Decrease In Frequency Of Contractions Research Articles

Flow-dependent regulation of rat mesenteric lymphatic vessel contractile response requires activation of endothelial TRPV4 channels.

The objective of our study is to evaluate the involvement of the transient receptor potential vanilloid 4 (TRPV4) in the alteration of lymphatic pumping in response to flow and determine the signaling pathways involved. We used immunofluorescence imaging and western blotting to assess TRPV4 expression in rat mesenteric lymphatic vessels. We examined inhibition of TRPV4 with HC067047, nitric oxide synthase (NOS) with L-NNA and cyclooxygenases (COXs) with indomethacin on the contractile response of pressurized lymphatic vessels to flow changes induced by a stepwise increase in pressure gradients, and the functionality of endothelial TRPV4 channels by measuring the intracellular Ca2+ response of primary lymphatic endothelial cell cultures to the selective agonist GSK1016790A. TRPV4 protein was expressed in both the endothelial and the smooth muscle layer of rat mesenteric lymphatics with high endothelial expression around the valve sites. When maintained under constant transmural pressure, most lymphatic vessels displayed a decrease in contraction frequency under conditions of flow and this effect was ablated through inhibition of NOS, COX or TRPV4. Our findings demonstrate a critical role for TRPV4 in the decrease in contraction frequency induced in lymphatic vessels by increases in flow rate via the production and action of nitric oxide and dilatory prostanoids.

Notch signaling is a novel regulator of visceral smooth muscle cell differentiation in the murine ureter.

The contractile phenotype of smooth muscle cells (SMCs) is transcriptionally controlled by a complex of the DNA-binding protein SRF and the transcriptional co-activator MYOCD. The pathways that activate expression of Myocd and of SMC structural genes in mesenchymal progenitors are diverse, reflecting different intrinsic and extrinsic signaling inputs. Taking the ureter as a model, we analyzed whether Notch signaling, a pathway previously implicated in vascular SMC development, also affects visceral SMC differentiation. We show that mice with a conditional deletion of the unique Notch mediator RBPJ in the undifferentiated ureteric mesenchyme exhibit altered ureter peristalsis with a delayed onset, and decreased contraction frequency and intensity at fetal stages. They also develop hydroureter 2 weeks after birth. Notch signaling is required for precise temporal activation of Myocd expression and, independently, for expression of a group of late SMC structural genes. Based on additional expression analyses, we suggest that a mesenchymal JAG1-NOTCH2/NOTCH3 module regulates visceral SMC differentiation in the ureter in a biphasic and bimodal manner, and that its molecular function differs from that in the vascular system.

P–362 The effect of nolasiban on uterine contractility at the time of embryo transfer in in vitro fertilisation patients

