Birgit Eichhorn

Vascular large conductance calcium-activated potassium channels: functional role and therapeutic potential.

Large-conductance Ca2+-activated K+ channels (BKCa or maxiK channels) are expressed in different cell types. They play an essential role in the regulation of various cell functions. In particular, BKCa channels have been extensively studied in vascular smooth muscle cells, where they contribute to the control of vascular tone. They facilitate the feedback regulation against the rise of intracellular Ca2+, membrane depolarization and vasoconstriction. BKCa channels promote a K+ outward current and lead to membrane hyperpolarization. In endothelial cells expression and function of BKCa channels play an important role in the regulation of the vascular smooth muscle activity. Endothelial BKCa channels modulate the biosyntheses and release of various vasoactive modulators and regulate the membrane potential. Because of their regulatory role in vascular tone, endothelial BKCa channels have been suggested as therapeutic targets for the treatment of cardiovascular diseases. Hypertension, atherosclerosis, and diabetes are associated with altered current amplitude, open probability, and Ca2+-sensing of BKCa channels. The properties of BKCa channels and their role in endothelial and vascular smooth muscle cells would address them as potential therapeutic targets. Further studies are necessary to identify the detailed molecular mechanisms of action and to investigate selective BKCa channels openers as possible therapeutic agents for clinical use.

Catecholamines Relax Detrusor through β2-Adrenoceptors in Mouse and β3-Adrenoceptors in Man

(-)-Isoproterenol [4-[1-hydroxy-2-[(1-methylethyl)amino]ethyl]-1,2-benzene diol hydrochloride] relaxes murine detrusor through β-adrenoceptors (ARs); however, the β-AR subtypes involved are unknown. β2-ARs have been associated with caveolae, plasma-lemmal scaffolding domains that are absent in caveolin-1 (cav-1) knockout (KO) mice. Here, we studied detrusor responses in the absence and presence of β-AR subtype-selective antagonists in wild-type (WT) and cav-1 KO mice. To inquire whether the murine detrusor model is relevant to man, β-AR subtypes that mediate (-)-isoproterenol-evoked human detrusor relaxation were investigated. In WT mice, (-)-isoproterenol concentration-dependently relaxed the KCl (40 mM)-precontracted detrusor (-logEC50M = 8.04, Emax = 62%). The effects of (-)-isoproterenol were surmountably antagonized by the β2-AR-selective antagonist ICI 118,551 [(±)-1-[2,3-(dihydro-7-methyl-1H-inden-4-yl)oxy]-3-[(1-methylethyl)amino]-2-butanol] (pKB = 9.28) but not affected by the β1-AR-selective antagonist CGP 20712 [1-[2-((3-carbamoyl-4-hydroxy)phenoxy)ethylamino]-3-[4-(1-methyl-4-trifluoromethyl-2-imidazolyl)phenoxy]-2-propanol] and β3-AR-selective L-748,337 [(S)-M-[4-[2-[3-[3-[acetamidomethyl)phenoxy)-2-hydroxypropyl]-amino]-ethyl]-phenylbenzsulfonamide)], suggesting involvement of β2-AR only. The cav-1 KO detrusor displayed significant contractile dysfunction. (-)-Isoproterenol was less potent and efficient in relaxing detrusor from cav-1 KO (-logEC50M, 7.76; Emax = 44%), but ICI 118,551 caused similar antagonism (pKB = 9.15), suggesting that β2-AR function persisted in cav-1 KO. The β3-AR-selective antagonist L-748,337 in the presence of ICI 118,551 and CGP 20712 caused additional blockade of (-)-isoproterenol effects in cav-1 KO, consistent with a β3-AR involvement during relaxation and suppression of this effect in WT. (-)-Isoproterenol relaxed human detrusor muscle precontracted with carbachol (-logEC50M = 6.39, Emax = 52%). However, the effects of (-)-isoproterenol in human detrusor were not blocked by CGP 20712 or ICI 118,551 but antagonized by L-748,337 (pKB = 7.65). We conclude that murine detrusor relaxation occurs via β2-AR, and loss of caveolae does not perturb β2-AR function but unmasks an additional activation of β3-AR. In contrast, detrusor relaxation in man is mediated exclusively via β3-AR.

Catecholamines relax detrusor through beta 2-adrenoceptors in mouse and beta 3-adrenoceptors in man.

