Smooth Muscle Membrane Research Articles

Contribution of transient receptor potential vanilloid 1 (TRPV1) channel to cholinergic contraction of rat bladder smooth muscle.

Transient receptor potential vanilloid 1 (TRPV1) is a calcium-permeable ion channel that is gated by the pungent constituent of red chili pepper, capsaicin, and by related chemicals from the group of vanilloids, in addition to noxious heat. It is expressed mostly in sensory neurons to act as a detector of painful stimuli produced by pungent chemicals and high temperatures. Although TRPV1 is also found outside the sensory nervous system, its expression and function in the bladder detrusor smooth muscle (DSM) remain controversial. Here, by using Ca2+ imaging and patch clamp on isolated rat DSM cells, in addition to tensiometry on multicellular DSM strips, we show that TRPV1 is expressed functionally in only a fraction of DSM cells, in which it acts as an endoplasmic reticulum Ca2+-release channel responsible for the capsaicin-activated [Ca2+]i rise. Carbachol-stimulated contractions of multicellular DSM strips contain a TRPV1-dependent component, which is negligible in the circular DSM but reaches ≤50% in the longitudinal DSM. Activation of TRPV1 in rat DSM during muscarinic cholinergic stimulation is ensured by phospholipase A2-catalysed derivation of arachidonic acid and its conversion by lipoxygenases to eicosanoids, which act as endogenous TRPV1 agonists. Immunofluorescence detection of TRPV1 protein in bladder sections and isolated DSM cells confirmed both its preferential expression in the longitudinal DSM sublayer and its targeting to the endoplasmic reticulum. We conclude that TRPV1 is an essential contributor to the cholinergic contraction of bladder longitudinal DSM, which might be important for producing spatial and/or temporal anisotropy of bladder wall deformation in different regions during parasympathetic stimulation. KEY POINTS: The transient receptor potential vanilloid 1 (TRPV1) heat/capsaicin receptor/channel is localized in the endoplasmic reticulum membrane of detrusor smooth muscle (DSM) cells of the rat bladder, operating as a calcium-release channel. Isolated DSM cells are separated into two nearly equal groups, within which the cells either show or do not show TRPV1-dependent [Ca2+]i rise. Carbachol-stimulated, muscarinic ACh receptor-mediated contractions of multicellular DSM strips contain a TRPV1-dependent component. This component is negligible in the circular DSM but reaches ≤50% in longitudinal DSM. Activation of TRPV1 in rat DSM during cholinergic stimulation involves phospholipase A2-catalysed derivation of arachidonic acid and its conversion by lipoxygenases to eicosanoids, which act as endogenous TRPV1 agonists.

Caveolae Disassembly using Methyl-β-cyclodextrin Causes the Abolition of Coupling of the Caveolae and the Sarcoplasmic Reticulum in the Rat Femoral Artery

Background: Caveolae are essential in regulating signal transduction mechanisms of ion channels in vascular tissue, including BKCa channels (maxi-K). The current study investigated the localization of maxi-K channels within caveolae. Methods: Sixteen rats were divided into two groups: A control group and a treated group, where arteries in the treated group were incubated with methyl-β-cyclodextrin (MβCD) to disassemble caveolae from artery tissue. Immunohistochemistry (IHC), immunocytochemistry (ICC), transmission electron microscopy (TEM) and Western blot techniques were used in this study. Result: IHC of intact arteries showed colocalization of maxi-K channels with caveolin-1 in smooth muscle and endothelial cells and colocalized of maxi-K channels with caveolin-3 in smooth muscle cells only. These findings were also corroborated with ICC in a single smooth muscle cell. TEM revealed caveolae covering most plasma membranes of smooth muscle and endothelial cells and showed that caveolae sit close to the sarcoplasmic reticulum only in smooth muscle cells. TEM showed incubating arteries with MβCD led to the disassembly of caveolae from artery tissue. This study concluded that maxi-K channels localize to caveolae and that caveolae abolishment by MβCD led to the abolition of the coupling of caveolae and the sarcoplasmic reticulum.

Extracellular glucose and dysfunctional insulin receptor signaling independently upregulate arterial smooth muscle TMEM16A expression.

