Panax Quinquefolium Saponins Attenuate Myocardial Dysfunction Induced by Chronic Ischemia.

BACKGROUND/AIMS: Previous studies in rat models of myocardial ischemia showed that Panax quinquefolium saponins (PQS) could attenuate ischemic/reperfusion injury, increase vessel density and improve cardiac function. In the current study, we examined whether PQS could attenuate myocardial dysfunction in a swine model of chronic myocardial ischemia (CMI). METHODS: CMI was established in Bama mini-pigs by placing amroid constrictor on the left anterior descending artery (LAD). Starting from 2 months after the surgery, pigs randomly received PQS (30 mg/kg/day), atorvastatin (1.5 mg/kg/day), or no drug for one month (n=6). A group of pigs receiving sham surgery was included as an additional control. Glucose utilization was assessed with positron emission tomography-computer tomography (PET-CT). Cardiac function was assessed with echocardiography. Myocyte size, nuclear density, and arteriolar density were examined in tissue section obtained from the ischemia area. Potential molecular targets of PQS were identified using proteomic analysis with isobaric tags for relative and absolute quantitation (iTARQ) and network pharmacology. RESULTS: In comparison to the sham controls, pigs implanted with ameroid constrictor had decreased ventricular wall motion, left ventricular ejection fraction (LVEF), and glucose utilization. PQS significantly increased cardiac function and glucose utilization. Arteriole density and myocyte nuclear density were increased. Myocyte diameter was decreased. PQS also attenuated the CMI-induced change of protein expression profile. The effects of atorvastatin were generally similar to that of PQS. However, PQS attenuated the reduction of left ventricular systolic WT induced by CMI more robustly than atorvastatin. CONCLUSION: The results from the current study supports the use of PQS in patients with coronary artery disease.

Biomonitoring the cardiorenal effects of Luehea divaricata Mart.: An ethnoguided approach.

ETHNOPHARMACOLOGICAL RELEVANCE: Luehea divaricata Mart. (Malvaceae) is an important medicinal species that is widely used as a diuretic in the Brazilian Pantanal region. An ethanolic supernatant that was obtained from an infusion of leaves of this species (ESLD) was recently shown to exert hypotensive and diuretic activity. Nevertheless, the secondary metabolites that are responsible for this activity and the molecular mechanisms of pharmacological action remain unknown. AIM: We performed a detailed study to identify possible active metabolites that are present in different ESLD fractions and investigated their effects on renal and peripheral arteriolar tone. We further evaluated their interrelations with sustained diuretic and hypotensive actions. MATERIALS AND METHODS: The ESLD was first obtained from L. divaricata leaves, and liquid-liquid fractionation was performed. The fractions were analyzed by liquid chromatography-mass spectrometry. An ethyl acetate fraction (AceFr), n-butanolic fraction (ButFr), and aqueous fraction (AqueFr) were then orally administered in male Wistar rats in a single dose or daily for 7 days. The doses were previously defined based on the yield that was obtained from each fraction. Hydrochlorothiazide was used as a positive control. Blood pressure, heart rate, urinary volume, pH, density, and urinary sodium, potassium, chloride, and calcium levels were measured. Serum levels of nitrite, thiobarbituric acid reactive species, nitrotyrosine, aldosterone, vasopressin, and plasma angiotensin converting enzyme activity were also measured. Finally, the direct effects of the ButFr on renal and mesenteric arteriolar tone and the role of nitric oxide and prostaglandins in the renal and hemodynamic effects were investigated. RESULTS: Of the fractions that were tested, only the ButFr exerted significant diuretic and saluretic effects. The AceFr and ButFr also had acute hypotensive effects, but only the ButFr maintained its response after 7 days of treatment. Prolonged treatment with the ButFr increased serum nitrite levels and significantly reduced oxidative and nitrosative markers of stress. Additionally, the ButFr caused a vasodilatory response in the renal and mesenteric arteriolar beds through the release of nitric oxide and prostaglandins. Finally, the diuretic and hypotensive effects of the ButFr were completely blocked by pretreatment with Nω-nitro-L-arginine methyl ester and indomethacin, thus demonstrating the direct involvement of nitric oxide and prostaglandins in these effects. CONCLUSION: The ButFr that was obtained from Luehea divaricata exerted sustained diuretic and hypotensive effects. These effects were apparently attributable to the release of nitric oxide and prostaglandins, which reduce renal and peripheral arteriolar tone and lead to an increase in the glomerular filtration rate and a reduction of global peripheral resistance. These findings suggest that the ButFr may be a potential complementary therapy for several conditions in which diuretic and hypotensive effects are required.

