in non-pregnant rat and to investigate the related action mechanism/s. The effects of the aromatic monoterpenes, β-ocimene and carvacrol, constituents of Prangoil, were also evaluated. Spontaneous contractions and contraction-induced by K+-depolarization and oxytocin in rat uterus were recorded in vitro, using organ bath technique. Prangoil reduced the amplitude of spontaneous contractions as well as responses to KCl and oxytocin. β-ocimene and carvacrol matched oil inhibitory effects. Prangoil effects were not affected by nitrergic and adenylyl cyclase inhibitors or non-specific potassium channel blocker, but they were reduced by nifedipine, L-type calcium channel inhibitor, or 2-aminoethoxydiphenylborate (2-APB), membrane-permeant inositol 1,4,5-triphosphate receptor inhibitor. The response to β-ocimene was reduced by nifedipine and by 2-APB (20 μM), whilst carvacrol inhibitory effect was attenuated only by nifedipine. In conclusion, Prangoil, and its components, β-ocimene and carvacrol, reduced spontaneous and KCl or oxytocin-induced contractions of rat myometrium, mainly modulating extracellular Ca2+ influx through L-Type channels and Ca2+ release from the intracellular store. Further studies could contribute to evaluate the potential use of Prangoil against disorders characterized by abnormal uterine contractions.

Electric field stimulation unmasks a subtle role for T-type calcium channels in regulating lymphatic contraction

We previously identified two isoforms of T-type, voltage-gated calcium (Cav3) channels (Cav3.1, Cav3.2) that are functionally expressed in murine lymphatic muscle cells; however, contractile tests of lymphatic vessels from single and double Cav3 knock-out (DKO) mice, exhibited nearly identical parameters of spontaneous twitch contractions as wild-type (WT) vessels, suggesting that Cav3 channels play no significant role. Here, we considered the possibility that the contribution of Cav3 channels might be too subtle to detect in standard contraction analyses. We compared the sensitivity of lymphatic vessels from WT and Cav3 DKO mice to the L-type calcium channel (Cav1.2) inhibitor nifedipine and found that the latter vessels were significantly more sensitive to inhibition, suggesting that the contribution of Cav3 channels might normally be masked by Cav1.2 channel activity. We hypothesized that shifting the resting membrane potential (Vm) of lymphatic muscle to a more negative voltage might enhance the contribution of Cav3 channels. Because even slight hyperpolarization is known to completely silence spontaneous contractions, we devised a method to evoke nerve-independent, twitch contractions from mouse lymphatic vessels using single, short pulses of electric field stimulation (EFS). TTX was present throughout to block the potential contributions of voltage-gated Na+ channels in perivascular nerves and lymphatic muscle. In WT vessels, EFS evoked single contractions that were comparable in amplitude and degree of entrainment to those occurring spontaneously. When Cav1.2 channels were blocked or deleted, only small residual EFS-evoked contractions (~ 5% of normal amplitude) were present. These residual, EFS-evoked contractions were enhanced (to 10–15%) by the KATP channel activator pinacidil (PIN) but were absent in Cav3 DKO vessels. Our results point to a subtle contribution of Cav3 channels to lymphatic contractions that can be unmasked in the absence of Cav1.2 channel activity and when the resting Vm is more hyperpolarized than normal.

The effect of abdominal bracing on respiration during a lifting task: a cross-sectional study

BackgroundAbdominal bracing is a maneuver widely used by rehabilitation specialists and sports trainers to improve spinal stability. This study aimed to investigate how lifting tasks with and without abdominal bracing affect the respiratory function of the diaphragm.MethodsM-mode ultrasonographic assessment of diaphragmatic motion combined with spirometry was performed on 31 healthy adults. Participants were asked to breathe continuously whilst lifting a load with spontaneous abdominal muscle contraction (natural loaded breathing) and abdominal bracing (AB loaded breathing).ResultsPearson’s correlations revealed strong correlations between ultrasonography and spirometry measures (p < 0.001) for all types of breathing: tidal breathing (r = 0.709, r2 = 0.503), natural loaded breathing (r = 0.731, r2 = 0.534) and AB loaded breathing (r = 0.795, r2 = 0.632). Using paired-samples t-tests, the natural loaded breathing ultrasonography revealed more caudal diaphragm positions during inspiration (p < 0.001) but not during expiration (p = .101). Spirometry demonstrated lower lung volumes (L) at the end of inspiration and expiration (p < 0.001), with no changes in total lung volume (p = 0.06). The AB loaded breathing ultrasonography revealed more caudal diaphragm positions during inspiration (p = 0.002) but not during expiration (p = 0.05). Spirometry demonstrated lower lung volumes at the end of inspiration (p < 0.001), expiration (p = 0.002), and total lung volumes (p = 0.019).ConclusionThis study demonstrated that abdominal bracing performed during a lifting task reduces lung volume despite an increase in diaphragmatic motion. Diaphragm excursions strongly correlate with lung volumes even under postural loading.Trial registrationThe study was prospectively registered on 8 April 2021 at ClinicalTrials.gov with identification number NCT04841109.