(-)-Isoproterenol [4-[1-hydroxy-2-[(1-methylethyl)amino]ethyl]-1,2-benzene diol hydrochloride] relaxes murine detrusor through β-adrenoceptors (ARs); however, the β-AR subtypes involved are unknown. β2-ARs have been associated with caveolae, plasma-lemmal scaffolding domains that are absent in caveolin-1 (cav-1) knockout (KO) mice. Here, we studied detrusor responses in the absence and presence of β-AR subtype-selective antagonists in wild-type (WT) and cav-1 KO mice. To inquire whether the murine detrusor model is relevant to man, β-AR subtypes that mediate (-)-isoproterenol-evoked human detrusor relaxation were investigated. In WT mice, (-)-isoproterenol concentration-dependently relaxed the KCl (40 mM)-precontracted detrusor (-logEC50M = 8.04, Emax = 62%). The effects of (-)-isoproterenol were surmountably antagonized by the β2-AR-selective antagonist ICI 118,551 [(±)-1-[2,3-(dihydro-7-methyl-1H-inden-4-yl)oxy]-3-[(1-methylethyl)amino]-2-butanol] (pKB = 9.28) but not affected by the β1-AR-selective antagonist CGP 20712 [1-[2-((3-carbamoyl-4-hydroxy)phenoxy)ethylamino]-3-[4-(1-methyl-4-trifluoromethyl-2-imidazolyl)phenoxy]-2-propanol] and β3-AR-selective L-748,337 [(S)-M-[4-[2-[3-[3-[acetamidomethyl)phenoxy)-2-hydroxypropyl]-amino]-ethyl]-phenylbenzsulfonamide)], suggesting involvement of β2-AR only. The cav-1 KO detrusor displayed significant contractile dysfunction. (-)-Isoproterenol was less potent and efficient in relaxing detrusor from cav-1 KO (-logEC50M, 7.76; Emax = 44%), but ICI 118,551 caused similar antagonism (pKB = 9.15), suggesting that β2-AR function persisted in cav-1 KO. The β3-AR-selective antagonist L-748,337 in the presence of ICI 118,551 and CGP 20712 caused additional blockade of (-)-isoproterenol effects in cav-1 KO, consistent with a β3-AR involvement during relaxation and suppression of this effect in WT. (-)-Isoproterenol relaxed human detrusor muscle precontracted with carbachol (-logEC50M = 6.39, Emax = 52%). However, the effects of (-)-isoproterenol in human detrusor were not blocked by CGP 20712 or ICI 118,551 but antagonized by L-748,337 (pKB = 7.65). We conclude that murine detrusor relaxation occurs via β2-AR, and loss of caveolae does not perturb β2-AR function but unmasks an additional activation of β3-AR. In contrast, detrusor relaxation in man is mediated exclusively via β3-AR.

Impaired vascular function in small resistance arteries of LOX-1 overexpressing mice on high-fat diet

AIMS LOX-1 is a major vascular receptor for oxidized low-density lipoprotein (oxLDL). In this study, we analysed the impact of LOX-1 overexpression and high dietary fat intake on vascular function in small resistance arteries. METHODS AND RESULTS Relaxation of mesenteric arteries was measured using a wire myograph. Compared with the control group, mice overexpressing LOX-1 on a high-fat diet (FD) had preserved vascular smooth muscle relaxation, but impaired endothelium-dependent relaxation via NO. Vascular NO availability was decreased by exaggerated formation of reactive oxygen species and decreased endothelial NO synthase expression. Endothelium-derived hyperpolarizing factor (EDHF)-mediated relaxation via cytochrome P450 metabolites was increased in LOX-1 + FD animals, but did not completely compensate for the loss of NO. Currents of calcium-activated potassium channels with large conductance (BKCa channels) were measured by the voltage-clamp method. The BKCa current amplitudes were not altered in endothelial cells, but highly increased in vascular smooth muscle cells from resistance arteries of LOX-1-overexpressing mice on FD. BK(Ca) currents were activated by low-dose H2O2 and cytochrome P450 metabolites 11,12-EET and 14,15-EET as EDHF in control mice. CONCLUSION LOX-1 overexpression and FD caused functional changes in endothelial and vascular smooth muscle cells of small resistance arteries.

