Video Dimension Analyzer Research Articles

Pregnancy-Induced Physiologic Adaptation of the Abdominal Aorta Is Associated with Changes in Gene Expression and Genomic Methylation

Background/Aims: Ten-eleven translocation 2 (Tet2), a DNA demethylase enzyme, has been identified as a master epigenetic regulator of vascular smooth muscle cell plasticity. We hypothesized that pregnancy will induce significant adaptive changes in aortic biomechanics that correlate with the Tet family gene expression. Methods: Abdominal aortas from pregnant and nonpregnant mice were dissected and cannulated. Intraluminal pressure was adjusted using a pressure-servo system while using a video dimension analyzer to measure the lumen diameter. Quantitative polymerase chain reaction and immunoblot was used to analyze the expression of Tet genes. Global genomic methylation was assessed with the luminometric methylation assay. Results: Compared to the nonpregnant (NP, 706 ± 8 µm) control group, the aortic luminal diameter was significantly increased in both E18.5 (836 ± 14 µm) and PP30 (889 ± 16 µm) mice. Distensibility was reduced in E18.5 (90 ± 4%) mice and returned to NP values (108 ± 2%) in PP30 (108 ± 3%) mice. Tet2 transcription decreased at the beginning of pregnancy and subsequently increased in late gestation, inversely corresponding to changes in global methylation. Conclusion: Physiologic changes in the aorta were accompanied by changes in gene expression and genomic methylation, suggesting an epigenetic component to maternal vascular remodeling during pregnancy.

207 The Vascular Effects of Resveratrol on Aged Mouse Femoral Arteries

<h3>Background</h3> In addition to its antioxidant properties, resveratrol (RV) is also known for its vasodilating effects. Although some of the vasodilating mechanisms of RV have been attributed to increases in NO, it has been previously demonstrated that in dietary-obese rats, in which endothelial function is impaired, RV is able to promote &gt;95% dilation. Given the above observations, we hypothesised that RV treatment restores endothelial function and flow mediated dilation of femoral arteries in old mice. <h3>Methods</h3> To establish vasodilator responses, C57BL/6 male mice (105 weeks of age) were humanely euthanised in accordance with the ‘Animals (Scientific Procedures) Act 1986’ and Institutional guidelines. Sections of the femoral artery were carefully dissected and mounted between two glass cannulae and fixed in place on a modified pressure myograph chamber. Arteries were initially pressurised to an intravascular (lumenal) pressure of 60 mmHg and maintained at that pressure using a pressure servo-control unit. Lumen and internal diameters of the vessel were constantly measured using a video dimension analyser. Constrictor responses (Phenylephrine (Phe) 10^–9–10^–3 M) and endothelial-dependent dilator responses to acetylcholine (Ach; 10^–9–10^–3 M) and flow (5–10 µL·min^–1) in pre-constricted vessels (Phe 10^–5 M) were assessed in femoral arteries incubated for 1 hr in either RV (30 µM) or physiological saline solution (PSS). The dilator responses were calculated as percentage of the pre-constrictor diameter, <h3>Results and conclusion</h3> RV treatment had no significant effect on Ach-dependent dilator responses. Whereas intraluminal flow of 8µL·min^–1 increased the diameter of the femoral arteries by up to 80% of the original diameter when arteries were pre-constricted and incubated in PSS, we observed no dilation after treatment with RV. Our data suggest that RV may have differential effects on Ach-dependent and flow mediated dilator responses in femoral arteries from aged mice.

Augmentation of Tendon Healing with Butyric Acid-Coated Suture: Biomechanical Evaluation in a Rabbit Model (SS-09)

