High dilutions of homeopathic remedies induce relaxation of rat aorta precontracted with Noradrenalin

BACKGROUND Homeopathic potencies have been reported to produce alteration of contraction in isolated rat ileum in an organ bath. Potentized homeopathic drugs like Lycopus V and Aurum met are used for the treatment of hypertension. AIM The purpose of this study is to see whether Lycopus V 30 CH and Aurum met 30 CH could produce relaxation of isolated rat aorta in the organ bath. METHODS The aorta of rats were dissected out, placed in Krebs-Henseleit solution, cleared of connective tissue and endothelium and cut into 2-2.5 mm long rings. The rings were fixed in organ baths with the upper end connected by a string to an isometric transducer which was finally attached through a data acquisitation equipment to a computer. Aurum met 30 CH Lycopus V 30 CH, and their medium 90% ethanol were added separately to the bathing fluid containing the aorta rings which were precontracted with noradrenalin (NA). RESULTS Both the drugs produced significant relaxation of the aorta (p

The Effect of 5-hydroxytryptamine on Smooth Muscles is Impacted by Broadband UV and LED UV and Blue Light

Due to their effects, similar to low-intensity therapy light sources such as light-emitting diodes (LED) and broadband spectrum lamps have recently become commonly used in the diagnosis and treatment of neurodegenerative pathologies, cancer, as well as ageing. Despite the proven positive effects of such therapies, deeper understanding of the light therapies’ biological effects remains unclear. Even more, the molecular mechanisms through which different neurotransmitters, namely serotonin (5-hydroxytryptamine, 5-HT), mediate the organism’s response to radiation are yet indistinct. In this paper, we present the design and development of a specialized system for irradiation of biological objects, which is composed of LED 365 nm and LED 470 nm and a broadband lamp source of UVA/B (350 nm) with intensity, power density and direction, which can be optimized experimentally. The system, named a “water organ bath (wob)”, is used in the current work to irradiate smooth muscle stomach strips of rats. The obtained results prove that the modulation of the spontaneous contractile smooth muscle activity and the potentiation of the effects of major neurotransmitters are executed by the emitted light. The probable explanation for the neurotransmitters photoactivation is that it is the resultant effect of electromagnetic radiation on intracellular enzymes signaling systems.

Irisin relaxes rat thoracic aorta: MEK1/2 signaling pathway, KV channels, SKCa channels, and BKCa channels are involved in irisin-induced vasodilation

In this study, it was aimed to investigate the effects of irisin on vascular smooth muscle contractility in rat thoracic aorta, and the hypothesis that mitogen-activated protein kinase kinase (MEK1/2) signalling pathway, voltage-gated potassium (KV) channels, small-conductance calcium-activated potassium (SKCa) channels, and large-conductance calcium-activated potassium (BKCa) channels may have roles in these effects. Isometric contraction-relaxation responses of isolated thoracic aorta rings were measured with an organ bath model. The steady contraction was induced with 10-5 M phenylephrine (PHE), and then the concentration-dependent responses of irisin (10-9-10-6 M) were examined. Irisin exerted the vasorelaxant effects at concentrations of 10-8, 10-7, and 10-6 M compared to the control group (p<0.001). Besides, MEK1/2 inhibitor U0126, KV channel blocker XE-991, SKCa channel blocker apamin, and BKCa channel blocker tetraethylammonium (TEA) incubations significantly inhibited the irisin-induced relaxation responses. In conclusion, the first physiological findings were obtained regarding the functional relaxing effects of irisin in rat thoracic aorta. The findings demonstrated that irisin induces relaxation responses in endothelium-intact aortic rings in a concentration-dependent manner. Furthermore, this study is the first to report that irisin-induced relaxation responses are related to the activity of the MEK1/2 pathway, KV channels, and calcium-activated K+ (SKCa and BKCa) channels.