Abstract Study question What is the effect of nolasiban on the uterine contractility of in-vitro fertilisation (IVF) patients prior to embryo transfer (ET) ? Summary answer A single oral dose of nolasiban 900 mg administered 4 h before ET significantly decreased contraction frequency and increased coordination compared to placebo. What is known already Nolasiban is an investigational oral oxytocin receptor antagonist (OTRa) being developed to improve the chance of pregnancy following ET. Increased uterine contraction frequency can influence embryo implantation, and the coordination of these uterine contractions is equally important. OTRa have been shown to decrease uterine contractions and increase endometrial perfusion. Recently, an automated and quantitative measurement tool using transvaginal ultrasound (TVUS) to better characterise uterine contractility has been developed which can be used to quantify the effect of nolasiban on uterine contractility. Study design, size, duration This study is part of a completed multi-centre randomised placebo-controlled trial (IMPLANT 1 – NCT02310802) in IVF patients (n = 247) carried out in 2015. Our study retrospectively assessed a sub-set of patients with good quality TVUS recordings to evaluate their mechanical uterine motion that were randomised to receive either nolasiban 900mg (n = 39) or placebo (n = 42). Participants/materials, setting, methods Subjects were < 37 years, undergoing ET on Day 3 following IVF/ICSI and with evidence of uterine contractions 4 h before ET. Nolasiban was administered 4 h before ET. Patients underwent TVUS immediately before drug administration and again immediately before ET. Uterine contraction frequency, amplitude, power and coordination were measured by applying dedicated speckle tracking and strain analysis. The Shapiro–Wilk test, followed by the Wilcoxon rank-sum test were applied to compare features between treatment groups. Main results and the role of chance The mean (SD) frequency of uterine contractions was 1.54 (0.25) in the nolasiban group versus 1.57 (0.12) in the placebo group (p = 0.016). The mean (SD) coordination was 0.10 (0.17) in the nolasiban group versus 0.02 (0.16) in the placebo group (p = 0.034). The coordination feature was measured by assessing the presence of simultaneous movements of the anterior and posterior uterine walls, a higher value reflects increased coordination. There was no difference in contraction amplitude or power. Limitations, reasons for caution This was a retrospective analysis of TVUS videos. The small sample size limits the generalisability of the findings. Furthermore, our initial results do not show how the changes in uterine motion may affect pregnancy rate after ET, meaning that the clinical relevance of our results remains to be proven. Wider implications of the findings: Our results show that in patients taking one 900mg dose of nolasiban prior to ET, beneficial uterine contractions are seen, which could be promising for embryo implantation and pregnancy in IVF patients. Our quantitative TVUS measurement tool can be applied to different patient populations to accurately quantify uterine motion. Trial registration number NCT02310802

The mechanism of action of oxytocin antagonist nolasiban in ART in healthy female volunteers

Research questionWhat are the effects of the oxytocin receptor (OTR) antagonist nolasiban on uterine contractions, endometrial perfusion and endometrial mRNA expression? DesignRandomized, double-blind, parallel-group, mechanism-of-action study with nolasiban. Forty-five healthy, pre-menopausal women were treated with placebo, 900 mg or 1800 mg nolasiban on the day corresponding to blastocyst transfer. Ultrasonographic uterine contraction frequency and endometrial perfusion were assessed, and endometrial biopsies analysed by next-generation sequencing. ResultsBoth doses of nolasiban showed decreased contraction frequency and increased endometrial perfusion depending on the time point assessed. At 1800 mg, 10 endometrial genes (DPP4, CNTNAP3, CNTN4, CXCL12, TNXB, CTSE, OLFM4, KRT5, KRT6A, IDO2) were significantly differentially expressed (adjusted P < 0.05). Of these, OLFM4, DPP4 and CXCL12 were regulated in the same direction as genes involved in implantation during the window of implantation. In addition, three genes (DPP4, CXCL12 and IDO2) were associated with decidualization and endometrial receptivity. ConclusionsThese data expand our knowledge of the mechanism of action of nolasiban in increasing pregnancy rates after embryo transfer. The results suggest more marked effects of nolasiban 1800 mg compared with the 900 mg dose, supporting testing at higher doses in IVF patients.

An Efficient Noninvasive Neuromodulation Modality for Overactive Bladder Using Time Interfering Current Method.

The present study aimed to evaluate a new tibial nerve stimulation (TNS) modality, which uses interferential currents, in terms of the stimulation electric field penetration efficiency into the body and physiological effectiveness. In silico experiments were performed to analyze the penetration efficiency of proposed interferential current therapy (ICT). Based on this, we performed in vivo experiments to measure excitation threshold of ICT for the tibial nerve, which is related to stimulation field near the nerve. Regarding analysis of the physiological effectiveness, in vivo ICT-TNS was performed, and changes in bladder contraction frequency and voiding volume were measured. The penetration efficiency and physiological effectiveness of ICT were evaluated by comparison with those of conventional TNS using transcutaneous electrical nerve stimulation (TENS). Simulation results showed that ICT has high penetration efficiency, thereby generating stronger field than TENS. These results are consistent with the in vivo results that nerve excitation threshold of ICT is lower than that of TENS. Moreover, ICT-TNS decreased contraction frequency and increased voiding volume, and its performance was profound compared with that of TENS-TNS. The proposed ICT is more efficient in inducing the stimulation field near the tibial nerve placed deep inside the body compared with conventional TENS and shows a good clinical effectiveness for TNS. The high efficiency of ICT increases the safety of noninvasive neurostimulation; therefore, it has clinical potential to become a promising modality for TNS to treat OAB and other peripheral neurostimulations.

