The bladders of intact rats were collected, and bladder strips were cut and left in a urothelial cell culture medium (Cat No 4321; ScienCell), in culture medium with 1.25 g/l of phenylephrine (P6126, Sigma) or in culture medium with 1.25 g/l of phenylephrine and 0.1 g/l of the alpha 1 A adrenoceptor blocker silodosin (commercial formulation available from Recordati Urorec®). After 24 h, the medium was collected to measure NGF by ELISA.
P6126
Manufactured by Merck Group
P6126 is a laboratory equipment product from Merck Group. It is designed for general laboratory use. The core function of this product is to provide a controlled environment for various experiments and research activities. Further details on the specific features and intended use are not available.
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Lab products found in correlation
7 protocols using p6126
1
Bladder Strip Phenylephrine Response
2
Aortic Ring Contractility Assay
3
Phenylephrine-Induced SOCE Activation
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crSTIM1−/− and littermate control male mice were injected subcutaneously with either sterile saline solution or phenylephrine (PE; 15 mg/kg; Sigma P6126), as described previously (van Berlo et al., 2011 (link)). Briefly, mice injected with saline and PE were either euthanized and hearts were snap‐frozen for subsequent kinomic and immunoblot analysis 15 min following treatment or used for electrocardiogram (ECG) studies (see below). PE, an α1‐adrenergic receptor agonist, has been shown to activate SOCE (Hunton et al., 2002 (link)) and induce ERK1/2 phosphorylation (van Berlo et al., 2011 (link)). The dose of 15 mg/kg PE and treatment time of 15 min were both selected due to previous reports demonstrating a robust increase in ERK1/2 phosphorylation in wild‐type mice (van Berlo et al., 2011 (link)).
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crSTIM1−/− and littermate control male mice were injected subcutaneously with either sterile saline solution or phenylephrine (PE; 15 mg/kg; Sigma P6126), as described previously (van Berlo et al., 2011 (link)). Briefly, mice injected with saline and PE were either euthanized and hearts were snap‐frozen for subsequent kinomic and immunoblot analysis 15 min following treatment or used for electrocardiogram (ECG) studies (see below). PE, an α1‐adrenergic receptor agonist, has been shown to activate SOCE (Hunton et al., 2002 (link)) and induce ERK1/2 phosphorylation (van Berlo et al., 2011 (link)). The dose of 15 mg/kg PE and treatment time of 15 min were both selected due to previous reports demonstrating a robust increase in ERK1/2 phosphorylation in wild‐type mice (van Berlo et al., 2011 (link)).
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Osmotic Minipump Implantation in Mice
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Myocardial Strip Contractility Assay
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Isometric Tension Measurement of Murine Aorta
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For isometric tension measurement, thoracic aortas were isolated from mice and rinsed in Krebs solution. After careful removal of connective tissue, the aorta was cut into 2-mm-long segments and mounted in a wire myography chamber (Danish Myo Technology, 620 M) filled with aerated (95% O2-5% CO2) Krebs solution and maintained at 37 °C. For equilibration, isolated aortic rings were stretched until receiving resting tension and exposed to 60 mM KCl Krebs solution to analyze contractility. For analysis of the contractile response to cumulatively administered vasoconstrictive agonists, each aorta was treated with PE (Sigma, P6126). Assessment of aorta tension was performed in a blinded trial.
7
Vascular Smooth Muscle Cell Contractility
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VSMC contractility was measured as described previously 17, 18 . Briefly, primary VSMCs were cultured for 4-5 days. In some experiments, estradiol (100 ng/ml, Sigma-Aldrich) or testosterone (6.5 ng/ml, Sigma-Aldrich) was added at the beginning of the culture. The plates were viewed in the chamber of a Zeiss microscope with environmental controls (37°C and 5% CO2). The cells were stimulated with (R)-(-)-phenylephrine hydrochloride (PE, 20 M, P6126, Sigma-Aldrich), and images were captured continuously for 15 min at 1 frame per min. Fifteen or more cells were selected randomly with their length measured at the time points indicated, using Carl Zeiss Axiovision software. The results are reported as contraction percentages (% contraction) calculated as follows: % contraction=100 x (cell length at time 0 -cell length at time X) / cell length at time 0.
Means and standard error of the means (SEM) of contraction percentages of 15 or more cells were presented.
Means and standard error of the means (SEM) of contraction percentages of 15 or more cells were presented.
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