IntroductionButyric acid (BA), in the appropriate concentration, has been shown to be angiogenic and to enhance transcriptional activity in tissue. These properties of BA have the potential to augment biologic healing in a repaired tendon. The purpose of this study is to evaluate this possibility through the comparison of biomechanical properties of healing tendons repaired with BA-coated suture versus control suture.MethodsA rabbit Achilles tendon model was used to evaluate the biomechanical strength of tendon healing at six weeks in twenty-three rabbits repaired either with BA-coated Ultrabraid (Smith & Nephew Endoscopy, Andover, MA) suture (n=14), or control Ultrabraid suture (n=9). Unilateral tendon defects were created in the middle bundle of the Achilles tendon of each rabbit, which were repaired equivalently with either BA-coated suture or control suture. The samples were harvested at six weeks post-repair and biomechanically tested using Video Dimension Analysis to determine sample stiffness.ResultsAt six weeks after repair, the tendon samples repaired with BA-coated suture had significantly increased stiffness relative to the tendons repaired with control suture. The Young's modulus was 84.9 ± 11.5 MPa in the BA group and 58.8 ± 15.2 MPa in the control group (p<0.05).ConclusionTendons repaired with BA-coated sutures demonstrated improved biomechanical properties at six weeks in a rabbit model. These findings are encouraging as a relatively simple alteration of suture material may augment early tendon healing to create a stronger repair construct during this time. IntroductionButyric acid (BA), in the appropriate concentration, has been shown to be angiogenic and to enhance transcriptional activity in tissue. These properties of BA have the potential to augment biologic healing in a repaired tendon. The purpose of this study is to evaluate this possibility through the comparison of biomechanical properties of healing tendons repaired with BA-coated suture versus control suture. Butyric acid (BA), in the appropriate concentration, has been shown to be angiogenic and to enhance transcriptional activity in tissue. These properties of BA have the potential to augment biologic healing in a repaired tendon. The purpose of this study is to evaluate this possibility through the comparison of biomechanical properties of healing tendons repaired with BA-coated suture versus control suture. MethodsA rabbit Achilles tendon model was used to evaluate the biomechanical strength of tendon healing at six weeks in twenty-three rabbits repaired either with BA-coated Ultrabraid (Smith & Nephew Endoscopy, Andover, MA) suture (n=14), or control Ultrabraid suture (n=9). Unilateral tendon defects were created in the middle bundle of the Achilles tendon of each rabbit, which were repaired equivalently with either BA-coated suture or control suture. The samples were harvested at six weeks post-repair and biomechanically tested using Video Dimension Analysis to determine sample stiffness. A rabbit Achilles tendon model was used to evaluate the biomechanical strength of tendon healing at six weeks in twenty-three rabbits repaired either with BA-coated Ultrabraid (Smith & Nephew Endoscopy, Andover, MA) suture (n=14), or control Ultrabraid suture (n=9). Unilateral tendon defects were created in the middle bundle of the Achilles tendon of each rabbit, which were repaired equivalently with either BA-coated suture or control suture. The samples were harvested at six weeks post-repair and biomechanically tested using Video Dimension Analysis to determine sample stiffness. ResultsAt six weeks after repair, the tendon samples repaired with BA-coated suture had significantly increased stiffness relative to the tendons repaired with control suture. The Young's modulus was 84.9 ± 11.5 MPa in the BA group and 58.8 ± 15.2 MPa in the control group (p<0.05). At six weeks after repair, the tendon samples repaired with BA-coated suture had significantly increased stiffness relative to the tendons repaired with control suture. The Young's modulus was 84.9 ± 11.5 MPa in the BA group and 58.8 ± 15.2 MPa in the control group (p<0.05). ConclusionTendons repaired with BA-coated sutures demonstrated improved biomechanical properties at six weeks in a rabbit model. These findings are encouraging as a relatively simple alteration of suture material may augment early tendon healing to create a stronger repair construct during this time. Tendons repaired with BA-coated sutures demonstrated improved biomechanical properties at six weeks in a rabbit model. These findings are encouraging as a relatively simple alteration of suture material may augment early tendon healing to create a stronger repair construct during this time.

Heparin-binding EGF-like growth factor is a potent dilator of terminal mesenteric arterioles