Diabetes alters the function of ion channels responsible for regulating arterial smooth muscle membrane potential, resulting in vasoconstriction. Our prior research demonstrated an elevation of TMEM16A in diabetic arteries. Here, we explored the mechanisms involved in Transmembrane protein 16A (TMEM16A) gene expression. Our data indicate that a Snail-mediated repressor complex regulates arterial TMEM16A gene transcription. Snail expression was reduced in diabetic arteries while TMEM16A expression was upregulated. The TMEM16A promoter contained three canonical E-box sites. Electrophoretic mobility and super shift assays revealed that the -154 nt E-box was the binding site of the Snail repressor complex and binding of the repressor complex decreased in diabetic arteries. High glucose induced a biphasic contractile response in pressurized nondiabetic mouse hindlimb arteries incubated ex vivo. Hindlimb arteries incubated in high glucose also showed decreased phospho-protein kinase D1 and TMEM16A expression. In hindlimb arteries from nondiabetic mice, administration of a bolus dose of glucose activated protein kinase D1 signaling to induce Snail degradation. In both in vivo and ex vivo conditions, Snail expression exhibited an inverse relationship with the expression of protein kinase D1 and TMEM16A. In diabetic mouse arteries, phospho-protein kinase D1 increased while Akt2 and pGSK3β levels declined. These results indicate that in nondiabetic mice, high glucose triggers a transient deactivation of the Snail repressor complex to increase arterial TMEM16A expression independently of insulin signaling. Conversely, insulin resistance activates GSK3β signaling and enhances arterial TMEM16A channel expression. These data have uncovered the Snail-mediated regulation of arterial TMEM16A expression and its dysfunction during diabetes.NEW & NOTEWORTHY The calcium-activated chloride channel, TMEM16A, is upregulated in the diabetic vasculature to cause increased vasoconstriction. In this paper, we have uncovered that the TMEM16A gene expression is controlled by a Snail-mediated repressor complex that uncouples with both insulin-dependent and -independent pathways to allow for upregulated arterial protein expression thereby causing vasoconstriction. The paper highlights the effect of short- and long-term glucose-induced dysfunction of an ion channel expression as a causative factor in diabetic vascular disease.

Nebulized caffeine alleviates airway hyperresponsiveness in a murine asthma model.

The clinical definition of "difficult asthma" has expanded recently to include an ever-growing subset of patients with symptoms that cannot be controlled by conventional means, forcing the medical community to develop innovative therapeutics. Beneficial effects of coffee for subjects with asthma, primarily the effect of methylxanthine components, have long been described. Methylxanthines, including theophylline and caffeine, inhibit phosphodiesterases and downstream cAMP signaling to prevent mast cell degranulation while promoting immunomodulation (Peleman RA, Kips JC, Pauwels RA. Clin Exp Allergy 28: 53-56, 1998; Deshpande DA, Wang WCH, McIlmoyle EL, Robinett KS, Schillinger RM, An SS, Sham JSK, Liggett SB. Nat Med 16: 1299-1304, 2010). Caffeine is also a bitter taste receptor agonist, binding to taste-sensing type 2 receptors (TAS2R) before releasing calcium to hyperpolarize airway smooth muscle membranes, inducing bronchodilation (Workman AD, Palmer JN, Adappa ND, Cohen NA. Curr Allergy Asthma Rep 15: 72, 2015; Devillier P, Naline E, Grassin-Delyle S. Pharmacol Ther 155: 11-21, 2015). Theophylline is conventionally used to treat asthma, whereas, according to the literature, the dosage required for orally administered caffeine has yielded modest improvement (Alfaro TM, Monteiro RA, Cunha RA, Cordeiro CR. Clin Respir J 12: 1283-1294, 2018). We sought to determine whether aerosolization of ultrafine caffeine particles (2.5-4 μm) directly to the lungs of susceptible A/J mice challenged with methacholine would improve pulmonary function via forced oscillation technique. In addition, we assessed whether nebulization of caffeine leads to changes in lung pathophysiology and bronchoalveolar lavage cell profiles. We found that mice that received aerosolized caffeine had statistically significant decreases in maximum airway resistance [6.3 vs. 3.9 cmH2O·s/mL at 62.5 mg/mL caffeine; confidence interval (CI) = -4.3, -0.4; P = 0.02] and significant delays in the time required to reach maximum resistance compared with that of controls (64.7 vs. 172.1 sec at 62.5 mg/mL caffeine, CI = 96.0, 118.9; P < 0.0001). Nebulized caffeine yielded a consistent effect on airway hyperresponsiveness at a range of doses without evidence of significant pathology relative to vehicle control.NEW & NOTEWORTHY For decades, coffee has been shown to improve symptoms in patients with asthma. One component, theophylline, is conventionally used to treat asthma, whereas the dosage required for orally administered caffeine has yielded modest improvement. We sought to determine whether aerosolization of caffeine directly to the lungs of susceptible A/J mice challenged with methacholine would alter pulmonary function via forced oscillation technique. We found nebulized caffeine yielded a consistent improvement on murine AHR.