Expression of ALS-linked SOD1 Mutation in Motoneurons or Myotubes Induces Differential Effects on Neuromuscular Function In vitro

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease that selectively affects upper and lower motoneurons. Dismantlement of the neuromuscular junction (NMJ) is an early pathological hallmark of the disease whose cellular origin remains still debated. We developed an in vitro NMJ model to investigate the differential contribution of motoneurons and muscle cells expressing ALS-causing mutation in the superoxide dismutase 1 (SOD1) to neuromuscular dysfunction. The primary co-culture system allows the formation of functional NMJs and fosters the expression of the ALS-sensitive fast fatigable type II-b myosin heavy chain (MHC) isoform. Expression of SOD1G93A in myotubes does not prevent the formation of a functional NMJ but leads to decreased contraction frequency and lowers the slow type I MHC isoform transcript levels. Expression of SOD1G93A in both motoneurons and myotubes or in motoneurons alone however alters the formation of a functional NMJ. Our results strongly suggest that motoneurons are a major factor involved in the process of NMJ dismantlement in an experimental model of ALS.

Monosodium glutamate impairs the contraction of uterine visceral smooth muscle ex vivo of rat through augmentation of acetylcholine and nitric oxide signaling pathways

The aim of the study was to examine the toxic effects of Monosodium glutamate (MSG), an extensively used food additive, on the contraction of uterine visceral smooth muscle (UVSM) in rat and to elucidate the probable neurocrine mechanism involved in it. MSG produced significant potentiation of the force and inhibition of frequency of uterus recorded ex vivo in chronic MSG exposure and in single dose acute experiments. MSG also produced significant potentiation of force of acetylcholine induced contraction and no alterations in atropine induced contraction of uterus. Further, MSG produced significant increase in force and frequency of contraction of neostigmine incubated uterus. We have found significant potentiation of the post pause force of contraction of uterus when MSG was applied in adrenaline incubated uterus. MSG also produced significant decrease in frequency of contraction of sodium nitroprusside incubated uterus; increase in frequency of N-ω-Nitro-l-Arginine Methyl Ester incubated uterus and no significant changes in frequency of contraction of methylene blue incubated uterus. These results indicate that MSG potentiates the force of contraction of UVSM predominantly by augmenting the activity of cholinergic intrinsic efferents and inhibits the frequency of contraction probably by augmenting the activity of nitrergic efferents. In conclusion, MSG potentiates the force and inhibits the frequency of contraction of UVSM, and the MSG induced effect is probably mediated through the augmentation of acetylcholine and nitric oxide signaling pathways.

A Validation Study of Near-Infrared Fluorescence Imaging of Lymphatic Vessels in Humans

Near-infrared fluorescence (NIRF) imaging is a new imaging technique that is used to visualize lymphatic vessels in humans. It has a high spatial and temporal resolution, allowing real-time visualization of lymphatic flow. The current study investigated the intra- and inter-individual variability of the technique, and how local hypo- and hyperthermia (20°C, 40°C), as well as exercise affect lymph transport. In this study, 10 healthy volunteers were studied twice, with 2 weeks between. NIRF imaging was conducted by using intradermal Indocyanine green injections and a custom-built camera setup. All data were blinded before analysis and presented as mean ± standard deviation. Mean contraction frequency and lymph propulsion velocity were 0.59 ± 0.13 minutes-1 and 1.51 ± 0.24 cm/s, respectively, with no significant difference during each 4 hours examination or between the two visits. The maximal pressure that the lymphatic flow in the vessels could overcome on test day 1 and 2 was 56 ± 9 mmHg and 57 ± 9 mmHg, respectively (p = 0.496). Local hyperthermia increased contraction frequency from 0.62 ± 0.4 minutes to 1.46 ± 0.5 minutes-1 (p < 0.05). Hypothermia caused no significant changes. Immediately after exercise (exercising at a simulated distance of 1.4 km on a cycle ergometer), an increase in lymph propulsion velocity from 1.5 ± 0.49 to 2.2 ± 0.63 cm/s was observed (p < 0.05); whereas contraction frequency was unaltered. A decrease in contraction frequency from 0.68 ± 0.25 minutes to 0.35 ± 0.19 minutes-1 was observed 10 minutes after exercise, without a change in velocity. NIRF imaging can be conducted for 4 hours without a change in lymphatic activity. Furthermore, it has the sensitivity to detect changes in lymphatic activity by local hyperthermia and exercise. No changes were seen after local hypothermia. Pumping pressure shows good repeatability, whereas the other parameters show poor repeatability.