Investigation of murine Vasodynamics by Fourier Domain Optical Coherence Tomography

In vivo imaging of blood vessels obtain useful insights in characterizing the dynamics of vasoconstriction and vasodilation. Fourier domain optical Coherence Tomography (FD-OCT) imaging technique permits in vivo investigation of blood vessels in their anatomical context without preparation traumata by temporal resolved image stacks. OCT is an optical, contact less imaging technique based on Michelson interferometry of short coherent near infrared light. Particularly by the possibility of a contact-less measurement and the high axial resolution up to 10 microns OCT is superior to an investigation by ultra sound measurement. Furthermore we obtain a high time resolution of vessel dynamic measurements with the used Fourier domain OCT-system by a high A-scan rate [1,22kHz]. In this study the model of saphenous artery was chosen for analyzing function and dynamics. The arteria saphena in the mouse is a suitable blood vessel due to the small inner diameter, a sensitive response to vasoactive stimuli and an advantageous anatomically position. Male wild type mice (C57BL/6) at the age of 8 weeks were fed control or high-fat diet for 10 weeks before analyzing the vasodynamics. The blood vessel was stimulated by dermal application of potassium to induce vasoconstriction or Sodium-Nitroprusside (SNP) to induce vasodilation. The morphology of the a. saphena and vein was determined by 3D image stacks. Time series (72 seconds, 300x512 pixel per frame) of cross-sectional images were analysed using semi automatic image processing software. Time course of dynamic parameters of the vessel was measured.

Mucosa of human detrusor impairs contraction and β‐adrenoceptor‐mediated relaxation

To elucidate the impact of the mucosa on detrusor muscle function by investigating force of contraction under various stimulatory conditions and during subsequent relaxation using catecholamines.

Catecholamines Relax Detrusor through 2-Adrenoceptors in Mouse and 3-Adrenoceptors in Man

(-)-Isoproterenol [4-[1-hydroxy-2-[(1-methylethyl)amino]ethyl]-1,2-benzene diol hydrochloride] relaxes murine detrusor through β-adrenoceptors (ARs); however, the β-AR subtypes involved are unknown. β2-ARs have been associated with caveolae, plasma-lemmal scaffolding domains that are absent in caveolin-1 (cav-1) knockout (KO) mice. Here, we studied detrusor responses in the absence and presence of β-AR subtype-selective antagonists in wild-type (WT) and cav-1 KO mice. To inquire whether the murine detrusor model is relevant to man, β-AR subtypes that mediate (-)-isoproterenol-evoked human detrusor relaxation were investigated. In WT mice, (-)-isoproterenol concentration-dependently relaxed the KCl (40 mM)-precontracted detrusor (-logEC50M = 8.04, Emax = 62%). The effects of (-)-isoproterenol were surmountably antagonized by the β2-AR-selective antagonist ICI 118,551 [(±)-1-[2,3-(dihydro-7-methyl-1H-inden-4-yl)oxy]-3-[(1-methylethyl)amino]-2-butanol] (pKB = 9.28) but not affected by the β1-AR-selective antagonist CGP 20712 [1-[2-((3-carbamoyl-4-hydroxy)phenoxy)ethylamino]-3-[4-(1-methyl-4-trifluoromethyl-2-imidazolyl)phenoxy]-2-propanol] and β3-AR-selective L-748,337 [(S)-M-[4-[2-[3-[3-[acetamidomethyl)phenoxy)-2-hydroxypropyl]-amino]-ethyl]-phenylbenzsulfonamide)], suggesting involvement of β2-AR only. The cav-1 KO detrusor displayed significant contractile dysfunction. (-)-Isoproterenol was less potent and efficient in relaxing detrusor from cav-1 KO (-logEC50M, 7.76; Emax = 44%), but ICI 118,551 caused similar antagonism (pKB = 9.15), suggesting that β2-AR function persisted in cav-1 KO. The β3-AR-selective antagonist L-748,337 in the presence of ICI 118,551 and CGP 20712 caused additional blockade of (-)-isoproterenol effects in cav-1 KO, consistent with a β3-AR involvement during relaxation and suppression of this effect in WT. (-)-Isoproterenol relaxed human detrusor muscle precontracted with carbachol (-logEC50M = 6.39, Emax = 52%). However, the effects of (-)-isoproterenol in human detrusor were not blocked by CGP 20712 or ICI 118,551 but antagonized by L-748,337 (pKB = 7.65). We conclude that murine detrusor relaxation occurs via β2-AR, and loss of caveolae does not perturb β2-AR function but unmasks an additional activation of β3-AR. In contrast, detrusor relaxation in man is mediated exclusively via β3-AR.