Objective We have previously shown that heparin-binding EGF-like growth factor (HB-EGF) protects the intestines from multiple forms of injury via direct cytoprotective effects on the intestinal mucosa. In this study, we examined the effects of HB-EGF on the hemodynamics of intestinal arterioles, the major resistance vessels that regulate blood flow to the intestines, as an additional mechanism of HB-EGF-mediated intestinal protection. Methods The hemodynamic effects of HB-EGF in rodent terminal mesenteric arterioles and human submucosal arterioles were examined ex vivo using a video dimension analyzer. Cultured human intestinal microvascular endothelial cells (HIMEC) were used to elucidate the mechanisms of HB-EGF-induced vasodilation. Results HB-EGF significantly increased vessel diameter under conditions of increasing intraluminal pressure and increased flow rate. These HB-EGF-mediated vasodilatory effects were observed in terminal mesenteric arterioles from adult rats and 3 day old rat pups. These effects were confirmed in submucosal arterioles from human intestine. Furthermore, HB-EGF significantly reduced endothelin-1-induced mesenteric arteriolar vasoconstriction. The vasodilatory effects of HB-EGF were blocked by ET B receptor antagonism in adult rat arterioles, and also by nitric oxide synthase inhibition in rat pup and human infant arterioles. In HIMEC, HB-EGF significantly increased endothelin B (ET B) receptor protein expression and provoked intracellular calcium mobilization. Conclusions HB-EGF is a potent vasodilator of the intestinal microvasculature, further supporting its use in diseases manifested by decreased intestinal blood flow, including necrotizing enterocolitis.

Mechanical behavior of arteries under inflation and extension

In order to understand the blood vessel of artery responding to medical treatments, there is a need to quantify the biomechanical behavior of the vessel. Using video dimension analyser, mechanical pressure-diameter relationships of passive carotid artery with and without balloon inflated at various axial loads were measured. Inversion test was also analytically modeled to address the heterogeneity of artery due to constituents. The pressure-diameter curves and stress distribution across the arterial wall were calculated. The simulation showed that the inversion tests, in comparison to other mechanical tests, provided additional information to develop the strain energy function describing arterial behavior

Radial metrology application to whole-body measurement on hyperelastic tubular samples

This paper focuses on a one-camera/one-shot procedure able to get the whole deformation map of hyperelastic tubular samples. A challenging application of this approach is the investigation of the highly anisotropic and inhomogeneous arterial tissue mechanical response during inflation/extension tests. To address this issue, full field optical methods based on digital correlation (DIC), fringe projection (FP) and stereo-photogrammetry (SP) have been already proposed in literature to overcome limitations of the most widely adopted 2-D video dimension analyzer (VDA) systems. In this paper, the feasibility of a very straightforward full-field procedure that uses radial metrology concepts has been studied. The rationale behind the proposed method relies on the relation existing between image deformation of a world point reflected by a 45° concave conical mirror and the relative position of this point with respect to the specular surface. Under certain assumptions reasonably true for the application of interest, by using simple relationships, it is possible to retrieve the position of markers applied onto the sample surface with great precision. This procedure has several advantages such as the retrieval of the whole 360° surface map in one shot, the ease of application, the use of one single camera, the real-time measurement capability. Conversely, the proposed approach is suitable only for geometries with smooth transversal sections, needs sample preparation and its spatial resolution is limited by the sparsity of the surface control points. The paper describes first the theoretical basis of the procedure; then results of experimental tests on calibration samples and latex tubular specimens are presented and discussed. Further set-up improvements will allow the present procedure to be implemented for in-vitro inflation/extension tests on vascular segments.

Vasoactivity of epoxyeicosatrienoic acids (EETs) in murine renal arcuate arteries

EETs mediate adenosine-induced vasodilation in rat preglomerular microvessels (PGMV); 11, 12-EET is the candidate mediator of PGMV dilation. We report on the response of the mouse arcuate artery to EETs. Male C57BL/6J mice (age 9–12 weeks) were anaesthetized and their kidneys excised and hemisected. Arcuate arteries (85 to 100 μm) were microdissected (4°C), cannulated and pressurized to 80 mmHg. Vessels were superfused with oxygenated Krebs’ solution (at 37°C) containing L-NAME (200 μM) and indomethacin (10μM). Vessels were then preconstricted with phenylephrine (20nM) producing an internal diameter (i.d.) of 46±6 μm. EETs were added to the superfusate and the i.d. was monitored by a video dimension analyzer. The threshold concentration of 11, 12-EET that dilated the arcuate artery (Δ13±2 μm i.d.) was 10−8M. Maximum dilation, Δ29±5 μm i.d., was achieved by 10−6M 11, 12-EET. The 5,6-, 8,9- and 14,15-EETs were equipotent and were ca 50% less active than 11, 12-EET. The ability of 11, 12-EET to increase the i.d. of the arcuate artery was compromised after incubation of arteries with an inhibitor of adenylyl cyclase, SQ 22536, for 15 min. 11, 12-EET increased i.d. by 29±5 μm before and by 7±2 μm (baseline 34±9 μM) after SQ 22536. Thus, the arcuate artery of the mouse resembles that of the rat in terms of responsiveness to EETs. It will serve as a model to delineate the regulation of EET-dependent renal vascular mechanisms.