Postprandial lymphatic pump function after a high-fat meal: a characterization of contractility, flow, and viscosity.

Dietary lipids are transported from the intestine through contractile lymphatics. Chronic lipid loads can adversely affect lymphatic function. However, the acute lymphatic pump response in the mesentery to a postprandial lipid meal has gone unexplored. In this study, we used the rat mesenteric collecting vessel as an in vivo model to quantify the effect of lipoproteins on vessel function. Lipid load was continuously monitored by using the intensity of a fluorescent fatty-acid analog, which we infused along with a fat emulsion through a duodenal cannula. The vessel contractility was simultaneously quantified. We demonstrated for the first time that collecting lymphatic vessels respond to an acute lipid load by reducing pump function. High lipid levels decreased contraction frequency and amplitude. We also showed a strong tonic response through a reduction in the end-diastolic and systolic diameters. We further characterized the changes in flow rate and viscosity and showed that both increase postprandially. In addition, shear-mediated Ca(2+) signaling in lymphatic endothelial cells differed when cultured with lipoproteins. Together these results show that the in vivo response could be both shear and lipid mediated and provide the first evidence that high postprandial lipid has an immediate negative effect on lymphatic function even in the acute setting.

Role of sex hormones in gastrointestinal motility in pregnant and non-pregnant rats.

To correlate gastric contractility, gastrointestinal transit, and hormone levels in non-pregnant (estrous cycle) and pregnant rats using noninvasive techniques. Female rats (n = 23) were randomly divided into (1) non-pregnant, (contractility, n = 6; transit, n = 6); and (2) pregnant (contractility, n = 5; transit, n = 6). In each estrous cycle phase or at 0, 7, 14, and 20 d after the confirmation of pregnancy, gastrointestinal transit was recorded by AC biosusceptometry (ACB), and gastric contractility was recorded by ACB and electromyography. After each recording, blood samples were obtained for progesterone and estradiol determination. In the estrous cycle, despite fluctuations of sex hormone levels, no significant changes in gastrointestinal motility were observed. Days 7 and 14 of pregnancy were characterized by significant changes in the frequency of contractions (3.90 ± 0.42 cpm and 3.60 ± 0.36 cpm vs 4.33 ± 0.25 cpm) and gastric emptying (168 ± 17 min and 165 ± 15 min vs 113 ± 15 min) compared with day 0. On these same days, progesterone levels significantly increased compared with control (54.23 ± 15.14 ng/mL and 129.96 ± 30.52 ng/mL vs 13.25 ± 6.31 ng/mL). On day 14, we observed the highest level of progesterone and the lowest level of estradiol compared with day 0 (44.3 ± 15.18 pg/mL vs 24.96 ± 5.96 pg/mL). Gastrointestinal motility was unaffected by the estrous cycle. In our data, high progesterone and low estradiol levels can be associated with decreased contraction frequency and slow gastric emptying.

Aging-related anatomical and biochemical changes in lymphatic collectors impair lymph transport, fluid homeostasis, and pathogen clearance.