Physiological Effects of Condensed Tannins from Black Currant (Ribes nigrum L.) on Isolated Rat Duodenal Contraction

Condensed tannins (CTs) extracted from various plants have been shown to possess antioxidant, antidiabetic, anthelmintic, anti-palatable and anti-diarrhea activity. Black currant (Ribes nigrum L.), a native plant of northern Europe and Asia, is rich in phenolic compounds, including CTs. Among the biological activities of CTs, their astringent property is likely to affect gastrointestinal motility. This study aimed to investigate the physiological effect of CTs from black currant (R. nigrum L.) leaves on isolated rat duodenal contraction. Duodenal segments were fixed in organ baths containing carbogen aerated Krebs solution at the resting tension of 0.7 - 0.8 g. The frequency, amplitude, and tone of duodenal contraction were recorded. Either CTs or acetylcholine (ACh) were cumulatively added into the bath at the concentration of between 0.001 - 10 µg/mL and 10–8 - 10–4 M, respectively. The mechanisms of CTs and ACh actions were studied using muscarinic receptor antagonist (atropine, 1.55×10–5 M) and calcium channel blocker (verapamil, 10–6 M). It is found that CTs at the concentration between 0.001 - 10 µg/mL had no direct effect on duodenal frequency, amplitude, and tone of contraction, whereas ACh showed a significant increase in tonic contraction, was suppressed by atropine. Interestingly, in the presence of atropine and verapamil, CTs showed a further significant decrease in the amplitude of duodenal contraction compared to the effect of these 2 blockers alone. It is concluded that CTs would synergize the activity of the muscarinic receptor antagonist and the calcium channel blocker at duodenal enteric neurons or smooth muscle membrane. However, the use of CTs from black currant (R. nigrum L.) leaves to treat gastro-intestinal disorders while having muscarinic receptor antagonist or calcium channel blocker need cautions.&#x0D; HIGHLIGHTS&#x0D; &#x0D; Black currant (Ribes nigrum L.), a native plant of northern Europe and Asia, is rich in phenolic compounds including condensed tannins (CTs) and their astringent property is likely to affect gastrointestinal motility&#x0D; CTs isolated from leaves of black currant (R. nigrum L.) at the physiological doses (0.001 - 10.0 µg/mL) have no direct effect on isolated rat duodenal contraction&#x0D; CTs may possess the synergistic or additive effect with either atropine, a nonselective muscarinic receptor, or verapamil, a calcium channel blocker on duodenal amplitude of contraction&#x0D; &#x0D; GRAPHICAL ABSTRACT

Morphofunctional features of the uterus of rodents: morphogenesis, structure, cyclic changes, impact of various negative environmental factors

The review highlights and generalises data on morphological and functional features of the rodent uterus. In all viviparous vertebrates, the development of embryos occurs in a specific organ – the uterus. The mammalian uterus is a primary paired organ that develops from the oviducts. In mammals with more complicated level of organisation (in all placentals), there is a tendency for the fusion of two uteri (partial, for example, in most rodents). To date, the morphofunctional features of the uterus of laboratory animals have been sufficiently studied in representatives of the order Rodentia. The data on the structural features of the uterus of most rodents from natural ecosystems are stated to be fragmentary and refer mainly to synanthropic species. In all rodents, the uterine wall is structured typically of mammals and consists of three membranes – mucous (endometrium), muscular (myometrium) and serous (perimetry). The mucous membrane of the uterus is lined with a single layer of ciliated epithelium. Cyclic changes in the mucous membrane of the uterus and vagina of rodents results in a complex of rearrangements. Thus, the integumentary epithelium of the uterine mucosa is represented by high cylindrical cells during periods of physiological activity, the cell nuclei are arranged in two to three rows, and the height of the epithelium decreases during periods of physiological rest. The leading role in the functional regulation of the uterus belongs to female sex hormones; their secretion, in turn, depends on the pituitary and hypothalamic hormones. The paper provides data on the negative impact of various organic substances, metal compounds, physical factors, including electromagnetic effects, on the structural components of the uterine wall (integumentary and glandular epithelium, smooth muscle membrane myocytes, cellular elements of connective tissue).