Isometric pressure myography enables measurement of maximal vascular vasoconstriction and authentic EC 50

Both isometric wire and isobaric pressure myography have limitations. Wire myography distorts the vessel, may damage the endothelium, and limits intraluminal perfusion. On the other hand, occlusion of the vessel lumen may preclude measurement of maximal responses in the pressure myograph. We developed a system whereby the response of pressurized vessels could be measured under isometric conditions. We used the video dimension analyzer and servo controller of a classical pressure myograph (Living Systems Instrumentation) to maintain mesenteric and tail artery segments at a constant diameter by changing the intraluminal pressure. Concentration-response curves for phenylephrine showed a classical sigmoidal form (pressure vs concentration), from which we were able to determine maximal response and EC50. The EC50 for the mesenteric arteries was significantly lower by isometric pressure myography (4.5±0.7x10−7M, n=5) than by wire myography (Kent Scientific) (20.4±2.0x10−6M, n=7). On the isometric pressure myograph system, there was a rightward shift of the curve in the presence of prazosin. We propose that isometric pressure myography, which may be readily achieved with minimal modification of existing equipment, offers significant advantages over classical isobaric pressure myography. It allows for estimation of a true maximal constrictive response, and calculation of an authentic EC50. CIHR

Stimulatory Effect of Erythropoietin on Endothelial Repair After Vascular Injury

Erythropoietin (EPO) fosters tissue oxygenation by stimulating erythropoiesis. More recently, EPO is recognized as a tissue protective cytokine. In this study, we tested the hypothesis that EPO promotes recovery of injured vascular endothelium. A murine model of wire-induced injury of common carotid artery was used to examine the therapeutic effect of EPO. Recombinant human EPO (1000 U/kg, s.c., biweekly) was administered for 2 weeks after which vascular reactivity of isolated carotid arteries was studied in vitro under pressurized condition using a video dimension analyzer. Changes in vascular structure were histologically assessed. Injured arteries exhibited impairment in endothelium-dependent relaxations to acetylcholine and increased thickness of the medial cross sectional area (P<0.05; n=5–7). EPO normalized the vasorelaxant response of injured arteries to acetylcholine and prevented the increase in medial cross sectional area (30±1 x 10-3 mm2; P<0.05 vs. without EPO: 37±3 x 10-3 mm2; n=6). The number of circulating endothelial progenitor cells (EPCs), defined as CD34+/Flk-1+ cells in the peripheral blood, was significantly reduced 2 weeks after vascular injury (0.016±0.005%; P<0.05 vs. non-injured control: 0.09±0.01%), but this was entirely restored by EPO (0.212±0.08%; P<0.05; n=5). Our results indicate that EPO accelerates recovery of the injured carotid artery endothelium, and such salutary effects of EPO may reflect augmentation in circulating EPCs induced by EPO.

Whole 3D shape reconstruction of vascular segments under pressure via fringe projection techniques

Understanding and modelling vascular wall mechanics is a primary issue in the study of circulatory diseases. Although theoretical and numerical studies on arteries compliance are continuously increasing, relatively little work has been documented on the use of non-invasive imaging techniques for monitoring 3D vascular wall deformations. Usually, 2D video dimension analyzer (VDA) systems recover diameter and length variations during inflation/extension tests by tracking position changes of few markers put on the blood vessel surface. Then, strain determination relies on the assumption of axisymmetric deformations. However, more rigorous evaluations of whole wall deformation map are required for properly modelling the highly anisotropic and inhomogeneous vascular tissue mechanical response. This paper describes the development and application of a fringe projection (FP)-based procedure for the 360° 3D shape reconstruction of tubular samples subjected to internal pressure. A specially designed fixture for mounting and inflating the tubular segment allows specimen rotation about its axis. Movement is controlled by a high-precision rotational stage. This yields accurate positioning of the surface to be investigated with respect to the viewing direction. Data point clouds obtained from multiple recorded images are then processed and merged in a CAD environment, thus providing the whole shape of the sample with very high spatial resolution. The entire procedure has successfully been applied to latex specimens and porcine vascular segments. Further improvements will make the present procedure suitable for in vitro tests under more closely reproduced physiological conditions.