Ultra-Slow Single-Vessel BOLD and CBV-Based fMRI Spatiotemporal Dynamics and Their Correlation with Neuronal Intracellular Calcium Signals.

Functional MRI has been used to map brain activity and functional connectivity based on the strength and temporal coherence of neurovascular-coupled hemodynamic signals. Here, single-vessel fMRI reveals vessel-specific correlation patterns in both rodents and humans. In anesthetized rats, fluctuations in the vessel-specific fMRI signal are correlated with the intracellular calcium signal measured in neighboring neurons. Further, the blood-oxygen-level-dependent (BOLD) signal from individual venules and the cerebral-blood-volume signal from individual arterioles show correlations at ultra-slow (<0.1 Hz), anesthetic-modulated rhythms. These data support a model that links neuronal activity to intrinsic oscillations in the cerebral vasculature, with a spatial correlation length of ∼2 mm for arterioles. In complementary data from awake human subjects, the BOLD signal is spatially correlated among sulcus veins and specified intracortical veins of the visual cortex at similar ultra-slow rhythms. These data support the use of fMRI to resolve functional connectivity at the level of single vessels.

Rapid versus slow ascending vasodilatation: intercellular conduction versus flow-mediated signalling with tetanic versus rhythmic muscle contractions.

KEY POINTS: In response to exercise, vasodilatation ascends from downstream arterioles into upstream feed arteries (FAs). We hypothesized that the signalling events underlying ascending vasodilatation variy with the intensity and duration of skeletal muscle contraction. In the gluteus maximus muscle of C57BL/6 mice, brief tetanic contraction evoked rapid onset vasodilatation (ROV) (<1 s) throughout the resistance network. Selective damage to endothelium midway between FAs and primary arterioles eliminated ROV only in FAs. Blocking SKCa and IKCa channels attenuated ROV, implicating hyperpolarization as the underlying signal. During rhythmic twitch contractions, slow onset vasodilatation (10-15 s) in FAs remained intact following loss of ROV and was eliminated following nitric oxide synthase inhibition. Tetanic contraction initiates hyperpolarization that conducts along endothelium into FAs. Rhythmic twitch contractions stimulate FA endothelium to release nitric oxide in response to elevated shear stress secondary to metabolic dilatation of arterioles. Complementary endothelial signalling pathways for ascending vasodilatation ensure increased oxygen delivery to active skeletal muscle. ABSTRACT: In response to exercise, vasodilatation initiated within the microcirculation of skeletal muscle ascends the resistance network into upstream feed arteries (FAs) located external to the tissue. Ascending vasodilatation (AVD) is essential for reducing FA resistance that otherwise restricts blood flow into the microcirculation. In the present study, we tested the hypothesis that signalling events underlying AVD vary with the intensity and duration of muscle contraction. In the gluteus maximus muscle of anaesthetized male C57BL/6 mice (aged 3-4 months), brief tetanic contraction (100 Hz for 500 ms) evoked rapid onset vasodilatation (ROV) in FAs that peaked within 4 s. By contrast, during rhythmic twitch contractions (4 Hz), slow onset vasodilatation (SOV) of FAs began after ∼10 s and plateaued within 30 s. Selectively damaging the endothelium with light-dye treatment midway between a FA and its primary arteriole eliminated ROV in the FA along with conducted vasodilatation of the FA initiated on the arteriole using ACh microiontophoresis. Superfusion of SKCa and IKCa channel blockers UCL 1684 + TRAM 34 attenuated ROV, implicating endothelial hyperpolarization as the underlying signal. Nevertheless, the SOV of FAs during rhythmic contractions persisted until inhibition of nitric oxide synthase with Nω -nitro-l-arginine methyl ester. Thus, ROV of FAs reflects hyperpolarization of downstream arterioles that conducts along the endothelium into proximal FAs. By contrast, SOV of FAs reflects the local production of nitric oxide by the endothelium in response to luminal shear stress, which increases secondary to arteriolar dilatation downstream. Thus, AVD ensures increased oxygen delivery to active muscle fibres by reducing upstream resistance via complementary signalling pathways that reflect the intensity and duration of muscle contraction.

Astrocyte contributions to flow/pressure-evoked parenchymal arteriole vasoconstriction.