EndoFLIP Topography: Motor Patterns in an Obstructed Esophagus

EndoFLIP Topography: Motor Patterns in an Obstructed Esophagus

Diminished Pulmonary Arterial Smooth Muscle Cell Membrane Cholesterol Unmasks EGFR‐dependent Vasoconstriction

IntroductionChronic hypoxia (CH) leads to increased pulmonary vascular resistance and resultant pulmonary hypertension (PH) in patients with chronic obstructive pulmonary disease, sleep apnea, or in residents at high altitude. Recent studies from our laboratory have demonstrated that CH augments vasoconstrictor reactivity to a variety of stimuli, including pulmonary arterial smooth muscle cell (PASMC) stretch, membrane depolarization, and stimulation of G protein‐coupled receptors. This effect of CH is mediated by a switch in PASMC signaling from Ca2+‐dependent mechanisms towards a novel epidermal growth factor receptor (EGFR)‐NADPH oxidase 2 (NOX2)‐Rho kinase (ROK) Ca2+ sensitization pathway. The overall objective of this study is to understand the mechanism by which CH mediates this switch in signaling. We have recently demonstrated that CH decreases membrane cholesterol, a component of lipid membrane domains that can regulate both EGFR and NOX2 activity in other cell types. Therefore, we hypothesized that this EGFR‐NOX2‐ROK signaling pathway is intact in PASMC from control animals but quiescent due to inhibition by membrane cholesterol.MethodsWe first validated our ability to manipulate membrane cholesterol levels by treating primary cultures of PASMCs from Sprague Dawley rats with either cholesterol (2 mM), epicholesterol (an epimeric form of cholesterol; 2 mM), or MβCD (cholesterol chelator; 10 mM). Membrane cholesterol content was quantified by filipin (50 mg/ml) fluorescence using confocal microscopy. Subsequently, we examined vasoconstrictor responses to increasing concentrations of epidermal growth factor (EGF) (10‐10 – 10‐6 M) in pressurized, endothelium‐disrupted rat pulmonary arteries (~150 mm diameter). The effect of diminished membrane cholesterol on vasomotor responses was tested by substituting native cholesterol with epicholesterol (2 mM), which lacks the ability participate in protein‐sterol interactions while maintaining normal membrane fluidity. Vessels were additionally treated with the selective NOX2 inhibitor gp91ds‐tat (50 mM), the selective Rho kinase inhibitor fasudil (10 mM), or their respective vehicles.ResultsCholesterol supplementation significantly increased PASMC membrane cholesterol as indicated by filipin fluorescence. Conversely, treatment with either MβCD or epicholesterol significantly decreased membrane cholesterol compared to vehicle‐treated cells (Figure 1). In agreement with our hypothesis, epicholesterol‐dependent substitution of cholesterol revealed concentration‐dependent vasoconstriction to EGF in pulmonary arteries (Figure 2A). Treatment with either fasudil (Figure 2A) or gp91ds‐tat (Figure 2B) attenuated this effect.ConclusionsThe EGFR‐NOX2‐RhoA signaling cascade is present in arteries from normoxic animals but is normally inhibited by membrane cholesterol. The results of this study will allow for improved mechanistic analysis of how cholesterol regulates this pathway that is central to enhanced vasoconstrictor reactivity in PH.

Effects on Endothelin‐1 on Irisin Production By Myocytes Derived From C2C12 Cells

Skeletal muscle is one of the most metabolically active tissues. Consequently, it is highly susceptible to hormonal variations, whether by endocrine, paracrine, or autocrine pathways. This muscle is recognized as an endocrine organ that releases myokines, including irisin. Irisin acts as a signal that induces energy consumption, communicating directly with the adipose tissue, modulating the metabolic profile and increasing energy expenditure. Endothelin‐1 (ET‐1), a peptide with pro‐inflammatory activity, is a potent vasoconstrictor that interacts with receptors on the vascular smooth muscle membrane, stimulating its contraction. However, the functional relationship between ET‐1 and skeletal muscle remains unclear. The objective of this work was to study the effects of ET‐1 on cell count, cell viability, and irisin production in C2C12 cell culture. The cells were cultured for 3 and 5 days, at an initial density (day 0) of 1 x 103 cells/well (48‐well culture plates), in 200 µL of culture medium containing equine serum, antibiotics, and antifungal, in the absence of ET‐1and in the presence of ET‐1 at two concentrations (10‐10 M and 10‐6 M). The assays were performed in triplicates, with exchange of the medium every 48 h. Cell counting was performed with a Bio‐Rad TC20™ cell counter. The MTT reduction assay was used to determine cell viability. Irisin in the culture medium was determined using an ELISA kit. The results were presented as mean ± SEM. Statistical analysis of the data employed ANOVA and Fisher’s test for multiple comparisons, adopting significance level of 5% (p&lt;0.05). Both time‐ and dose‐dependent effects on cell counts were observed. Culturing for 5 days led to a higher number of cells (111,367 ± 42,813 cells), compared to 3‐day culture (22,422 ± 9,141 cells). Furthermore, stimulation by ET‐1 at 10‐6 M resulted in higher numbers of cells for culture times of both 3 days (64,971 ± 41,841 cells) and 5 days (174,478 ± 49,152 cells), compared to the same culture times in the absence of ET‐1. Cell viability was higher for the 5‐day cultures of the control group (0.1984 ± 0.04) and the 10‐10 M ET‐1 group (0.2633 ± 0.0311), compared to the 3‐day cultures (control group: 0.1681 ± 0.003; 10‐10 M ET‐1 group: 0.1421 ± 0.0135). Biologically, in the absence of ET‐1, there was a time‐dependent reduction in irisin production (3‐day culture: 0.106 ± 0.0085 ng/mL; 5‐day culture: 0.097 ± 0.09 ng/mL). Therefore, the findings suggested that the multiplication activity of the C2C12 cells cultivated according to the proposed study model was modulated by time and by ET‐1, and that cell viability was modulated by the culture time. The time‐dependent decreased in irisin production suggested that there were negative effects on cell multiplication and viability, and that irisin could modulate the intracellular activity pathways for the mechanisms of cell differentiation.