The role of lymphatic vessels is to transport fluid, soluble molecules, and immune cells to the draining lymph nodes. Here, we analyze how the aging process affects the functionality of the lymphatic collectors and the dynamics of lymph flow. Ultrastructural, biochemical, and proteomic analysis indicates a loss of matrix proteins, and smooth muscle cells in aged collectors resulting in a decrease in contraction frequency, systolic lymph flow velocity, and pumping activity, as measured in vivo in lymphatic collectors. Functionally, this impairment also translated into a reduced ability for in vivo bacterial transport as determined by time-lapse microscopy. Ultrastructural and proteomic analysis also indicates a decrease in the thickness of the endothelial cell glycocalyx and loss of gap junction proteins in aged lymph collectors. Redox proteomic analysis mapped an aging-related increase in the glycation and carboxylation of lymphatic’s endothelial cell and matrix proteins. Functionally, these modifications translate into apparent hyperpermeability of the lymphatics with pathogen escaping from the collectors into the surrounding tissue and a decreased ability to control tissue fluid homeostasis. Altogether, our data provide a mechanistic analysis of how the anatomical and biochemical changes, occurring in aged lymphatic vessels, compromise lymph flow, tissue fluid homeostasis, and pathogen transport.

The influence of leukotrienes C4 and D4 on the contractility of an inflamed porcine uterus

The effect of leukotriene (LT) C4 (LTC4) and LTD4 on the contractility of an inflamed porcine uterus was investigated. On Day 3 of the estrous cycle (Day 0 of the study), either saline or Escherichia coli suspension was injected into each uterine horn. Although acute endometritis developed in all bacteria-inoculated gilts, a severe acute endometritis was noted more often on Day 8 than on Day 16. Myometrial and endometrial/myometrial strips were incubated with LTC4 or LTD4 alone, or together with a cysteinyl-LT receptor antagonist (BAY-u9773). Leukotriene C4 increased the contraction intensity in the saline- and bacteria-treated uteri on Day 8; however, its effect was lower in the myometrium of inflamed uteri. Contraction frequency was found to decrease in the saline-treated uteri as opposed to inflamed ones, in which it was elevated. On Day 16, contraction intensity increased in response to LTC4 in the saline-treated uteri but was reduced in the inflamed organs. The value of this parameter was lower in the inflamed uteri than that in the saline-treated ones. Leukotriene D4 (Days 8 and 16) augmented contractility in the saline-treated uteri, but despite increasing its intensity in the inflamed organs, it decreased contraction frequency. Leukotriene C4 or LTD4, added to BAY-u9773–pretreated saline- and bacteria-treated uteri on both days, decreased the contraction intensity. On Day 16 after treatment with BAY-u9773 and LTC4, contraction intensity in the endometrium/myometrium of the inflamed uteri was lower than that in the saline-treated organs. Data show that both LTC4 and LTD4 affect the contractility of inflamed porcine uteri, though LTC4 exerts a weaker contractile effect.

Related neuropeptides use different balances of unitary mechanisms to modulate the cardiac neuromuscular system in the American lobster, Homarus americanus.

To produce flexible outputs, neural networks controlling rhythmic motor behaviors can be modulated at multiple levels, including the pattern generator itself, sensory feedback, and the response of the muscle to a given pattern of motor output. We examined the role of two related neuropeptides, GYSDRNYLRFamide (GYS) and SGRNFLRFamide (SGRN), in modulating the neurogenic lobster heartbeat, which is controlled by the cardiac ganglion (CG). When perfused though an isolated whole heart at low concentrations, both peptides elicited increases in contraction amplitude and frequency. At higher concentrations, both peptides continued to elicit increases in contraction amplitude, but GYS caused a decrease in contraction frequency, while SGRN did not alter frequency. To determine the sites at which these peptides induce their effects, we examined the effects of the peptides on the periphery and on the isolated CG. When we removed the CG and stimulated the motor nerve with constant bursts of stimuli, both GYS and SGRN increased contraction amplitude, indicating that each peptide modulates the muscle or the neuromuscular junction. When applied to the isolated CG, neither peptide altered burst frequency at low peptide concentrations; at higher concentrations, SGRN decreased burst frequency, whereas GYS continued to have no effect on frequency. Together, these data suggest that the two peptides elicit some of their effects using different mechanisms; in particular, given the known feedback pathways within this system, the importance of the negative (nitric oxide) relative to the positive (stretch) feedback pathways may differ in the presence of the two peptides.