Effect of Caffeine on Cerebral Blood Flow Response to Somatosensory Stimulation

Despite caffeine's wide consumption and well-documented psychoactive effects, little is known regarding the effects of caffeine on neurovascular coupling. In the present study, we evaluated the effects of caffeine, an adenosine receptor antagonist, on intracerebral arterioles in vitro and subsequently, on the pial circulation in vivo during cortical activation induced by contralateral sciatic nerve stimulation (SNS). In our in vitro studies, we utilized isolated intracerebral arterioles to determine the effects of caffeine (10 or 50 micromol/L) on adenosine-induced vasodilatation. At the lower concentration, caffeine was without effect, but at the higher concentration, caffeine produced significant attenuation. In our in vivo studies, we determined the cerebrospinal fluid (CSF) caffeine concentrations at 15, 30, and 60 mins after intravenous administration of 5, 10 and 40 mg/kg. At the latter two concentrations, CSF levels exceeded 10 micromol/L. We then evaluated the pial arteriolar response during cortical activation caused by contralateral SNS after administering caffeine intravenously (0, 5, 10, 20 30, and 40 mg/kg). The pial circulation was observed through a closed cranial window in chloralose-anesthetized Sprague-Dawley rats. The contralateral sciatic nerve was isolated, positioned on silver electrodes and stimulated for 20 secs (0.20 V, 0.5 ms, and 5 Hz). Arteriolar diameter was quantified using an automated video dimension analyzer. Contralateral SNS resulted in a 23.8% +/-3.9% increase in pial arteriolar diameter in the hindlimb sensory cortex under control conditions. Intravenous administration of caffeine at the lowest dose studied (5 mg/kg) had no effect on either resting arteriolar diameter or SNS-induced vasodilatation. However, at higher doses (10, 20, 30, and 40 mg/kg, intravenously), caffeine significantly (P < 0.05; n = 6) attenuated both resting diameter and cerebral blood flow (CBF) responses to somatosensory stimulation. Intravenous administration of theophylline (10, 20, and 40 mg/kg), another adenosine receptor antagonist, also significantly reduced SNS-induced vasodilatation in a dose-dependent manner. Hypercarbic vasodilatation was unaffected by either caffeine or theophylline. The results of the present study show that caffeine significantly reduces cerebrovascular responses to both adenosine and to somatosensory stimulation and supports a role of adenosine in the regulation of CBF during functional neuronal activity.

Effects of separate application of three growth factors (TGF-beta1, EGF, and PDGF-BB) on mechanical properties of the in situ frozen–thawed anterior cruciate ligament

Objective. To clarify effects of a separate application of TGF-beta1, EGF, and PDGF-BB on the material properties of the in situ frozen–thawed anterior cruciate ligament. Design. Twenty-eight rabbits were divided into four groups after undergoing the in situ freeze–thaw treatment in the right anterior cruciate ligament. In 3 of the 4 groups, 4 ng TGF-beta1, 20 ng EGF, and 4 μg PDGF-BB was applied to the frozen anterior cruciate ligament, respectively. In the remaining sham treatment group, only fibrin sealant as a vehicle was applied. Each animal was sacrificed at 12 weeks after surgery. Background. If the role of growth factors in ligament healing and remodeling is understood, better therapies can be designed for ligament trauma. Methods. The freeze–thaw treatment was performed three times using the originally developed cryo-probe. The cross-sectional area of the anterior cruciate ligament was measured by the optical non-contact method. After preconditioning, each specimen was stretched to failure. The ligament strain was determined with a video dimension analyzer. Results. The tensile strength and the tangent modulus of the anterior cruciate ligament in the TGF-beta1 group was significantly higher than in the sham group, but significantly lower than in the normal control group. There were no significant differences in the strength and the modulus between the EGF group, the PDGF-BB group, and the sham group. Conclusions. In this model, an application of 4 ng TGF-beta1 significantly inhibited some of the material deterioration that occurs in the in situ frozen–thawed anterior cruciate ligament, while an application of 20 ng EGF or 4 μg PDGF-BB did not significantly affect the deterioration. Relevance This information will be useful in the future to develop a new biological therapy for ligament reconstruction to prevent the graft deterioration after transplantation.