Basal and activity-dependent cerebral blood flow changes are coordinated by the action of critical processes, including cerebral autoregulation, endothelial-mediated signaling, and neurovascular coupling. The goal of our study was to determine whether astrocytes contribute to the regulation of parenchymal arteriole (PA) tone in response to hemodynamic stimuli (pressure/flow). Cortical PA vascular responses and astrocytic Ca(2+) dynamics were measured using an in vitro rat/mouse brain slice model of perfused/pressurized PAs; studies were supplemented with in vivo astrocytic Ca(2+) imaging. In vitro, astrocytes responded to PA flow/pressure increases with an increase in intracellular Ca(2+). Astrocytic Ca(2+) responses were corroborated in vivo, where acute systemic phenylephrine-induced increases in blood pressure evoked a significant increase in astrocytic Ca(2+). In vitro, flow/pressure-evoked vasoconstriction was blunted when the astrocytic syncytium was loaded with BAPTA (chelating intracellular Ca(2+)) and enhanced when high Ca(2+) or ATP were introduced to the astrocytic syncytium. Bath application of either the TRPV4 channel blocker HC067047 or purinergic receptor antagonist suramin blunted flow/pressure-evoked vasoconstriction, whereas K(+) and 20-HETE signaling blockade showed no effect. Importantly, we found TRPV4 channel expression to be restricted to astrocytes and not the endothelium of PA. We present evidence for a novel role of astrocytes in PA flow/pressure-evoked vasoconstriction. Our data suggest that astrocytic TRPV4 channels are key molecular sensors of hemodynamic stimuli and that a purinergic, glial-derived signal contributes to flow/pressure-induced adjustments in PA tone. Together our results support bidirectional signaling within the neurovascular unit and astrocytes as key modulators of PA tone.

Effects of physical stimulation of spontaneous arteriolar vasomotion in patients of various ages undergoing rehabilitation.

On two samples of rehabilitation patients of different age groups (approx. 38 years and approx. 51 years), via a placebo-controlled study series using representative features of microcirculation, the complementary therapeutic success of additional treatment complementing the biorhythmically defined physical vasomotion stimulation was determined. The results showed that in older rehabilitation patients the amounts of characteristic microcirculatory changes were greater than in younger persons undergoing rehabilitation, but they would subside faster after termination of the additional treatment than in the younger group.

Advanced in vitro approach to study neurovascular coupling mechanisms in the brain microcirculation.

An understanding of the signalling events underlying neurovascular coupling mechanisms in the brain is a crucial step in the development of novel therapeutic approaches for the treatment of cerebrovascular-associated disorders. In this study we present an enhanced in vitro brain slice preparation from male Wistar rat cortical slices that incorporates haemodynamic variables (flow and pressure) into parenchymal arterioles resulting in the development of myogenic tone (28% from maximum dilatation). Moreover, we characterized flow-induced vascular responses, resulting in various degrees of vasoconstrictions and the response to 10 mM K(+) or astrocytic activation with the mGluR agonist, t-ACPD (100 µM), resulting in vasodilatations of 33.6±4.7% and 38.6±4.6%, respectively. Using fluorescence recovery, we determined perfusate velocity to calculate diameter changes under different experimental pH conditions. Using this approach, we demonstrate no significant differences between diameter changes measured using videomicroscopy or predicted from the velocity values obtained using fluorescence recovery after photobleaching. The model is further validated by demonstrating our ability to cannulate arterioles in two brain regions (cortex and supraoptic nucleus of the hypothalamus). Altogether, we believe this is the first study demonstrating successful cannulation and perfusion of parenchymal arterioles while monitoring/estimating luminal diameter and pressure under conditions where flow rates are controlled.

Effects of hot and cold foods on signals of heart rate variability and nail fold microcirculation of healthy young humans: a pilot study.

