Adrenergic Contraction Research Articles

Role of NaV1.7 in postganglionic sympathetic nerve function in human and guinea-pig arteries.

NaV1.7 plays a crucial role in inducing and conducting action potentials in pain-transducing sensory nociceptor fibres, suggesting that NaV1.7 blockers could be effective non-opioid analgesics. While SCN9A is expressed in both sensory and autonomic neurons, its functional role in the autonomic system remains less established. Our single neuron rt-PCR analysis revealed that 82% of sympathetic neurons isolated from guinea-pig stellate ganglia expressed NaV1.7 mRNA, with NaV1.3 being the only other tetrodotoxin-sensitive channel expressed in approximately 50% of neurons. We investigated the role of NaV1.7 in conducting action potentials in postganglionic sympathetic nerves and in the sympathetic adrenergic contractions of blood vessels using selective NaV1.7 inhibitors. Two highly selective NaV1.7 blockers, GNE8493 and PF05089771, significantly inhibited postganglionic compound action potentials by approximately 70% (P<0.01), with residual activity being blocked by the NaV1.3 inhibitor, ICA121431. Electrical field stimulation (EFS) induced rapid contractions in guinea-pig isolated aorta, pulmonary arteries, and human isolated pulmonary arteries via stimulation of intrinsic nerves, which were inhibited by prazosin or the NaV1 blocker tetrodotoxin. Our results demonstrated that blocking NaV1.7 with GNE8493, PF05089771, or ST2262 abolished or strongly inhibited sympathetic adrenergic responses in guinea-pigs and human vascular smooth muscle. These findings support the hypothesis that pharmacologically inhibiting NaV1.7 could potentially reduce sympathetic and parasympathetic function in specific vascular beds and airways. KEY POINTS: 82% of sympathetic neurons isolated from the stellate ganglion predominantly express NaV1.7 mRNA. NaV1.7 blockers inhibit action potential conduction in postganglionic sympathetic nerves. NaV1.7 blockade substantially inhibits sympathetic nerve-mediated adrenergic contractions in human and guinea-pig blood vessels. Pharmacologically blocking NaV1.7 profoundly affects sympathetic and parasympathetic responses in addition to sensory fibres, prompting exploration into the broader physiological consequences of NaV1.7 mutations on autonomic nerve activity.

Effect of NaV1.7 blockers on post-ganglionic sympathetic nerve function

NaV1.7, also known as the sodium voltage-gated channel alpha subunit 9 (SCN9A), plays a critical role in action potential induction and conduction in pain-causing nociceptors. Accordingly, NaV1.7 blockers may be effective non-opioids analgesics. SCN9A is also expressed in autonomic neurons, but its functional role in the autonomic system is less established. We employed three highly selective NaV1.7 inhibitors to investigate the role of NaV1.7 in action potential conduction in postganglionic sympathetic nerves and in sympathetic adrenergic contractions of blood vessels. Our single neuron rt-PCR analysis revealed that nearly all neurons (32/39) isolated from guinea pig stellate ganglia expressed NaV1.7 mRNA. Genentech Pharmaceuticals provided a highly selective NaV1.7 blocker, GNE8493, which exhibits a selectivity ranging from 500 to 5000-fold for NaV1.7 over all other NaV subtypes. SiteOne Pharmaceuticals contributed another mechanistically distinct and highly selective NaV1.7 blocker, ST2262, with a selectivity of 1000-fold for human NaV1.7 over other NaV subtypes (as reported in Sci Rep. 10:14791, 2020). Unfortunately, ST2262 has little affnity for rodent and guinea pig NaV1.7. We quantified the effect of GNE8493 on postganglionic compound actional potentials (CAP) within the sympathetic trunk of superior cervical ganglia (SCG). GNE8493 inhibited the CAP in the postganglionic neurons of SCG by ~ 70% (n=5, p&lt;0.01). With such a significant impact of the NaV1.7 blocker on post-ganglionic actional potential conduction, we were prompted to explore its potential physiological relevance. Our study involved the isolated pulmonary arteries from anonymous human lung donors (obtained from IIAM) and isolated pulmonary arteries and the abdominal aorta from guinea pigs. We measured smooth muscle tension using standard tissue bath techniques. Stimulation of intrinsic nerves was achieved through electrical field stimulation (EFS) at 12V, 1msec, and 10Hz for 30 seconds, resulting in rapid contractions in all tissues. These contractions were completely prevented by the nonselective NaV1 blocker tetrodotoxin and by the alpha adrenoceptor antagonist prazosin. The contractions were quantified as a percentage of the maximum adrenergic contraction achievable, induced by the addition of 100 μM phenylephrine at the end of the experiment. In guinea pigs, GNE8493 completely blocked sympathetic contractions of the isolated pulmonary arteries (IC50 of -log (M) 5.8) and abdominal aorta (IC50 of -log (M) 6.4 ± 0.3). In human pulmonary arteries, GNE8493 (1 μM) and ST2262 inhibited the sympathetic contractions 94% and 87%, respectively (p&lt;0.01, n=6). These findings support the hypothesis that pharmacological inhibition of NaV1.7 using selective inhibitors has the potential to reduce sympathetic function in specific vascular beds. R35HL155671 - BJU F32HL170490 - JSK. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Selective inhibition of neurogenic, but not agonist-induced contractions by phospholipase A2 inhibitors points to presynaptic phospholipase A2 functions in contractile neurotransmission to human prostate smooth muscle.