Medical Transfer of Patients in Preterm Labor: Treatments and Tocolytics

Objective. To examine the epidemiology, effectiveness, and safety of tocolytics, and the variation in use of standard treatments and predictive testing for women in preterm labor (PTL) transported to tertiary care. Methods. This was a health record review of consecutive PTL patients (<38 weeks gestation) transported to a tertiary care facility by Ontario's air and land critical care transport service between January 1, 2006 and January 1, 2011. The primary outcome was the effectiveness of tocolytics in decreasing the frequency of contractions and incidence of delivery. Secondary outcomes included the type of tocolytics used, adverse events (defined a priori), use of standard treatments (corticosteroid, antibiotic), and use of predictive tests (cervical length measurement, fetal fibronectin). We report descriptive statistics and relative risk of contractions decreasing with tocolytics with 95% confidence intervals. Results. Of the 510 transports reviewed, 488 met all inclusion criteria with the following characteristics: mean age 26.1 years, mean gestational age 31.2 weeks, mean transport time 80 minutes, 61.0% multiparous, 13.3% twins, mean initial dilatation 1.8 cm, contraction <8 min apart 67.7%, and 66.8% from Northern Ontario. Tocolytics were used in 206 (42.2%), with nonsteroidal anti-inflammatory drugs and nitroglycerine being used most frequently. Eleven (2.3%) patients delivered during transport, 4 (36.4%) of which received tocolytics with relative risk (RR) 0.8 (95%CI 0–2.0), and number needed to treat (NNT) 165 (31.5–299.3). Among the 311 (63.7%) patients for which change in contractions was documented, 140 (45%) received tocolytics and of these patients, contractions decreased in 94 (67%), with an RR 0.6 (95%CI 0.3–0.9) and NNT 4.6 (3.1–6.2). Adverse events were documented in 67 (14%) patients (most commonly tachycardia 5.8%). Steroids were appropriately used in 268 (54.9%) patients and antibiotics were appropriately used in 286 (58.6%) patients. Predictive testing was performed in only 19 (3.9%) patients. Conclusions. Tocolytics were associated with decreased contraction frequency in more than two-thirds of those treated. However, nearly half those who did not receive the treatment also had decreased contractions, and the effect did not reach statistical significance. Variation in the type of tocolytic drug used suggests a need for clinical practice guidelines for tocolytic use during transport of PTL patients.

Dexmedetomidine modifies uterine contractions in pregnancy terms of rats

Objectives:The present study was aimed at determining the effective doses of Dexmedetomidine (Dex) involved in amplitude of contraction-force and frequency of uterine rings in pregnancy terms of rats. All experiments involving animal subjects were carried out with the approval of animal care and use Ethical Committee of Cukurova University. Experiments were performed on female Albino-Wistar rats (200-260 g; n = 40).Materials and Methods:Uterine rings from pregnant rats were placed in organ bath with Krebs and calcium ion (Ca2+)-free solutions to record and exposed to serially increasing log10 concentrations of Dex.Results:In Krebs solution, while Dex caused an increase in the spontaneous contraction-forces in all pregnancy terms of rats in a significant dose-dependent manner, it led to a decrease in contraction-frequency in late-pregnancy term of rats. In Ca2+ -free, the spontaneous contraction-force decreased in late-pregnancy term and increased in early and middle-pregnancy terms. In addition, while Dex increased the contraction-frequency in early and middle-pregnancy terms, it decreased in late-pregnancy term in a dose-dependent manner.Statistical Analysis Used:The data were subjected to one-way analysis of variance. Repeated measures were employed for comparison of several group means through the Tukey post-hoc test (SPSS 10.00 Inc., Chicago, Ill, USA). P < 0.05 was considered statistically significant.Conclusions:This study suggested that Dex might differently alter the spontaneous contraction-forces and contraction-frequencies of uterine rings in all pregnancy terms of rats in Krebs and Ca2+-free solutions.