Genetic ablation of smooth muscle KIR2.1 is inconsequential to the function of mouse cerebral arteries

Cerebral blood flow is a finely tuned process dependent on coordinated changes in arterial tone. These changes are strongly tied to smooth muscle membrane potential and inwardly rectifying K+ (KIR) channels are thought to be a key determinant. To elucidate the role of KIR2.1 in cerebral arterial tone development, this study examined the electrical and functional properties of cells, vessels and living tissue from tamoxifen-induced smooth muscle cell (SMC)-specific KIR2.1 knockout mice. Patch-clamp electrophysiology revealed a robust Ba2+-sensitive inwardly rectifying K+ current in cerebral arterial myocytes irrespective of KIR2.1 knockout. Immunolabeling clarified that KIR2.1 expression was low in SMCs while KIR2.2 labeling was remarkably abundant at the membrane. In alignment with these observations, pressure myography revealed that the myogenic response and K+-induced dilation were intact in cerebral arteries post knockout. At the whole organ level, this translated to a maintenance of brain perfusion in SMC KIR2.1−/− mice, as assessed with arterial spin-labeling MRI. We confirmed these findings in superior epigastric arteries and implicated KIR2.2 as more functionally relevant in SMCs. Together, these results suggest that subunits other than KIR2.1 play a significant role in setting native current in SMCs and driving arterial tone.

Zinc drives vasorelaxation by acting in sensory nerves, endothelium and smooth muscle

Zinc, an abundant transition metal, serves as a signalling molecule in several biological systems. Zinc transporters are genetically associated with cardiovascular diseases but the function of zinc in vascular tone regulation is unknown. We found that elevating cytoplasmic zinc using ionophores relaxed rat and human isolated blood vessels and caused hyperpolarization of smooth muscle membrane. Furthermore, zinc ionophores lowered blood pressure in anaesthetized rats and increased blood flow without affecting heart rate. Conversely, intracellular zinc chelation induced contraction of selected vessels from rats and humans and depolarized vascular smooth muscle membrane potential. We demonstrate three mechanisms for zinc-induced vasorelaxation: (1) activation of transient receptor potential ankyrin 1 to increase calcitonin gene-related peptide signalling from perivascular sensory nerves; (2) enhancement of cyclooxygenase-sensitive vasodilatory prostanoid signalling in the endothelium; and (3) inhibition of voltage-gated calcium channels in the smooth muscle. These data introduce zinc as a new target for vascular therapeutics.

Remodeling of Arterial Tone Regulation in Postnatal Development: Focus on Smooth Muscle Cell Potassium Channels.

Maturation of the cardiovascular system is associated with crucial structural and functional remodeling. Thickening of the arterial wall, maturation of the sympathetic innervation, and switching of the mechanisms of arterial contraction from calcium-independent to calcium-dependent occur during postnatal development. All these processes promote an almost doubling of blood pressure from the moment of birth to reaching adulthood. This review focuses on the developmental alterations of potassium channels functioning as key smooth muscle membrane potential determinants and, consequently, vascular tone regulators. We present evidence that the pattern of potassium channel contribution to vascular control changes from Kir2, Kv1, Kv7 and TASK-1 channels to BKCa channels with maturation. The differences in the contribution of potassium channels to vasomotor tone at different stages of postnatal life should be considered in treatment strategies of cardiovascular diseases associated with potassium channel malfunction.

Regulation of capillary hemodynamics by KATP channels in resting skeletal muscle.