Phospholipases A2 (PLA2 ) may be involved in α1 -adrenergic contraction by formation of thromboxane A2 in different smooth muscle types. However, whether this mechanism occurs with α1 -adrenergic contractions of the prostate, is still unknown. While α1 -adrenoceptor antagonists are the first line option for medical treatment of voiding symptoms in benign prostatic hyperplasia (BPH), improvements are limited, probably by nonadrenergic contractions including thromboxane A2 . Here, we examined effects of PLA2 inhibitors on contractions of human prostate tissues. Prostate tissues were obtained from radical prostatectomy. Contractions were induced by electric field stimulation (EFS) and by α1 -adrenergic agonists in an organ bath, after application of the cytosolic PLA2 inhibitors ASB14780 and AACOCF3, the secretory PLA2 inhibitor YM26734, the leukotriene receptor antagonist montelukast, or of solvent to controls. Frequency-dependent contractions of human prostate tissues induced by EFS were inhibited by 25% at 8 Hz, 38% at 16 Hz and 37% at 32 Hz by ASB14780 (1 µM), and by 32% at 16 Hz and 22% at 32 Hz by AACOCF3 (10 µM). None of both inhibitors affected contractions induced by noradrenaline, phenylephrine or methoxamine. YM26734 (3 µM) and montelukast (0.3and 1 µM) neither affected EFS-induced contractions, nor contractions by α1 -adrenergic agonists, while all contractions were substantially inhibited by silodosin (100 nM). Our findings suggest presynaptic PLA2 functions in prostate smooth muscle contraction, while contractions induced by α1 -adrenergic agonists occur PLA2 -independent. Lacking sensitivity to montelukast excludes an involvement of PLA2 -derived leukotrienes in promotion of contractile neurotransmission.

PD02-03 ARF1 GEF INHIBITOR GOLGICIDE A INHIBITS ADRENERGIC AND NON-ADRENERGIC PROSTATE SMOOTH MUSCLE CONTRACTION

You have accessJournal of UrologyCME1 Apr 2023PD02-03 ARF1 GEF INHIBITOR GOLGICIDE A INHIBITS ADRENERGIC AND NON-ADRENERGIC PROSTATE SMOOTH MUSCLE CONTRACTION Alexander Tamalunas, Amin Wendt, Ru Huang, Ruixiao Wang, Yuhan Liu, Beata Rutz, Anna Ciotkowska, Giuseppe Magistro, Christian Stief, and Martin Hennenberg Alexander TamalunasAlexander Tamalunas More articles by this author , Amin WendtAmin Wendt More articles by this author , Ru HuangRu Huang More articles by this author , Ruixiao WangRuixiao Wang More articles by this author , Yuhan LiuYuhan Liu More articles by this author , Beata RutzBeata Rutz More articles by this author , Anna CiotkowskaAnna Ciotkowska More articles by this author , Giuseppe MagistroGiuseppe Magistro More articles by this author , Christian StiefChristian Stief More articles by this author , and Martin HennenbergMartin Hennenberg More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000003219.03AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: Prostate smooth muscle contraction is one of the main targets for medical treatment in lower urinary tract symptoms (LUTS) secondary to benign prostatic hyperplasia (BPH). Due to an unfavorable balance between side effects and limited efficacy, current medications cause high discontinuation rates. Agonist-induced smooth muscle contractions are mediated by activation of a1-adrenoceptors, endothelin and thromboxane, and their intracellular signaling pathways including monomeric GTPases. In general, monomeric GTPases are activated by GEFs, and deactivated by GAPs. Recently, we could show, that the small monomeric GTPase ARF6 is involved in prostate smooth muscle contraction. Here, we examined the effect of the ARF1 GEF inhibitor golgicide A on agonist-induced human prostate smooth muscle contraction. METHODS: Prostate tissues were obtained from patients undergoing radical prostatectomy for prostate cancer (n=30 patients). Contractility of prostate strips was then assessed in an organ bath. Data are presented as mean ±standard deviation, with Emax and EC50 values, while group p-value is given in the diagrams. RESULTS: Noradrenaline-induced adrenergic contractions were decreased by 47±12.1% with golgicide A (10µM) for concentrations of 1-100 µM (Figure 1A), by 30±6.6% for 1-100 µM of phenylephrine-induced contractions (Figure 1B), and by 61±10.1% for 1-100 µM of methoxamine-induced contractions (Figure 1C). Additionally, golgicide A inhibited non-adrenergic contraction by thromboxane A2 analogue U46619 by 82±19.6% for 1-300 µM of U46619-induced contractions (Figure 1D), endothelin-1 induced contraction by 53±7.0% for 1-30 µM of endothelin-1-induced contractions (Figure 1E), and EFS-induced neurogenic contractions by 76±16.6% for 2-32 Hz (Figure 1F). CONCLUSIONS: Golgicide A-sensitive GEFs are involved in regulation of human prostate smooth muscle contraction, by control of ARF1 GEF activity. Knowledge about GEFs and GAPs involved in smooth muscle contraction is still limited. However, increased understanding of prostate smooth muscle contraction may assist in developing new pharmaceutical targets for LUTS medication in the future. Source of Funding: None © 2023 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 209Issue Supplement 4April 2023Page: e69 Advertisement Copyright & Permissions© 2023 by American Urological Association Education and Research, Inc.MetricsAuthor Information Alexander Tamalunas More articles by this author Amin Wendt More articles by this author Ru Huang More articles by this author Ruixiao Wang More articles by this author Yuhan Liu More articles by this author Beata Rutz More articles by this author Anna Ciotkowska More articles by this author Giuseppe Magistro More articles by this author Christian Stief More articles by this author Martin Hennenberg More articles by this author Expand All Advertisement PDF downloadLoading ...