Mechanisms of aging-induced impairment of endothelium-dependent relaxation: role of tetrahydrobiopterin.

Oxidative stress has been implicated as an important mechanism of vascular endothelial dysfunction induced by aging. Previous studies suggested that tetrahydrobiopterin (BH4), an essential cofactor of endothelial NO synthase, could be a molecular target for oxidation. We tested the hypothesis that oxidative stress, in particular oxidation of BH4, may contribute to attenuation of endothelium-dependent relaxation in aged mice. Vasomotor function of isolated carotid arteries was studied using a video dimension analyzer. Vascular levels of BH4 and its oxidation products were measured via HPLC. In aged mice (age, 95 +/- 2 wk), endothelium-dependent relaxation to ACh (10(-5) to 10(-9) M) as well as endothelium-independent relaxation to the NO donor diethylammonium (Z)-1-(N,N-diethylamino)diazen-1-ium-1,2-diolate (DEA-NONOate, 10(-5) to 10(-9) M) were significantly reduced compared with relaxation detected in young mice (age, 23 +/- 0.5 wk). Incubation of aged mouse carotid arteries with the cell-permeable SOD mimetic Mn(III)tetra(4-benzoic acid)porphyrin chloride normalized relaxation to ACh and DEA-NONOate. Furthermore, production of superoxide anion in aorta and serum levels of amyloid P component, which is the murine analog of C-reactive protein, was increased in old mice. In aorta, neither the concentration of BH4 nor the ratio of reduced BH4 to the oxidation products were different between young and aged mice. Our results demonstrate that in mice, aging impairs relaxation mediated by NO most likely by increased formation of superoxide anion. Oxidation of BH4 does not appear to be an important mechanism underlying vasomotor dysfunction in aged mouse arteries.

Neurogenic vasodilation of dural blood vessels is not mediated by cholinergic transmission in the anaesthetised rat

Dural vessel dilation induced by activation of trigeminal sensory fibres may be responsible for some component of the migraine attack. The presence in some patients with migraine and cluster headache of clinical features, such as lacrimation, suggests cranial parasympathetic activation and poses the question as to whether neurogenic meningeal dilatation has a cholinergic component. Rats were prepared in order to record on-line the diameter of a middle meningeal artery branch through a closed cranial window using an intravital microscope coupled to a video dimension analyser. Acetylcholine (1 μg, intravenously, i.v.) was administered before and after muscarinic receptor inhibition ( n=5) with scopolamine (2 mg/kg, i.v.) or nicotinic receptor inhibition ( n=6) with mecamylamine (4 mg/kg, i.v.). Further vasodilation was induced by electrical stimulation of the cranial window surface before and after muscarinic receptor inhibition with i.v. scopolamine ( n=8). The mean dural vessel percentage increase caused by acetylcholine stimulation was significantly different before and after muscarinic receptor inhibition ( P=0.045). Moreover, there was no difference between the post receptor inhibition values and those obtained after vehicle infusion ( P=0.431). In contrast, no difference was detected in the effect of acetylcholine before and after nicotinic receptor inhibition ( P=0.688). In the second experiment, where the effect of muscarinic receptor inhibition on the neurogenic dilation model was assessed, no significant difference was demonstrated ( P=0.538). Cholinergic dilation of the rat dural arteries is mediated by muscarinic receptors, but this mechanism does not play a significant role in the rat dural vessel dilation induced by closed cranial window electrical stimulation.

Treatment with irbesartan counteracts the functional remodeling of ventricular myocytes from hypertensive rats.