ATP‐sensitive K+ channels (KATP) have been implicated in the regulation of resting vascular smooth muscle membrane potential and tone. However, whether KATP channels modulate skeletal muscle microvascular hemodynamics at the capillary level (the primary site for blood‐myocyte O2 exchange) remains unknown. We tested the hypothesis that KATP channel inhibition would reduce the proportion of capillaries supporting continuous red blood cell (RBC) flow and impair RBC hemodynamics and distribution in perfused capillaries within resting skeletal muscle. RBC flux (f RBC), velocity (V RBC), and capillary tube hematocrit (Hctcap) were assessed via intravital microscopy of the rat spinotrapezius muscle (n = 6) under control (CON) and glibenclamide (GLI; KATP channel antagonist; 10 µM) superfusion conditions. There were no differences in mean arterial pressure (CON:120 ± 5, GLI:124 ± 5 mmHg; p > 0.05) or heart rate (CON:322 ± 32, GLI:337 ± 33 beats/min; p > 0.05) between conditions. The %RBC‐flowing capillaries were not altered between conditions (CON:87 ± 2, GLI:85 ± 1%; p > 0.05). In RBC‐perfused capillaries, GLI reduced f RBC (CON:20.1 ± 1.8, GLI:14.6 ± 1.3 cells/s; p < 0.05) and V RBC (CON:240 ± 17, GLI:182 ± 17 µm/s; p < 0.05) but not Hctcap (CON:0.26 ± 0.01, GLI:0.26 ± 0.01; p > 0.05). The absence of GLI effects on the %RBC‐flowing capillaries and Hctcap indicates preserved muscle O2 diffusing capacity (DO2m). In contrast, GLI lowered both f RBC and V RBC thus impairing perfusive microvascular O2 transport (Q̇m) and lengthening RBC capillary transit times, respectively. Given the interdependence between diffusive and perfusive O2 conductances (i.e., %O2 extraction∝DO2m/Q̇m), such GLI alterations are expected to elevate muscle %O2 extraction to sustain a given metabolic rate. These results support that KATP channels regulate capillary hemodynamics and, therefore, microvascular gas exchange in resting skeletal muscle.

Evaluating the use of magnesium sulphate intravenous for the Asthma in pediatric patients

Background: One of the proposed mechanisms for the action of the magnesium sulphate includes relaxation of the bronchial smooth muscle by the inhibition of the calcium across the smooth muscle membranes. The present study was conducted to investigate the safety and efficacy of magnesium sulphate administration for the severity of acute asthma in pediatric patients. Methodology: The present retrospective study was conducted at Department of Tuberculosis and Reparatory Medicine, GMERS Medical College, Gandhinagar, Gujarat. Inclusion criteria included the patients under the age of 18 years or younger and who were treated with intravenous magnesium sulphate for the treatment of asthma attacks in the department of emergency of the hospital. Data collection form was used to assess the details of the patients. Results: Total of 60 paediatric patients were included in the present study. All the paediatric patients did received the magnesium sulphate through intravenous administration. Total of 40 patients required a PICU admission after the magnesium sulphate treatment. It was found that total of 96% of the patient did required the hospital admission. Around 5 patients were discharged directly from the emergency department. The serum concentrations of the magnesium sulphate were slight above the normal range that is 1.5 to 2.5 mg/dl. Conclusions: Monitoring of the intravenous magnesium sulphate and the related practised in various medical institutions did vary. Further studies are required to determine the regimen of the dose of intravenous magnesium sulphate for the paediatric patients. Keywords: Asthma; Intravenous; Magnesium sulphate; Paediatric patients.

Experimental study on the repair of ureteral functional regeneration with highly bioactive extracellular matrix stent.

The research of extracellular matrix stent (ECM) has made some progress in the repair of urethra and bladder defects. To observe the effects of highly bioactive ECM scaffold on the regeneration and repair of defects in long-segment ureteral replacement. An animal model of long-segment ureteral defect was established and four-layer tubular highly bioactive ECM materials were prepared. After the ureteral defect was repaired through surgery, the rabbits in the negative control group were administered a non-bioactive stent, and rabbits in the observation group were treated with an ECM stent. Comparison of macro-indicators: The negative control group had a higher infection rate, a lower survival rate and more complications than the observation group (p < 0.05). The frequency of ureteral peristalsis in the negative control group was lower than in the observation group. In addition, the rate of urinary dysfunction was higher, and the ratio of ureteral diameter was lower in the negative control group than in the observation group (all p < 0.05). Comparison of histopathology: Three months after the operation, the vascular, smooth muscle and mucous membrane of the ureter in the observation group regenerated to close to normal ureteral tissue. There was no significant difference between the ureter regeneration in the repair area and the normal ureter tissue in the observation group 3 months after the operation. The number of regenerated muscle fibers in the observation group was significantly higher than that of the negative control group. Compared with the negative control group, the fibrous capsule was thicker, the percentages of CD31, CD3, CD68, CD80+, and CD163+ were higher, the scope of new smooth muscle fiber was expanded, fusion with the host muscle fibers was higher, and the neuromuscular junction (NMJ) structure was stronger in the observation group (all p < 0.05). A highly bioactive ECM stent can better regenerate the local anatomical structure and physiological function.