Possible role of transient receptor potential melastatin 4 channels in adrenergic contractions in mouse prostate smooth muscles.

Transient receptor potential melastatin 4 (TRPM4) cation channels are expressed in prostate glands. However, the precise role of these channels in prostate contractility remains unclear. In this study, we examined whether TRPM4 channels were involved in adrenergic contractions in the mouse prostate gland. Adrenergic contractile responses elicited by noradrenaline or electrical field stimulation of the sympathetic nerve were isometrically recorded, and the effects of 9-phenanthrol, a specific TRPM4 channel inhibitor, on those contractile responses were investigated in mouse ventral prostate preparations. 9-phenanthrol (10 or 30 μM) inhibited noradrenaline- and sympathetic nerve-evoked contractions in a concentration-dependent manner. A similar inhibitory effect was observed with another TRPM4 channel inhibitor, 4-chloro-2-(2-(naphthalene-1-yloxy) acetamido) benzoic acid (NBA; 10 μM). Inhibition by 9-phenanthrol and NBA were much greater at lower noradrenaline concentrations and lower stimulus frequencies than those of higher concentrations or frequencies. However, 9-phenanthrol did not inhibit the noradrenaline-induced contractile response when the membrane potential was decreased to approximately 0 mV in the 140 mM K+ medium. Moreover, 9-phenanthrol does not affect noradrenaline-induced increases in spontaneous contractions of cardiac atrial preparation. This agent inhibited noradrenaline-induced contractions in the posterior aorta preparation. However, the inhibitory effect was significantly weaker than that observed in the prostate gland. These results suggest that TRPM4 channels are involved in adrenergic contractions in the mouse prostate gland, possibly through membrane depolarization by their opening; therefore, they might be potential candidates for treating benign prostatic hyperplasia.

STIM/Orai Inhibition as a Strategy for Alleviating Diabetic Erectile Dysfunction Through Modulation of Rat and Human Penile Tissue Contractility and in vivo Potentiation of Erectile Responses

Stromal interaction molecule (STIM)/Orai calcium entry system appears to have a role in erectile dysfunction (ED) pathophysiology but its specific contribution to diabetic ED was not elucidated. To evaluate STIM/Orai inhibition on functional alterations associated with diabetic ED in rat and human penile tissues and on in vivo erectile responses in diabetic rats. Rat corpus cavernosum (RCC) strips from nondiabetic (No DM) and streptozotocin-induced diabetic (DM) rats and human penile resistance arteries (HPRA) and corpus cavernosum (HCC) from ED patients undergoing penile prosthesis insertion were functionally evaluated in organ chambers and wire myographs. Erectile function in vivo in rats was assessed by intracavernosal pressure (ICP) responses to cavernous nerve electrical stimulation (CNES). Expression of STIM/Orai elements in HCC was determined by immunofluorescence and immunoblot. Functional responses in RCC, HCC and HPRA and STIM/Orai protein expression in HCC. In vivo erectile responses to CNES. Inhibition of Orai channels with YM-58483 (20 µM) significantly reduced adrenergic contractions in RCC but more effectively in DM. Thromboxane-induced and neurogenic contractions were reduced by STIM/Orai inhibition while defective endothelial, neurogenic and PDE5 inhibitor-induced relaxations were enhanced by YM-58483 (10 µM) in RCC from DM rats. In vivo, YM-58483 caused erections and attenuated diabetes-related impairment of erectile responses. YM-58483 potentiated the effects of PDE5 inhibition. In human tissues, STIM/Orai inhibition depressed adrenergic and thromboxane-induced contractions in ED patients more effectively in those with type 2 diabetes. Diabetes was associated with increased expression of Orai1 and Orai3 in ED patients. Targeting STIM/Orai to alleviate diabetes-related functional alterations of penile vascular tissue could improve erectile function and potentiate therapeutic effects of PDE5 inhibitors in diabetic ED. Improving effects of STIM/Orai inhibition on diabetes-related functional impairment was evidenced in vitro and in vivo in an animal model and validated in human tissues from ED patients. Functional findings were complemented with expression results. Main limitation was low numbers of human experiments due to limited human tissue availability. STIM/Orai inhibition alleviated alterations of functional responses in vitro and improved erectile responses in vivo in diabetic rats, potentiating the effects of PDE5 inhibition. STIM/Orai inhibition was validated as a target to modulate functional alterations of human penile vascular tissue in diabetic ED where Orai1 and Orai3 channels were upregulated. STIM/Orai inhibition could be a potential therapeutic strategy to overcome poor response to conventional ED therapy in diabetic patients. Sevilleja-Ortiz A, El Assar M, García-Gómez B, et al. STIM/Orai Inhibition as a Strategy for Alleviating Diabetic Erectile Dysfunction Through Modulation of Rat and Human Penile Tissue Contractility and in vivo Potentiation of Erectile Responses. J Sex Med 2022;19:1733-1749.

Vasoactive Effects of Chronic Treatment with Fructose and Slow-Releasing H2S Donor GYY-4137 in Spontaneously Hypertensive Rats: The Role of Nitroso and Sulfide Signalization.