Changes in electrophysiological (action potential prolongation, decrease in transient outward current I(to), occurrence of the hyperpolarization-activated current I(f)) and contractile properties develop in hypertrophied ventricular myocytes, likely implicated in the increased propensity to arrhythmias. Angiotensin II is a key signal for myocyte hypertrophy; the effect of 8-week treatment with irbesartan, a type 1 angiotensin II receptor (AT(1)) antagonist, on cardiac remodeling was tested. Sixteen-month-old hypertensive rats (SHRs) were treated with irbesartan (20 mg/kg/d) or saline for 8 weeks. At the end of treatment, systolic blood pressure and heart weight to body weight ratio were reduced in irbesartan-treated compared with nontreated SHRs. Electrical and contractile properties were measured in isolated ventricular myocytes, by patch-clamp or video-dimension analysis, respectively. Action potential duration was significantly shorter in irbesartan-treated than in nontreated SHRs (at -60 mV: 119 +/- 24 ms vs 187 +/- 20 ms); correspondingly, maximal I(to) density was larger in irbesartan-treated than in nontreated SHRs (25.4 +/- 2.8 pA/pF vs 18.5 +/- 1.5 pA/pF). Maximal specific conductance of I(f) was lower in irbesartan-treated vs nontreated SHRs (24.8 +/- 3.0 pS/pF vs 35.2 +/- 4.0 pS/pF). Finally, the relaxation rate of shortening in field-stimulated intact myocytes was significantly faster in irbesartan-treated than in nontreated SHRs (7.3 +/- 0.5/s vs 5.7 +/- 0.3/s). Thus, AT(1) blockade with irbesartan, at an oral daily dosage that gave a slight but significant reduction of systolic blood pressure, largely counteracts the development of myocyte hypertrophy and associated functional alterations.

Protective effect of chronic vitamin C treatment on endothelial function of apolipoprotein E-deficient mouse carotid artery.

Endothelium-dependent relaxations are impaired in carotid artery of apolipoprotein E-deficient (apoE-/-) mice. This impairment seems to be due to increased formation of superoxide anions and inactivation of endothelial nitric oxide (NO). In the present study, we tested hypothesis that chronic treatment with vitamin C may prevent endothelial dysfunction by increasing release of NO from endothelial cells. C57BL/6 and apoE-/- mice were treated for 26 weeks with Western-type fat diet with and without 1% vitamin C. Vasomotor function of isolated carotid arteries was studied by video dimension analyzer. Expression of endothelial NO synthase (eNOS) and platelet-endothelial cell adhesion molecule-1 (PECAM-1) protein were evaluated by Western blotting. Levels of cGMP and cAMP were measured by radioimmunoassay. In apoE-/- mice, vitamin C significantly augmented relaxations to acetylcholine (10-9-10-5 mol/l), but did not affect relaxations to NO donor diethylammonium-(Z)-1-(N,N-diethylamino) diazen-1-1,2-diolate (DEA-NONOate; 10-9-10-5 mol/l). In contrast, vitamin C reduced relaxations to acetylcholine and DEA-NONOate in C57BL/6 mice. Interestingly, vitamin C significantly increased basal cGMP levels in C57BL/6 mice but did not affect cGMP formation in apoE-/-. Vitamin C treatment did not affect expression of eNOS protein, whereas elevated expression of PECAM-1 protein in apoE-/- mice was returned to normal level. Our findings demonstrate that chronic treatment with vitamin C prevents endothelial dysfunction of carotid artery induced by hypercholesterolemia. This effect seems to be mediated by preservation of NO bioavailability in endothelial cells.

The ORL-1 (NOP 1) receptor ligand nociceptin/orphanin FQ (N/OFQ) inhibits neurogenic dural vasodilatation in the rat

The effects of the ORL-1 (NOP 1) receptor ligand nociceptin (N/OFQ) and the nociceptin antagonists [Nphe 1]N/OFQ-(1-13)-NH 2 (Nphe) and nocistatin (NST) on neurogenic dural vasodilatation (NDV) in the rat dura mater evoked by electrical stimulation of a closed cranial window were studied. The middle meningeal artery was visualised using intravital microscopy, and the vessel diameter analysed using a video dimension analyser. N/OFQ (1, 10, 100 nmol kg −1; i.v., n=10) significantly and dose-dependently suppressed NDV maximally by 65% ( P<0.01). Neither Nphe (100 nmol kg −1; n=5) nor NST (100 nmol kg −1; n=4) alone had an effect on NDV ( P>0.05). Baseline vessel diameter was not significantly affected by application of N/OFQ, NST or Nphe. Application of the selective N/OFQ antagonist Nphe (10, 100 nmol kg −1 i.v., n=8) dose-dependently and significantly ( P<0.01) reversed the inhibition of NDV induced by application of N/OFQ (10 nmol kg −1). NST (10, 100 nmol kg −1; n=7) failed to reverse the effects elicited by N/OFQ. Application of N/OFQ elicited a dose-dependent transient decrease in arterial blood pressure ( P<0.01). Nphe dose-dependently reversed the cardiovascular effects induced by application of N/OFQ (10 nmol kg −1) ( P<0.01),while NST did not alter the blood pressure reaction elicited by N/OFQ. The results show that N/OFQ inhibits NDV, an effect which is antagonised by Nphe, but not by NST. ORL-1 (NOP 1) receptors located on trigeminal sensory fibres may be involved in the regulation of dural vessel diameter and hence may play a role in migraine pathophysiology.