Endothelial Contribution to Warfarin-Induced Arterial Media Calcification in Mice

Arterial media calcification (AMC) is predominantly regulated by vascular smooth muscle cells (VSMCs), which transdifferentiate into pro-calcifying cells. In contrast, there is little evidence for endothelial cells playing a role in the disease. The current study investigates cellular functioning and molecular pathways underlying AMC, respectively by, an ex vivo isometric organ bath set-up to explore the interaction between VSMCs and ECs and quantitative proteomics followed by functional pathway interpretation. AMC development, which was induced in mice by dietary warfarin administration, was proved by positive Von Kossa staining and a significantly increased calcium content in the aorta compared to that of control mice. The ex vivo organ bath set-up showed calcified aortic segments to be significantly more sensitive to phenylephrine induced contraction, compared to control segments. This, together with the fact that calcified segments as compared to control segments, showed a significantly smaller contraction in the absence of extracellular calcium, argues for a reduced basal NO production in the calcified segments. Moreover, proteomic data revealed a reduced eNOS activation to be part of the vascular calcification process. In summary, this study identifies a poor endothelial function, next to classic pro-calcifying stimuli, as a possible initiator of arterial calcification.

Graft Preservation Solution DuraGraft® Alleviates Vascular Dysfunction Following In Vitro Ischemia/Reperfusion Injury in Rats

Vascular ischemia/reperfusion injury (IRI) in patients undergoing coronary artery bypass grafting can result in graft failure and the need for repeat revascularization procedures. DuraGraft® has been shown to protect structure and function in saphenous vein grafts against IRI. We compared the effect of DuraGraft® to saline solution on arterial grafts submitted to IRI. Rat thoracic aortic rings were harvested and immediately mounted in organ bath chambers (control, n = 7 rats) or underwent cold ischemic preservation either in saline (IR, n = 9 rats) or DuraGraft® (IR+Dura, n = 9 rats). Vascular function was measured ex vivo and immunohistochemistry was performed. Impaired maximum vasorelaxation (Rmax) to ACh in the IR-group compared to controls was ameliorated by DuraGraft®, indicating an improvement in endothelial function (Rmax to ACh (%): IR + Dura 73 ± 2 vs. IR 48 ± 3, p < 0.05). Additionally, decreased aortic ring sensitivity to ACh (pD2-value: -log 50% maximum response) seen after IR in the saline group was increased by DuraGraft® (pD2 to ACh: IR+Dura 7.1 ± 0.1 vs. IR 6.3 ± 0.2, p < 0.05). Impaired maximum contractile response to phenylephrine and high potassium chloride concentrations in the IR group compared to controls was significantly improved by DuraGraft®. DuraGraft® alleviates vascular dysfunction following IRI by reducing nitro-oxidative stress and the expression of ICAM-1, without leukocytes engagement.

Complementary mechanisms of modulation of spontaneous phasic contractions by the gaseous signalling molecules NO, H2S, HNO and the polysulfide Na2S3 in the rat colon

Objectives Reactive oxygen and nitrogen species may be produced during inflammation leading to the formation of NO, H2S or HNO. Enzymes such as iNOS, CSE and CBS might also be responsible for polysulfide production. Since these signalling molecules might have an impact on colonic motility, the aim of this study was to compare their effect on rat colonic slow phasic contractions (SPC). Methods Organ bath measurements with strips obtained from rat proximal colon were performed using the polysulfide Na2S3, sodium nitroprusside (NaNP), sodium hydrogen sulfide (NaHS), Angeli’s salt as NO, H2S, and HNO donors, respectively. TTX (1 µM) was used to block neuronal activity. Results All four molecules, concentration-dependently, inhibited the amplitude and frequency of SPC both in the circular and longitudinal muscle layer. The relative potency was NaNP>Angeli’s salt>NaHS>Na2S3. The inhibitory response induced by NaNP (1 µM) and Angeli’s salt (50 µM) was reversed by ODQ (10 µM) whereas the inhibitory effect of NaHS (1 mM) was reversed by apamin (1 µM) and glibenclamide (10 µM). Na2S3 (1 mM) response was partially reversed by apamin (1 µM) and glibenclamide (10 µM). High concentrations of Na2S3 caused an increase in tone. Low concentrations of NaHS or Na2S3 did not potentiate NaNP responses. Conclusions All signalling molecules inhibit SPC in both muscle layers. The effect is independent of neural activity and involves guanylyl cyclase (NO and HNO) and SKCa and KATP channels (NaHS or Na2S3). Other pathways might also be involved in Na2S3 responses. Accordingly, complementary mechanisms of inhibition might be attributable to these signalling molecules.