Endothelin-1 and angiotensin II modulate rate and contraction amplitude in a subpopulation of mouse embryonic stem cell-derived cardiomyocyte-containing bodies

Embryonic stem cell-derived cardiomyocytes (ESC-CMs) have applications in understanding cardiac disease pathophysiology, pharmacology, and toxicology. Comprehensive characterization of their basic physiological and pharmacological properties is critical in determining the suitability of ESC-CMs as models of cardiac activity. In this study we use video microscopy and quantitative PCR to investigate the responses of mouse ESC-CMs to adrenoceptor, muscarinic, angiotensin II (Ang II), and endothelin-1 (ET-1) receptor activation. Isoprenaline (10 nM–10 μM) increased beating rate and contraction amplitude in all beating bodies (BBs), whereas carbachol (up to 1 μM) and the I f channel blocker ZD-7288 (10 μM) decreased contraction frequency. ET-1 (0.01–100 nM) reduced contraction amplitude in all BBs and increased contraction frequency in 50% of BBs; these effects were blocked by the ET A receptor antagonist BQ123 (250 nM). Ang II (0.01 nM–1 μM) increased both contraction amplitude (all BBs) and frequency (in 50% of BBs), effects blocked, respectively, by losartan (100 nM) and PD123,319 (200 nM). These results indicate the presence of functional ET A and both AT 1 and AT 2 receptors in murine ESC-CMs, but their expression and or activity appears to be evident only in a limited set of BBs.

Effect of thyroid motilin on migrating motor complex in fasting rats

Objective:To investigate the effects of thyroid motilin(MTL) on migrating motor complex(MMC) in fasting rats.Methods: The distribution and quantitative expression of motilin in the thyroids were detected by immunohistochemistry and radioimmunoassay.We also observed the effect of thyriod motilin on the MMC by recording the gastrointestinal motility in conscious rats.Results: Tail vein injection of MTL increased MMC in antrum and duodenum in a dose-dependent manner,as manifested by increased contraction frequency and amplitude(P0.05,P0.01).However,the contraction frequency and amplitude were significantly decreased after injection of MTL-R antagonist GM109(P0.05).GM109 could obviously block the promoting effect of MTL on MMC.Removing left thyroid significantly increased MMC cycle,contraction amplitude,and decreased contraction frequency(P0.05).Injection of anti-MTL serum into the thyroids could significantly promote MMC and increase the frequency and amplitude of contraction(P0.05).Immunohistochemistry demonstrated that motilin immunoreactive(MTL-IR) cells,together with calcitonin(CT),were observed in the thyroids,indicating the expression of MTL in thyroid C-cells.Radioimmunoassay analyses displayed that MTL level in the duodenal mucosa was 128.02 and 100.57 times those in the antrum and thyroid,respectively.Conclusion: MTL is expressed in the thyroid C-cell and participates in the regulation of MMC.

Hawthorn (Crataegus monogyna Jacq.) extract exhibits atropine-sensitive activity in a cultured cardiomyocyte assay