Increased fructose consumption induces metabolic-syndrome-like pathologies and modulates vasoactivity and the participation of nitric oxide (NO) and hydrogen sulfide (H2S). We investigated whether a slow-releasing H2S donor, GYY-4137, could exert beneficial activity in these conditions. We examined the effect of eight weeks of fructose intake on the blood pressure, biometric parameters, vasoactive responses, and NO and H2S pathways in fructose-fed spontaneously hypertensive rats with or without three weeks of GYY-4137 i.p. application. GYY-4137 reduced triacylglycerol levels and blood pressure, but not adiposity, and all were increased by fructose intake. Fructose intake generally enhanced endothelium-dependent vasorelaxation, decreased adrenergic contraction, and increased protein expression of interleukin-6 (IL-6), tumor necrosis factor alpha (TNFα), and concentration of conjugated dienes in the left ventricle (LV). Although GYY-4137 administration did not affect vasorelaxant responses, it restored disturbed contractility, LV oxidative damage and decreased protein expression of TNFα in fructose-fed rats. While the participation of endogenous H2S in vasoactive responses was not affected by fructose treatment, the expression of H2S-producing enzyme cystathionine β-synthase in the LV was increased, and the stimulation of the NO signaling pathway improved endothelial function in the mesenteric artery. On the other hand, chronic treatment with GYY-4137 increased the expression of H2S-producing enzyme cystathionine γ-lyase in the LV and stimulated the beneficial pro-relaxant and anti-contractile activity of endogenous H2S in thoracic aorta. Our results suggest that sulfide and nitroso signaling pathways could trigger compensatory vasoactive responses in hypertensive rats with metabolic disorder. A slow H2S-releasing donor could partially amend metabolic-related changes and trigger beneficial activity of endogenous H2S.

Vascular Effects of Polyphenols from Agrimonia eupatoria L. and Role of Isoquercitrin in Its Vasorelaxant Potential in Human Arteries

Agrimonia eupatoria L. has been traditionally used for the treatment of inflammatory diseases but also as a hypotensive. To our knowledge, only one study has previously suggested an improvement in vascular endothelial function in diabetic conditions, as the underlying mechanisms and responsible compounds are unknown. In this study, we aimed to assess the direct vascular effects of Agrimonia eupatoria L. in human arteries. The infusion elicited a mild increase in basal vascular tone and a significant potentiation of the adrenergic contraction of 49.18% at 0.02 mg/mL, suggesting the presence of compounds with mild vasoconstrictor activity. In contrast, the ethyl acetate fraction inhibited adrenergic contraction by 80.65% at 2 mg/mL and elicited no effect on basal vascular tone. A potent concentration-dependent vasorelaxation was observed for both the infusion and the ethyl acetate fraction (maximal relaxation above 76% and 47%, respectively). Inhibition of nitric oxide synthase and cyclooxygenase elicited significant decreases in the vasorelaxation to the infusion, as, for the ethyl acetate fraction, only the cyclooxygenase pathway appeared to be involved. Isoquercitrin elicited a vasoactivity consistent with the ethyl acetate fraction, suggesting this is a major component responsible for the vasorelaxant properties of A. eupatoria. Further research is warranted to fully evaluate its vasoprotective properties with therapeutic potential in several conditions, e.g., atherosclerosis.

Effects of the Rho Kinase Inhibitor Fasudil on Adrenergic Contractions of Rat Aorta and Mouse Spleen

G‐protein coupled receptors such as α1‐adrenergic receptors (AR) are linked via the G‐protein Gq/G11 to both Ca2+ entry and release of Ca2+ from stores, but can also activate the small GTP binding protein RhoA and Rho kinase to produce contractions involving Ca2+ sensitization. In rat portal vein, tonic but not phasic components of contractions to the α1‐AR agonist phenylephrine involve predominantly the α1B‐AR subtype, and are at least partly mediated by mechanisms involving Rho kinase sensitive to fasudil (Alsufyani &amp; Docherty, 2021). In this study we examined the involvement of Rho Kinase in contractions involving α1‐AR in two further tissues to investigate the predominant α1‐AR subtype involved. We have examined the Rho kinase inhibitor fasudil in rat aorta and mouse spleen, tissues containing multiple α1‐AR subtypes. Male Wistar rats (250‐300g) and C57 mice were killed by CO2 overdose and cervical dislocation, and rat aortic rings and mouse spleens were mounted in organ baths under 0.5 g tension, and bathed with Krebs‐Henseleit solution at 37oC. Tissues were contracted with cumulative additions of norepinephrine (NE) in 0.5 log unit increments, over the concentration range 10‐8‐10‐3M. Both α2‐ and α1‐ARs mediate contractions of mouse spleen and the major α1‐AR involved is the α1B‐adrenoceptor, and this receptor interacts with an α1D‐AR at low, and an α1A‐AR at high, NE concentrations (Alsufyani et al., 2021). In mouse spleen, fasudil (3 μM) reduced contractions to both low and high concentrations of NE. In rat aorta, contractions to NE are reported to be mediated by two subtypes of α1‐AR: α1B‐ and α1D‐ARs, with responses to low but not high concentrations of NE being sensitive to the antagonist BMY7378. Responses to high concentrations of NE in rat aorta are sensitive to the non‐selective α1‐AR antagonist prazosin and the putative α1B‐AR antagonist cyclazosin. Fasudil (3 and 10 μM) significantly reduced contractions to both low and high concentrations of NE in rat aorta in a dose dependent manner in terms of maximum response. It is concluded that α1‐AR mediated contractions to NE in both rat aorta and mouse spleen are sensitive to non‐competitive block by the Rho kinase inhibitor fasudil in concentrations of 3‐10 μM. Although multiple subtypes of α1‐adrenoceptor are involved in contractions of both rat aorta and mouse spleen, the evidence suggests that, as previously reported in rat portal vein, α1B‐adrenoceptors are most likely linked to the Rho kinase pathwayAlsufyani HA &amp; Docherty, J.R. Can J Physiol Pharmacol. 2021 Jun;99(6):654‐659. Alsufyani HA, et al. Basic Clin Pharmacol Toxicol. 2021 Dec;129(6):416‐426.