Spasmolytic Effects of Aphanizomenon Flos Aquae (AFA) Extract on the Human Colon Contractility

The blue-green algae Aphanizomenon flos aquae (AFA), rich in beneficial nutrients, exerts various beneficial effects, acting in different organs including the gut. Klamin® is an AFA extract particularly rich in β-PEA, a trace-amine considered a neuromodulator in the central nervous system. To date, it is not clear if β-PEA exerts a role in the enteric nervous system. The aims of the present study were to investigate the effects induced by Klamin® on the human distal colon mechanical activity, to analyze the mechanism of action, and to verify a β-PEA involvement. The organ bath technique, RT-PCR, and immunohistochemistry (IHC) were used. Klamin® reduced, in a concentration-dependent manner, the amplitude of the spontaneous contractions. EPPTB, a trace-amine receptor (TAAR1) antagonist, significantly antagonized the inhibitory effects of both Klamin® and exogenous β-PEA, suggesting a trace-amine involvement in the Klamin® effects. Accordingly, AphaMax®, an AFA extract containing lesser amount of β-PEA, failed to modify colon contractility. Moreover, the Klamin® effects were abolished by tetrodotoxin, a neural blocker, but not by L-NAME, a nitric oxide-synthase inhibitor. On the contrary methysergide, a serotonin receptor antagonist, significantly antagonized the Klamin® effects, as well as the contractility reduction induced by 5-HT. The RT-PCR analysis revealed TAAR1 gene expression in the colon and the IHC experiments showed that 5-HT-positive neurons are co-expressed with TAAR1 positive neurons. In conclusion, the results of this study suggest that Klamin® exerts spasmolytic effects in human colon contractility through β-PEA, that, by activating neural TAAR1, induce serotonin release from serotoninergic neurons of the myenteric plexus.

Therapeutic Effects of Berberine Hydrochloride on Stress-Induced Diarrhea-Predominant Irritable Bowel Syndrome Rats by Inhibiting Neurotransmission in Colonic Smooth Muscle

The etiology of diarrhea-predominant irritable bowel syndrome (IBS-D) is complicated and closely related to neurotransmission in the gastrointestinal (GI) tract. Developing new strategies for treating this disease is a major challenge for IBS-D research. Berberine hydrochloride (BBH), the derivative of berberine, is a herbal constituent used to treat IBS. Previous studies have shown that BBH has potential anti-inflammatory, antibacterial, analgesic, and antidiarrheal effects and a wide range of biological activities, especially in regulating the release of some neurotransmitters. A modified IBS-D rat model induced by chronic restraint stress was used in all experiments to study the effects of BBH on the GI tract. This study measured the abdominal withdrawal reflex (AWR) response to graded colorectal distention (CRD; 20, 40, 60, and 80 mmHg) and observed the fecal areas of stress-induced IBS-D model. Experiments were conducted using organ bath techniques, which were performed in vitro using strips of colonic longitudinal smooth muscle. Inhibitory and excitatory neurotransmitter agents were added to each organ bath to observe contractile responses on the strips and the treatment effect exerted by BBH. The IBS-D rat model was successfully induced by chronic restraint stress, which resulted in an increased defecation frequency and visceral hypersensitivity similar to that of humans. BBH could reduce 4-h fecal areas and AWR response to CRD in IBS-D. The stress-induced IBS-D model showed upregulated colonic mRNA expression levels of 5-hydroxytryptamine-3A receptor and downregulated expression levels of neuronal nitric oxide synthase. Meanwhile, BBH could reverse this outcome. The responses of substances that regulate the contraction induced by related neurotransmission in the longitudinal smooth muscle of IBS-D colon (including the agonist of acetylcholine, carbachol; NOS inhibitor, L-NAME; and P2Y1 receptor antagonist, MRS2500) can be inhibited by BBH. In summary, BBH promotes defecation frequency and visceral hypersensitivity in IBS-D and exerts inhibitory effects on contractile responses in colonic longitudinal smooth muscle. Thus, BBH may represent a new therapeutic approach for treating IBS-D.