ERG K+ channels mediate a major component of action potential repolarization in lymphatic muscle

Smooth muscle cells in the walls of collecting lymphatic vessels fire spontaneous action potentials (APs), which conduct rapidly over the muscle layer to initiate contractions that propel lymph. Several ion channels have been implicated in the currents underlying the AP spike and the preceding diastolic depolarization, but the molecular identities of K+ channels involved in AP repolarization are unknown. Based on previous studies of other rhythmically active smooth muscles, we hypothesized that ether-a-go-go related gene (ERG) K+ channels (Kv11) play an important role in repolarization of the AP in lymphatic muscle. Message for one or more ERG channel isoforms was detected by RT-PCR analysis of lymphatic vessels from mice, rats and humans. Membrane potential recordings in smooth muscle cells of rat and human lymphatics revealed that nanomolar concentrations of ERG-1 inhibitors (E-4031 and BeKm-1) prolonged the duration of the AP plateau (normally ~ 1 s in duration) and induced multiple spikes, whereas ERG-1 activators (ICA-105574 and RPR-260243) shortened the plateau and could completely inhibit spontaneous APs. At relatively high inhibitor concentrations, the AP plateau duration lasted as long as 24 s. ERG activators reversed the effects of ERG inhibitors and vice-versa. In pressure myograph studies, ERG channel inhibition prolonged the diastolic repolarization phase of the contraction cycle and reduced the frequency of spontaneous contractions. This is the first evidence for a specific K+ channel contributing to the AP in lymphatic muscle. Our results imply that lymphatic contractile dysfunction may occur in long QT type II patients with mutations that result in ERG channel loss-of-function or impaired trafficking of the channel to the cell membrane.

Comparison of Intracervical Foley's Catheter With Vaginal Misoprostol Versus Intravaginal Misoprostol Alone for Cervical Ripening and Induction of Labor.

Introduction Induction of labor implies stimulation of contractions before the spontaneous onset of labor, with or without membranes. Augmentation refers to the enhancement of spontaneous contractions that are considered inadequate because of failed cervical and fetal descent. This study compared the effectiveness of intracervical Foley catheter insertion and vaginal misoprostol versus only vaginal misoprostol in the induction of labor and other outcomes relted to it. Methods The present study was a randomized controlled trial that included 148 women divided into two groups: (i) Group A, which received intracervical Foley catheter insertion and vaginal misoprostol (25 µg), and (ii) Group B, which received intravaginal administration of tablet misoprostol (25 µg) alone. We compared the median time from the time of induction to vaginal delivery, incidence of cesarean delivery, chorioamnionitis, puerperal infection, uterine tachysystole, neonatal information at delivery, and discharge status (i.e., birth weight, neonatal intensive care unit (NICU) admission, and neonatal death) between groups. Results We found that the rates of puerperal infection (n=36; 48.6%) and meconium-stained amniotic fluid (n=45; 60.8%) were higher in Group B than in Group A (n=20; 27.0% and n=25; 33.8%, respectively), which were statistically significant differences (p=0.0066 and p=0.0009, respectively). In addition, NICU admission was higher in Group B (n=47; 63.5%) than in Group A (n=30; 40.5%), which was a statistically significant difference (p=0.0051). Conclusion An intracervical Foley catheter with 25 µg of misoprostol was more effective for induction of labor than 25 µg of intravaginal misoprostol alone every six hours for a maximum of four doses in terms of induction to delivery interval, meconium-stained amniotic fluid, mode of delivery, intrapartum complications, and puerperal infection.

Modelling Lymph Propulsion in a 3D Model of Murine Collecting Vessel with Three Lymphangions in Series

The lymphatic system maintains tissue homeostasis by transporting the excess fluid from the interstitium and ultimately returning it to the venous circulation against an adverse pressure gradient and gravitational force. The spontaneous contractions of lymphangions, the building blocks of collecting vessels, and the secondary lymphatic valves play key roles in lymph propulsion. The aim of this study was to investigate lymph propulsion in a series of three contracting lymphangions in a 3D reconstructed model segmented from micro-CT scans of the collecting lymphatics in the hind limb of mice. Computational Fluid Dynamics and Fluid–Structure Interaction were used to study the behavior of flow within the collecting vessel, as well as the behavior and deformations of the vessel wall and the poroelastic interstitium. The secondary valves were modelled as porous membranes with closed or open states depending on their permeability. A sensitivity study revealed that the parameters having the most impact on the total volume of lymph propelled by active contraction of the lymphangions were the elastic modulus of the interstitium and the permeability of the secondary valves during the open states.