Hydrogen Sulfide Relaxes Human Uterine Artery via Activating Smooth Muscle BKCa Channels.

Opening of large conductance calcium-activated and voltage-dependent potassium (BKCa) channels hyperpolarizes plasma membranes of smooth muscle (SM) to cause vasodilation, underling a key mechanism for mediating uterine artery (UA) dilation in pregnancy. Hydrogen sulfide (H2S) has been recently identified as a new UA vasodilator, yet the mechanism underlying H2S-induced UA dilation is unknown. Here, we tested whether H2S activated BKCa channels in human UA smooth muscle cells (hUASMC) to mediate UA relaxation. Multiple BKCa subunits were found in human UA in vitro and hUASMC in vitro, and high β1 and γ1 proteins were localized in SM cells in human UA. Baseline outward currents, recorded by whole-cell and single-channel patch clamps, were significantly inhibited by specific BKCa blockers iberiotoxin (IBTX) or tetraethylammonium, showing specific BKCa activity in hUASMC. H2S dose (NaHS, 1–1000 µM)-dependently potentiated BKCa currents and open probability. Co-incubation with a Ca2+ blocker nifedipine (5 µM) or a chelator (ethylene glycol-bis (β-aminoethyl ether)-N,N,N′,N′-tetraacetic acid (EGTA), 5 mM) did not alter H2S-potentiated BKCa currents and open probability. NaHS also dose-dependently relaxed phenylephrine pre-constricted freshly prepared human UA rings, which was inhibited by IBTX. Thus, H2S stimulated human UA relaxation at least partially via activating SM BKCa channels independent of extracellular Ca2+.

Galectin-7 overexpression destroys airway epithelial barrier in transgenic mice.

When the integrity of airway epithelium is destroyed, the ordered airway barrier no longer exists and increases sensitivity to viral infections and allergens, leading to the occurrence of airway inflammation such as asthma. Here, we found that galectin-7 transgenic(+) mice exhibited abnormal airway structures as embryos and after birth. These abnormalities included absent or substantially reduced pseudostratified columnar ciliated epithelium and increased monolayer cells with irregular arrangement and widening of intercellular spaces. Moreover, airway tissue from galectin-7 transgenic(+) mice showed evidence of impaired cell-cell junctions and decreased expression of zonula occludens-1(ZO-1) and E-cadherin. When treated with respiratory syncytial virus (RSV) or ovalbumin (OVA), galectin-7 transgenic(+) mice developed substantially increased bronchial epithelial detachment and apoptosis, airway smooth muscle and basement membrane thickening, and enhanced airway responsiveness. We found that Galectin-7 localized in the cytoplasm and nucleus of bronchial epithelial cells, and that increased apoptosis was mediated through mitochondrial release of cytochrome c and upregulated JNK1 activation and expression of caspase-3 in galectin-7 Tg(+) mice. These findings suggested that Galectin-7 causes airway structural defects and destroys airway epithelium barrier, which predispose the airways to RSV or OVA-induced epithelial apoptosis, injury, and other asthma responses.

Endothelial Nitric Oxide Suppresses Action-Potential-Like Transient Spikes and Vasospasm in Small Resistance Arteries.

Endothelial dysfunction in small arteries is a ubiquitous, early feature of cardiovascular disease, including hypertension. Dysfunction reflects reduced bioavailability of endothelium-derived nitric oxide (NO) and depressed endothelium-dependent hyperpolarization that enhances vasoreactivity. We measured smooth muscle membrane potential and tension, smooth muscle calcium, and used real-time quantitative polymerase chain reaction in small arteries and isolated tubes of endothelium to investigate how dysfunction enhances vasoreactivity. Rat nonmyogenic mesenteric resistance arteries developed vasomotion to micromolar phenylephrine (α1-adrenoceptor agonist); symmetrical vasoconstrictor oscillations mediated by L-type voltage-gated Ca2+ channels (VGCCs). Inhibiting NO synthesis abolished vasomotion so nanomolar phenylephrine now stimulated rapid, transient depolarizing spikes in the smooth muscle associated with chaotic vasomotion/vasospasm. Endothelium-dependent hyperpolarization block also enabled phenylephrine-vasospasm but without spikes or chaotic vasomotion. Depolarizing spikes were Ca2+-based and abolished by either T-type or L-type VGCCs blockers with depressed vasoconstriction. Removing NO also enabled transient spikes/vasoconstriction to Bay K-8644 (L-type VGCC activator). However, these were abolished by the L-type VGCC blocker nifedipine but not T-type VGCC block. Phenylephrine also initiated T-type VGCC-transient spikes and enhanced vasoconstriction after NO loss in nonmyogenic arteries from spontaneously hypertensive rats. In contrast to mesenteric arteries, myogenic coronary arteries displayed transient spikes and further vasoconstriction spontaneously on loss of NO. T-type VGCC block abolished these spikes and additional vasoconstriction but not myogenic tone. Therefore, in myogenic and nonmyogenic small arteries, reduced NO bioavailability engages T-type VGCCs, triggering transient depolarizing spikes in normally quiescent vascular smooth muscle to cause vasospasm. T-type block may offer a means to suppress vasospasm without inhibiting myogenic tone mediated by L-type VGCCs.