Influence of pole‐like kinase 1 on controlling contractility of the urinary tract and gene activated patterns of human bladder smooth muscle cells

BackgroundPole like kinase (PLK)‐mediated promotion of proliferation and smooth muscle contraction in the lower urinary tract has been recently suggested and may offer putative targets and novel compounds for treatments of lower urinary tract symptoms. However, PLK function for contractility of smooth muscle and proliferation of bladder smooth muscle cells are unknown and the specificity of PLK inhibitors has been questioned.MethodsHere, human bladder smooth muscle cells (hBSMCs) inhibited with five PLK inhibitors (TAK960, MLN0905, Wortmannin, GW843682X, Centrinone‐B) and hBSMCs cultured in smooth muscle medium were compared. hBSMCs characteristics according to criteria set by the International Society of Cellular Therapy were confirmed by flow cytometry. Proliferation and viability were assessed by CCK‐8 assay. Contractions of human ureterovesical junction smooth muscle from donation after circulatory death renal transplantation were studied in an organ bath, where they were induced by the α1‐adrenoceptor agonists noradrenaline, phenylephrine, and electric field stimulation. Transcriptome sequencing of hBSMCs were obtained using NEBNext®UltraTM RNA Library Prep Kit for Illumina®. Gene set enrichment analysis was performed using the Kyoto Encyclopedia of Genes and Genomes and gene ontology terms.ResultsViability and proliferation of hBSMCs were reduced in a concentration‐ and time‐dependent manner. TAK960 inhibits adrenergic contractions of human ureterovesical junction smooth muscle, and proliferation of hBSMCs. No significant differences in other PLK inhibitors combined with control were found. Compared to hBSMCs in control medium, 323 upregulated genes and 160 downregulated genes were significantly altered by TAK960 24hours 500nM, respectively (FDR=0.05, FC=1.5). Genes most prominently and significantly upregulated by PLK inhibiting condition were IFI30, GBP4, VTN, SCG2, OAS2, RRAD, TNFSF10, KRT18, SFN, meanwhile, downregulated was CCL11 (log2FC ≥1.5 or ≤ ‐1.5). The most significantly relative signaling pathway included cytokine‐cytokine receptor interaction, systemic lupus erythematosus and transcriptional misregulation cancer (enrichment factor ≥0.05). Accession to cite for these sequences read archive data is PRJNA714246.ConclusionOur findings point to a certain role of PLK‐1 in of proliferation in hBSMCs and contractility in ureterovesical junction smooth muscle, which may be of relevance in lower urinary tract symptom. PLK‐1 inhibitors block these effects and show shared and divergent effects. PLK‐1 regulation of bladder smooth muscle growth and contraction may include genomic and nongenomic mechanisms.

Mouse aortic biomechanics are affected by short-term defective autophagy in vascular smooth muscle cells

The physiology of vascular smooth muscle (VSMC) cells is affected by autophagy, a catabolic cellular mechanism responsible for nutrient recycling. Autophagy-inducing compounds may reverse arterial stiffening, whereas congenital VSMC-specific autophagy deficiency promotes arterial stiffening. The elevated aortic stiffness in 3.5-month-old C57Bl/6 mice, in which the essential autophagy-related gene Atg7 was specifically deleted in the VSMCs (Atg7F/F SM22α-Cre+ mice) was mainly due to passive aortic wall remodeling. The present study investigated whether aortic stiffness was also modulated by a shorter duration of autophagy deficiency. Therefore, aortic segments of 2-month-old Atg7F/F SM22α-Cre+ mice were studied. Similarly to the older mice, autophagy deficiency in VSMCs promoted aortic stiffening by elastin degradation and elastin breaks, and increased the expression of the calcium binding protein S100A4 (+ 157%), the aortic wall thickness (+ 27%), the sensitivity of the VSMCs to depolarization and the contribution of VGCC mediated Ca2+ influx to α1 adrenergic contractions. Hence, all these phenomena occurred before the age of 2 months. When compared to autophagy deficiency in VSMCs at 3.5 months, shorter term autophagy deficiency led to higher segment diameter at 80 mmHg (+ 7% versus − 2%), normal baseline tonus (versus increased), unchanged IP3-mediated phasic contractions (versus enhanced), and enhanced endothelial cell function (versus normal). Overall, and because in vivo cardiac parameters or aortic pulse wave velocity were not affected, these observations indicate that congenital autophagy deficiency in VSMCs of Atg7F/F SM22α-Cre+ mice initiates compensatory mechanisms to maintain circulatory homeostasis.