Antispasmodic Effect of Asperidine B, a Pyrrolidine Derivative, through Inhibition of L-Type Ca2+ Channel in Rat Ileal Smooth Muscle

Antispasmodic agents are used for modulating gastrointestinal motility. Several compounds isolated from terrestrial plants have antispasmodic properties. This study aimed to explore the inhibitory effect of the pyrrolidine derivative, asperidine B, isolated from the soil-derived fungus Aspergillus sclerotiorum PSU-RSPG178, on spasmodic activity. Isolated rat ileum was set up in an organ bath. The contractile responses of asperidine B (0.3 to 30 µM) on potassium chloride and acetylcholine-induced contractions were recorded. To investigate its antispasmodic mechanism, CaCl2, acetylcholine, Nω-nitro-l-arginine methyl ester (l-NAME), nifedipine, methylene blue and tetraethylammonium chloride (TEA) were tested in the absence or in the presence of asperidine B. Cumulative concentrations of asperidine B reduced the ileal contraction by ~37%. The calcium chloride and acetylcholine-induced ileal contraction was suppressed by asperidine B. The effects of asperidine B combined with nifedipine, atropine or TEA were similar to those treated with nifedipine, atropine or TEA, respectively. In contrast, in the presence of l-NAME and methylene blue, the antispasmodic effect of asperidine B was unaltered. These results suggest that the antispasmodic property of asperidine B is probably due to the blockage of the L-type Ca2+ channel and is associated with K+ channels and muscarinic receptor, possibly by affecting non-selective cation channels and/or releasing intracellular calcium.

Abstract P171: Comparison Of Modulation Of Renal Arterial Tone By Perivascular Adipose Tissue Between Two Animal Models Of Metabolic Syndrome

Sex differences have recently been noticed in the regulation of arterial tone by perivascular adipose tissue (PVAT). In SHRSP.Z- Lepr fa /IzmDmcr (SHRSP.ZF) rats, an animal model of metabolic syndrome (MetS), we demonstrated that mesenteric and renal PVAT in female rats consistently have an enhancing effect on vasodilation at 23 weeks, an age by which the effect of PVAT is impaired in the male rats. This could explain the sex differences in the prevalence of cardiovascular complications in patients with MetS. Therefore, we determined whether the sex difference in PVAT response also occurs in another animal model of MetS, SHR/NDmcr-cp (SHR-cp) rats.Renal arteries were isolated from male and female 23-week-old SHR-cp rats, and ring preparations with and without PVAT were made. After a stable contraction was obtained by phenylephrine administration, vasodilation in response to acetylcholine was examined using organ bath methods.Vasodilation in arteries without PVAT from female rats was smaller than that in arteries without PVAT from male rats, and presence of PVAT in arteries from female rats increased vasodilation to the same level as that observed in arteries without PVAT from male rats. Furthermore, renal PVAT in male rats was shown to have an enhancing effect on vasodilation.The present study did not identify sex differences in renal PVAT-mediated modulation in SHR-cp rats because the enhancing effects of PVAT did not disappear in male SHR-cp rats, in contrast to that observed in male SHRSP.ZF rats at the same age. The difference in PVAT response in male rats between two MetS models may be due to differences in the severity of MetS symptoms, especially blood pressure, between the models.