Hawthorn (Crataegus spp.) plant extract is used as a herbal alternative medicine for the prevention and treatment of various cardiovascular diseases. Recently, it was shown that hawthorn extract preparations caused negative chronotropic effects in a cultured neonatal murine cardiomyocyte assay, independent of beta-adrenergic receptor blockade. The aim of this study was to further characterize the effect of hawthorn extract to decrease the contraction rate of cultured cardiomyocytes. To test the hypothesis that hawthorn is acting via muscarinic receptors, the effect of hawthorn extract on atrial versus ventricular cardiomyocytes in culture was evaluated. As would be expected for activation of muscarinic receptors, hawthorn extract had a greater effect in atrial cells. Atrial and/or ventricular cardiomyocytes were then treated with hawthorn extract in the presence of atropine or himbacine. Changes in the contraction rate of cultured cardiomyocytes revealed that both muscarinic antagonists significantly attenuated the negative chronotropic activity of hawthorn extract. Using quinuclidinyl benzilate, L-[benzylic-4,4'-(3)H] ([(3)H]-QNB) as a radioligand antagonist, the effect of a partially purified hawthorn extract fraction to inhibit muscarinic receptor binding was quantified. Hawthorn extract fraction 3 dose-dependently inhibited [(3)H]-QNB binding to mouse heart membranes. Taken together, these findings suggest that decreased contraction frequency by hawthorn extracts in neonatal murine cardiomyocytes may be mediated via muscarinic receptor activation.

Functional and Molecular Adaptation of Lymphatic Vessels

We developed a bovine model of mesenteric venous hypertension (MVH)by surgically occluding mesenteric veins for 3 days to investigate the functional and molecular adaptations of the mesenteric lymphatic vessels. The MVH model was characterized by an increase in mesenteric vein pressure from 10 to 30 mm Hg, a decrease in lymph protein concentration from 6 to 5 g/dL and an increase in intestinal water content from 72 to 78%. After 3‐days of MVH, isolated mesenteric lymphatic vessels demonstrated decreased contraction frequency, decreased ejection fraction and decreased stroke volume at each transmural pressure tested. Microarray analysis revealed that 117 genes were upregulated and 79 genes were down regulated in MVH lymphatics. A large number of the regulated genes are muscle contractile proteins, consistent with the changes in function. We further characterized changes in the mRNA of actin isoforms expressed in lymphatic muscle in response to venous hypertension. The smooth muscle actins (alpha and gamma) and skeletal actin were increased and cardiac actin decreased in lymphatics from the MHV model. Actin protein levels were analyzed by western blot analysis and revealed similar directional changes in response to MHV. These data suggest that lymphatic vessels undergo functional and molecular adaptation in response to changes in lymphdynamics in a time frame that may be important in resolution of edema. This work was supported by a LSI grant from Texas A&amp;M University and CDC grant 623086.

Characterization of the molecular and electrophysiological properties of the T‐type calcium channel in human myometrium

Rises in intracellular calcium are essential for contraction of human myometrial smooth muscle (HMSM) and hence parturition. The T-type calcium channel may play a role in this process. The aim was to investigate the role of the T-type calcium channel in HMSM by characterizing mRNA expression, protein localization, electrophysiological properties and function of the channel subunits Cav3.1(alpha1G), Cav3.2(alpha1H), and Cav3.3(alpha1I). QRT-PCR, immunohistochemistry, electrophysiology and invitro contractility were performed on human myometrial samples from term, preterm, labour and not in labour. QRT-PCR analysis of Cav3.1, Cav3.2 and Cav3.3 demonstrated expression of Cav3.1 and Cav3.2 with no significant change (P>0.05) associated with gestation or labour status. Immunohistochemistry localized Cav3.1 to myometrial and vascular smooth muscle cells whilst Cav3.2 localized to vascular endothelial cells and invading leucocytes. Voltage clamp studies demonstrated a T-type current in 55% of cells. Nickel block of T-type current was voltage sensitive (IC50 of 118.57+/-68.9 microM at -30 mV). Activation and inactivation curves of ICa currents in cells expressing T-type channels overlapped demonstrating steady state window currents at the resting membrane potential of myometrium at term. Current clamp analysis demonstrated that hyperpolarizing pulses to a membrane potential greater than -80 mV elicited rebound calcium spikes that were blocked reversibly by 100 microM nickel. Contractility studies demonstrated a reversible decrease in contraction frequency during application of 100 microM nickel (P<0.05). We conclude that the primary T-type subunit expressed in some MSMCs is Cav3.1. We found that application of 100 microM nickel to spontaneously contracting human myometrium reversibly slows contraction frequency.