Possible involvement of transient receptor potential melastatin 4 channels in the adrenergic contraction in mouse prostate smooth muscles

Possible involvement of transient receptor potential melastatin 4 channels in the adrenergic contraction in mouse prostate smooth muscles

Lenalidomide and pomalidomide inhibit growth of prostate stromal cells and human prostate smooth muscle contraction

AimsMedical treatment for lower urinary tract symptoms secondary to benign prostatic hyperplasia is characterized by an unfavorable balance between limited efficacy and pronounced side effects. We recently reported, that thalidomide reduces prostate smooth muscle contraction and inhibits cell growth. Like thalidomide, its analogs lenalidomide and pomalidomide are also in clinical use. Therefore, we investigated the effects of lenalidomide and pomalidomide on human prostate smooth muscle contraction, cytoskeletal organization, and growth-related functions in stromal cells. Materials and methodsProliferation was assessed by EdU assay and colony formation, cytoskeletal organization by phalloidin staining, cell viability by CCK8, and apoptosis and cell death by flow cytometry in cultured prostate stromal cells (WPMY-1). Contractions of human prostate tissues from radical prostatectomy were induced by methoxamine, noradrenaline, phenylephrine, endothelin-1, U46619, or electric field stimulation (EFS) in an organ bath. Key findingsProliferation of WPMY-1 cells was significantly reduced by lenalidomide (5–200 μM) and pomalidomide (2.5–5 μM). In parallel, organization of actin filaments collapsed after treatment with lenalidomide and pomalidomide. Lenalidomide and pomalidomide inhibited both adrenergic contractions and non-adrenergic contractions as well as neurogenic contractions induced by EFS. Neither reduction in viability, nor increase in cell death or apoptosis was observed in WPMY-1 cells. SignificanceThalidomide-derivatives impair growth of human prostate stromal cells, without showing a decrease in cell viability and, in parallel, inhibit adrenergic, neurogenic, and non-adrenergic contractions by breakdown of the actin cytoskeleton. Urodynamic effects in vivo appear possible.

Picotamide inhibits a wide spectrum of agonist‐induced smooth muscle contractions in porcine renal interlobar and coronary arteries

Picotamide is a thromboxane A2 (TXA2) receptor antagonist and TXA2 synthase inhibitor. In clinical studies, it has been considered as a platelet aggregation inhibitor and improved renal function. In vitro studies suggested inhibition of smooth muscle contraction by picotamide, which is poorly understood. Here, we examined effects of picotamide on contractions of renal interlobar and coronary porcine arteries, induced by different vasoconstrictors. Contractions were induced in an organ bath by agonists or electric field stimulation (EFS). Picotamide inhibited EFS‐induced contractions of interlobar arteries around 50% using concentrations of 100 and 300 µM. In interlobar arteries, concentration response curves for contractions induced by three different α1‐adrenoceptor agonists were shifted to the right by picotamide (2–10‐fold increases in EC50). In coronary arteries, α1‐adrenergic contractions were inhibited without right shift (approx. 50%). Contractions induced by two different cholinergic agonists in coronary arteries were inhibited by picotamide (≥50%) withouth right shift. Inhibition of serotonin‐induced contractions by picotamide showed features of a right shift, whereas contractions induced by the TXA2 analog U46619, angiotensin‐II, and endothelin‐1 were inhibited by picotamide in interlobar and coronary arteries without right shifts and to different degree. Picotamide inhibits a wide spectrum of vasoconstrictor‐induced contractions in porcine interlobar and coronary arteries. Inhibition of vasocontraction may contribute to beneficial effects of picotamide in the cardiovascular system and kidney.

Fructose Intake Impairs the Synergistic Vasomotor Manifestation of Nitric Oxide and Hydrogen Sulfide in Rat Aorta.

In this study, we evaluated the effect of eight weeks of administration of 10% fructose solution to adult Wistar Kyoto (WKY) rats on systolic blood pressure (SBP), plasma and biometric parameters, vasoactive properties of the thoracic aorta (TA), NO synthase (NOS) activity, and the expression of enzymes producing NO and H2S. Eight weeks of fructose administration did not affect SBP, glycaemia, or the plasma levels of total cholesterol or low-density and high-density lipoprotein; however, it significantly increased the plasma levels of γ-glutamyl transferase and alanine transaminase. Chronic fructose intake deteriorated endothelium-dependent vasorelaxation (EDVR) and increased the sensitivity of adrenergic receptors to noradrenaline. Acute NOS inhibition evoked a reduction in EDVR that was similar between groups; however, it increased adrenergic contraction more in fructose-fed rats. CSE inhibition decreased EDVR in WKY but not in fructose-fed rats. The application of a H2S scavenger evoked a reduction in the EDVR in WKY rats and normalized the sensitivity of adrenergic receptors in rats treated with fructose. Fructose intake did not change NOS activity but reduced the expression of eNOS and CBS in the TA and CSE and CBS in the left ventricle. Based on our results, we could assume that the impaired vascular function induced by increased fructose intake was probably not directly associated with a decreased production of NO, but rather with impairment of the NO–H2S interaction and its manifestation in vasoactive responses.

Effects of trans- versus cis-resveratrol on adrenergic contractions of the rat tail artery and role of endothelium.