Altered Lipid Domains Facilitate Enhanced Pulmonary Vasoconstriction after Chronic Hypoxia.

Chronic hypoxia (CH) augments depolarization-induced pulmonary vasoconstriction through superoxide-dependent, Rho kinase-mediated Ca2+ sensitization. Nicotinamide adenine dinucleotide phosphate oxidase and EGFR (epidermal growth factor receptor) signaling contributes to this response. Caveolin-1 regulates the activity of a variety of proteins, including EGFR and nicotinamide adenine dinucleotide phosphate oxidase, and membrane cholesterol is an important regulator of caveolin-1 protein interactions. We hypothesized that derangement of these membrane lipid domain components augments depolarization-induced Ca2+ sensitization and resultant vasoconstriction after CH. Although exposure of rats to CH (4 wk, ∼380 mm Hg) did not alter caveolin-1 expression in intrapulmonary arteries or the incidence of caveolae in arterial smooth muscle, CH markedly reduced smooth muscle membrane cholesterol content as assessed by filipin fluorescence. Effects of CH on vasoreactivity and superoxide generation were examined using pressurized, Ca2+-permeabilized, endothelium-disrupted pulmonary arteries (∼150 μm inner diameter) from CH and control rats. Depolarizing concentrations of KCl evoked greater constriction in arteries from CH rats than in those obtained from control rats, and increased superoxide production as assessed by dihydroethidium fluorescence only in arteries from CH rats. Both cholesterol supplementation and the caveolin-1 scaffolding domain peptide antennapedia-Cav prevented these effects of CH, with each treatment restoring membrane cholesterol in CH arteries to control levels. Enhanced EGF-dependent vasoconstriction after CH similarly required reduced membrane cholesterol. However, these responses to CH were not associated with changes in EGFR expression or activity, suggesting that cholesterol regulates this signaling pathway downstream of EGFR. We conclude that alterations in membrane lipid domain signaling resulting from reduced cholesterol content facilitate enhanced depolarization- and EGF-induced pulmonary vasoconstriction after CH.

Diversity of neurogenic smooth muscle electrical rhythmicity in mouse proximal colon.

The mechanisms underlying electrical rhythmicity in smooth muscle of the proximal colon are incompletely understood. Our aim was to identify patterns of electrical rhythmicity in smooth muscle of the proximal region of isolated whole mouse colon and characterize their mechanisms of origin. Two independent extracellular recording electrodes were used to record the patterns of electrical activity in smooth muscle of the proximal region of whole isolated mouse colon. Cross-correlation analysis was used to quantify spatial coordination of these electrical activities over increasing electrode separation distances. Four distinct neurogenic patterns of electrical rhythmicity were identified in smooth muscle of the proximal colon, three of which have not been identified and consisted of bursts of rhythmic action potentials at 1-2 Hz that were abolished by hexamethonium. These neurogenic patterns of electrical rhythmicity in smooth muscle were spatially and temporally synchronized over large separation distances (≥2 mm rosto-caudal axis). Myogenic slow waves could be recorded from the same preparations, but they showed poor spatial and temporal coordination over even short distances (≤1 mm rostro-caudal axis). It is not commonly thought that electrical rhythmicity in gastrointestinal smooth muscle is dependent upon the enteric nervous system. Here, we identified neurogenic patterns of electrical rhythmicity in smooth muscle of the proximal region of isolated mouse colon, which are dependent on synaptic transmission in the enteric nervous system. If the whole colon is studied in vitro, recordings can preserve novel neurogenic patterns of electrical rhythmicity in smooth muscle.NEW & NOTEWORTHY Previously, it has not often been thought that electrical rhythmicity in smooth muscle of the gastrointestinal tract is dependent upon the enteric nervous system. We identified patterns of electrical rhythmicity in smooth muscle of the mouse proximal colon that were abolished by hexamethonium and involved the temporal synchronization of smooth muscle membrane potential over large spatial fields. We reveal different patterns of electrical rhythmicity in colonic smooth muscle that are dependent on the ENS.