The role of histamine in dural vessel dilation

The pain of migraine is often throbbing suggesting an important role for the cranial blood vessels and their innervation by the trigeminal nerve. It is proposed that clinically effective anti-migraine compounds, such as 5-HT 1B/1D agonists, have actions that include inhibiting calcitonin gene-related peptide (CGRP) release from trigeminal nerves. Human studies suggest that histamine can induce migraine possibly by activating nitric oxide (NO) synthase to promote endogenous NO production. The present studies investigated the effect of histamine and its antagonists on the cranial blood vessels using intravital microscopy to assess directly the diameter of dural arteries in sodium pentobarbitone anaesthetised rats. Electrical stimulation of a closed cranial window produces, by local depolarisation of nerves, dural vessel dilation that is monitored continuously on-line using video-microscopy and a video dimension analyser. Histamine infusion caused immediate and reproducible dilation of meningeal blood vessels (103.5±6%; n=40) that could be blocked by H 1- (mepyramine) and H 2 (famotidine)-receptor antagonists ( P<0.05), as well as a nitric oxide synthase inhibitor ( N G-nitro- l-arginine methylester; P<0.05). Neurogenic dural vasodilation was not inhibited by H 2-receptor antagonists, but was significantly inhibited by a H 1-receptor antagonist at the high dose of 10 mg/kg. The present studies demonstrate that histamine is likely to activate NO synthase to promote NO production. There is also evidence that H 1-receptors may be present on trigeminal neurones as the H 1-receptor antagonist inhibited neurogenic vasodilation, albeit at a large dose.

Contribution of nonendothelial nitric oxide to altered rat uterine resistance artery serotonin reactivity during pregnancy

Objective: The aims of this study were to evaluate the effects of pregnancy on the serotonin reactivity of the mesometrial arteries and to assess the relative contribution of endothelial and smooth muscle cells to the overall effect. Study Design: Arteries were obtained from nonpregnant (n = 20) and late pregnant (n = 28) Sprague-Dawley rats and were cannulated in the chamber of an arteriograph. Changes in lumen diameter were measured by a video dimension analyzer. Statistical analysis used 2-way analysis of variance, followed by Bonferroni test. Results: Intact vessels from late pregnant versus nonpregnant animals were significantly less sensitive to the constrictor effects of serotonin at concentrations <100 nmol/L. A role for endothelial nitric oxide was indicated on the basis of the effects of denudation and Nω-nitro-L-arginine. Furthermore, the additional effect of Nω-nitro-L-arginine in denuded vessels from pregnant animals suggests that another, nonendothelial source of nitric oxide also contributes to the effect. Conclusion: Pregnancy significantly decreases uterine resistance artery sensitivity to serotonin. This difference in sensitivity is due to the enhanced production of nitric oxide by endothelial cells and, most likely, by vascular smooth muscle. (Am J Obstet Gynecol 2002;187:463-8.)

Biomechanical characteristics of the lateral ligament of the ankle joint

The purpose of this study was to investigate the biomechanical properties of the lateral ankle ligaments and to correlate these properties with the distribution of mechanoreceptors in the ligaments reported previously. Biomechanical tensile tests were performed by using a video dimension analyzer to monitor the distortion of the lateral ligament at the pole and center in the animals. The mean upward slope of the load-elongation curves of the pole group was 38.1+/-14.2, and that of the center group was 79.1+/-6.5. The pole of the ligaments tended to be elongated more than the center. These biomechanical results suggest that mechanoreceptors in the pole of the ligaments, compared with those in the center, may receive more mechanical stress and be effective in monitoring of tension applied to the ligament.