The health benefits of the natural polyphenol trans‐resveratrol may play an important role in preventing a variety of diseases. Resveratrol has been shown to reduce blood pressure and improve metabolic diseases such as type 2 diabetes mellitus and obesity. Our previous studies examined the role of K+ channels in the vasorelaxation responses to trans‐resveratrol in the rat tail artery. During these studies, we uncovered a novel transient contraction prior to the sustained relaxation effect of trans‐resveratrol. Thus, the purpose of this study was to determine the role of the endothelium in these vascular contraction and relaxation responses to trans‐resveratrol. We additionally sought to determine if the cis‐isomer of resveratrol exerts any of the same vascular effects as the trans‐isomer. The vascular responses to trans‐resveratrol were examined in rat tail arteries with intact or denuded endothelium over a 2‐hr period. Additionally, the vascular responses to trans‐ and cis‐resveratrol were compared in rat tail arteries with intact endothelium. Both the transient contractile response and the persistent relaxation response to trans‐resveratrol were similar in the arterial rings with intact or denuded endothelium. There was a significant correlation between the initial contraction‐enhancing action of trans‐resveratrol and the magnitude of the sustained relaxation for vessels with both intact and denuded endothelium. Moreover, we demonstrated that cis‐resveratrol produced a significantly greater relaxation response as compared to trans‐resveratrol without the initial contractile response. These data demonstrate the role of the vascular smooth muscle in the vascular responses to resveratrol and the potential clinical benefits of the cis‐isomer of resveratrol as compared to the trans‐isomer.

MP59-04 SYSTEMIC VASCULAR FUNCTION OF KIDNEY DONOR INFLUENCES SHORT-TERM RENAL PERFORMANCE OF RECEPTOR

INTRODUCTION AND OBJECTIVE: Renal function is highly dependent on adequate vascular function. This is especially relevant when considering the increasing use of older and marginal kidney donors to overcome organ unavailability. Our aim was to evaluate the influence of donor’s vascular function at different vascular beds on renal function in kidney recipients. METHODS: Aorta, mesenteric microvessels (MM) and corpus cavernosum (CC) specimens were obtained at the time of kidney explant for transplantation and vascular function was ex vivo determined by evaluating adrenergic contractions to norepinephrine and endothelium-dependent vasodilations to bradykinin (BK, MM) or acetylcholine (ACh, CC). RESULTS: Vascular function in each donor (n=38) was correlated with serum creatinine levels (SC) and glomerular filtration rate (GFR) in each receptor(s) (n=49). Endothelium-dependent vasodilation of donor MM (pEC50 for BK) was significantly and negatively associated with SC (r2=0.245, p=0.003) while positively with GFR (r2=0.267, p=0.002) at 3 months after transplantation but significance was lost at 6 months (SC: r2=0.114, p=0.092; GFR: r2=0.131, p=0.069). Receptor’s GFR was also positively associated with endothelium-dependent relaxation (pEC50 for ACh) in CC from male donors (r2=0.310;p=0.006) and negatively with adrenergic contraction in aorta (r2=0.289, p=0.001) at 3 months. These associations were also lost at 6 months after transplantation. Moreover, receptors with GFR≥60 ml/min at 3 months received the kidney from donors displaying a significantly better endothelial vasodilation in MM (pEC50 for BK 7.60±0.25 vs. 6.71±0.15, p=0.004). Conversely, donors in the highest tertile of endothelium-dependent relaxation in MM or CC displayed lower SC and higher GFR at 3 and 6 months after transplantation. CONCLUSIONS: Systemic vascular function of kidney donor is a determinant for renal function in the receptor at short-term but the influence of receptor condition on renal performance probably prevails at later time. Source of Funding: No

MP06-14 INHIBITION OF PROSTATE SMOOTH MUSCLE CONTRACTION BY THALIDOMIDE: A NOVEL REMEDY IN LUTS?

You have accessJournal of UrologyBenign Prostatic Hyperplasia: Basic Research & Pathophysiology (MP06)1 Apr 2020MP06-14 INHIBITION OF PROSTATE SMOOTH MUSCLE CONTRACTION BY THALIDOMIDE: A NOVEL REMEDY IN LUTS? Alexander Tamalunas*, Cora Sauckel, Anna Ciotkowska, Christian Gratzke, Christian G. Stief, and Martin Hennenberg Alexander Tamalunas*Alexander Tamalunas* More articles by this author , Cora SauckelCora Sauckel More articles by this author , Anna CiotkowskaAnna Ciotkowska More articles by this author , Christian GratzkeChristian Gratzke More articles by this author , Christian G. StiefChristian G. Stief More articles by this author , and Martin HennenbergMartin Hennenberg More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000000820.014AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: Medical treatment in benign prostatic hyperplasia (BPH) includes reduction of smooth muscle tone for rapid relieve of lower urinary tract symptoms (LUTS). Current medications cause high rates of patient non-compliance due to an unfavourable balance between side effects and efficacy, with discontinuation rates peaking up to 90% for combination therapies. We have recently shown that thalidomide reduces prostate stromal cell growth and modulates cytoskeletal actin organization. Here, we addressed effects of thalidomide on adrenergic and non-adrenergic contractions of human prostate smooth muscle. METHODS: Prostate tissues were obtained from patients undergoing radical prostatectomy for prostate cancer (n=40 patients). Contractility of prostate strips was then assessed in an organ bath. RESULTS: Noradrenaline-induced adrenergic contractions were decreased up to over 56 % with thalidomide (100µM) for concentrations of 1-100 µM (p<0.05, p<0.03, p<0.02, p<0.01, p<0.02 for 1 µM, 3 µM, 10 µM, 30 µM and 100 µM noradrenaline, respectively, for thalidomide vs. control, fig. A). Thalidomide caused significant inhibition of up to 90 % at 1-100 µM for both methoxamine- and phenylephrine-induced contractions (p<0.04, p<0.03, p<0.04, p<0.04, p<0.03 and p<0.05, p<0.02, p<0.03, p<0.02, p<0.01 for 1 µM, 3 µM, 10 µM, 30 µM and 100 µM phenylephrine and methoxamine, respectively, for thalidomide vs. control, fig. B & C). Thalidomide inhibited non-adrenergic contraction with thromboxane A2 analogue U46619 at 0.1 µM, 0.3 µM, 3 µM, 10 µM and 30 µM (p<0.01 for 0.1 µM, 0.3 µM, 10 µM and 30 µM and p<0.02 for 3 µM U46619, respectively, for thalidomide vs. control) and up to 58 % of endothelin-1-induced contractions at 0.3 µM, 1 µM and 3 µM (p<0.02, p<0.03 and p<0.03 for 0.3 µM, 1 µM and 3 µM endothelin-1, respectively, for thalidomide vs. control, fig. D & E). Significant inhibition up to 73 % also occurred in EFS-induced neurogenic contractions for 2-32 Hz (p<0.01, p<0.05, p<0.04, p<0.03, p<0.03 for 2, 4, 8, 16 and 32 Hz, respectively, for thalidomide vs. control, fig. F). CONCLUSIONS: Thalidomide not only inhibits prostate stromal cell growth and modulates cytoskeletal development, but also inhibits adrenergic and non-adrenergic prostate smooth muscle contraction. Therefore, thalidomide may be superior to current medical treatment. Source of Funding: None. © 2020 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 203Issue Supplement 4April 2020Page: e56-e56 Advertisement Copyright & Permissions© 2020 by American Urological Association Education and Research, Inc.MetricsAuthor Information Alexander Tamalunas* More articles by this author Cora Sauckel More articles by this author Anna Ciotkowska More articles by this author Christian Gratzke More articles by this author Christian G. Stief More articles by this author Martin Hennenberg More articles by this author Expand All Advertisement PDF downloadLoading ...

Relaxant and antiadrenergic effects of ranolazine in human saphenous vein.

Ranolazine improves vascular function in animal models. We evaluate the effects of ranolazine on vascular function and adrenergic response in human saphenous vein. Rings from 53 patients undergoing coronary artery bypass grafting were mounted in organ baths. Concentration-response curves to ranolazine were constructed in rings precontracted with phenylephrine, endothelin-1, vasopressin, KCl and the thromboxane A2 analogue U-46619. In rings precontracted with phenylephrine, relaxation to ranolazine was tested in the absence and presence of endothelial factors inhibitors, K+ channel blockers and verapamil. The effects of ranolazine on frequency-response and concentration-response curves to phenylephrine were performed in the absence and presence of endothelial factors inhibitors and K+ channel blockers. Endothelial nitric oxide synthase, α1 adrenergic receptor and large conductance Ca2+-activated K+ channel protein expressions were measured by Western blotting. Ranolazine (10-9-10-4 M) produced a concentration-dependent relaxation only in rings precontracted with phenylephrine that was reduced by endothelial denudation, NG-nitro-l-arginine methyl ester (10-4 M), charybdotoxin (10-7 M) and verapamil (10-6 M). Ranolazine diminished adrenergic contractions induced by electrical field stimulation (2-4 Hz) and phenylephrine (10-9-10-5 M) that were prevented by tetraethylammonium (10-3 M) and charybdotoxin (10-7 M). Ranolazine significantly decreased α1 adrenergic receptor and increased large conductance Ca2+-activated K+ channel protein expression in the saphenous vein. Ranolazine diminishes the adrenergic vasoconstriction, acting as α1 antagonist, and by increasing large conductance Ca2+-activated K+ channel involvement. The relaxant effects of ranolazine are partially mediated by endothelial nitric oxide, large conductance Ca2+-activated K+ channels and the blockade of voltage-dependent Ca2+ channels.

A NAV2729-sensitive mechanism promotes adrenergic smooth muscle contraction and growth of stromal cells in the human prostate

Voiding symptoms in benign prostatic hyperplasia (BPH) are driven by prostate smooth muscle contraction and prostate growth. Smooth muscle contraction in the prostate and other organs critically depends on activation of the small monomeric GTPase RhoA and probably Rac1. A role of another GTPase, ADP-ribosylation factor 6 (ARF6), for smooth muscle contraction has been recently suggested by indirect evidence but remains to be proven for any organ. Here, we report effects of NAV2729, an inhibitor with assumed specificity for ARF6, in human prostate tissues and cultured prostate stromal cells (WPMY-1). NAV2729 (5 μm) inhibited neurogenic and α1-adrenergic contractions of human prostate tissues. Contractions induced by endothelin-1, by the thromboxane A2 agonist U46619, or by high molar KCl were not inhibited. Correlation analyses suggested up-regulation of prostatic ARF6 expression with increasing degree of BPH, as ARF6 expression increased with the content of prostate-specific antigen (PSA) of prostate tissues. NAV2729 inhibited ARF6 activity but not other GTPases (ARF1, RhoA, Rac1) in prostate tissues and in WPMY-1 cells. Proliferation of WPMY-1 cells was inhibited concentration-dependently by NAV2726, as reflected by decreased viability, 5-ethynyl-2'-deoxyuridine (EdU) assay, colony formation assay, and expression of Ki-67. Silencing of ARF6 expression mimicked effects of NAV2729 on viability and in the EdU assay. Effects of NAV2729 on viability and proliferation were attenuated in cells with silenced ARF6 expression. Our findings suggest that a NAV2729-sensitive mechanism promotes adrenergic contraction and stromal cell growth in the human prostate, which is probably ARF6-mediated. Similar actions in other organs and urodynamic effects of NAV2729 appear possible.