In traditional Chinese medicine, hot- and cold-attribute of food ingredients are a major part of dietary therapy. The aim of this study was to establish a suitable scientific methodology to define the attributes of food ingredients by investigating the relationship between food attributes and the physiological signals produced in healthy young subjects with different constitutions. Thirty subjects were grouped into hot and cold constitutions by Chinese medical doctors. Every subject took water, aged ginger tea and coconut water, which are well recognized as having neutral-, hot- and cold-attribute, respectively, on different visits. The different physiological signals induced by the samples were observed using skin and axillary temperature sensors, a heart rate variability analyzer and a laser Doppler anemometer. We found that the capillary red blood cell (RBC) velocity in nail fold microcirculation (NFM) of the subjects with hot constitution accelerated significantly after taking the hot-attribute aged ginger tea, which might be the result of elevated vagal activity leading to arteriole dilation in these subjects. In contrast, in subjects with cold constitution, capillary RBC velocity decelerated significantly and skin temperature decreased markedly after taking the cold-attribute coconut water, which might have been induced by sympathetic nerve activation causing the arteriole to be constricted. Accordingly, the use of capillary RBC velocity of NFM measured by laser Doppler anemometer may be a promising way to classify attributes of food ingredients commonly used in Chinese medicine dietary therapy in accordance with different personal constitutions.

Decreased arteriolar tetrahydrobiopterin is linked to superoxide generation from nitric oxide synthase in mice fed high salt.

OBJECTIVE: Impaired endothelium-dependent arteriolar dilation in mice fed high salt (HS) is due to local oxidation of nitric oxide (NO) by superoxide anion (O(2) (-)). We explored the possibility that "uncoupled" endothelial nitric oxide synthase (eNOS) is the source of this O(2) (-). METHODS: Levels of L-arginine (L-Arg), tetrahydrobiopterin (BH(4)), and O(2) (-) (hydroethidine oxidation) were measured in spinotrapezius muscle arterioles of mice fed normal salt (0.45%, NS) or (4%, HS) diets for 4 weeks, with or without dietary L-Arg supplementation. The contribution of NO to endothelium-dependent dilation was determined from the effect of N(omega)-nitro-L-arginine methyl ester (L-NAME) on responses to acetylcholine (ACh). RESULTS: Arterioles in HS mice had lower [BH(4)] and higher O(2) (-) levels than those in NS mice. ACh further increased arteriolar O(2) (-) in HS mice only. L-Arg supplementation prevented the reduction in [BH(4)] in arterioles of HS mice, and O(2) (-) was not elevated in these vessels. Compared to NS mice, arteriolar ACh responses were diminished and insensitive to L-NAME in HS mice, but not in HS mice supplemented with L-Arg. CONCLUSIONS: These findings suggest that eNOS uncoupling due to low [BH(4)] is responsible for O(2) (-) generation and reduced NO-dependent dilation in arterioles of mice fed a HS diet.

Increased NO bioavailability in aging male rats by genistein and exercise training: using 4, 5-diaminofluorescein diacetate.

BACKGROUND: Several kinds of anti-oxidants have drawn a lot of intention for their benefits on vascular protection. In addition, it has been demonstrated that exercise training could improve endothelial function by up-regulating endothelial nitric oxide synthase (eNOS) protein. Therefore, the present study aims to investigate the effects of genistein, a potent phyto-antioxidant, and exercise training on age-induced endothelial dysfunction in relation to NO bioavailability using in situ NO-sensitive fluorescent dye detection. METHODS: Male Wistar rats (20-22-month old) were divided into four groups: aged rats treated with corn oil, (Aged+Veh, n = 5), aged rats treated with genistein (Aged+Gen, n = 5, (0.25 mg/kg BW/day, s.c.)), aged rats with and without exercise training (Aged+Ex, n = 5, swimming 40 min/day, 5 days/week for 8 weeks) (Aged+Without-Ex, n = 5). Cremaster arterioles (15-35 micrometer) were visualized by fluorescein isothiocyanate labeled dextran (5 microgram/ml). The vascular response to acetylcholine (Ach; 10(-5)M, 5 ml/5 min) was accessed after 1-min norepinephrine preconstriction (10 micro molar). To determine NO bioavailability, the Krebs-Ringer buffer with 4, 5-diaminofluorescein-diacetate (3 micro molar DAF-2DA), and 10 micro- molar Ach saturated with 95%N2 and 5%CO2 were used. Changes of DAF-2T-intensities along the cremaster arterioles were analyzed by the Image Pro-Plus Software (Media Cybernatics, Inc, USA). Liver malondialdehyde (MDA) level was measured by thiobarbituric acid reaction and used as an indicator for oxidative stress. RESULTS: The results showed that means arterial blood pressure for both Aged+Gen and Aged+Ex groups were significantly reduced when compared to the Aged groups, Aged+Veh and Aged+Without-Ex (P < 0.05). Among the treated groups, Ach-induced vasodilatation were significantly increased (P < 0.05) and was associated with increased NO-associated fluorescent intensities (P < 0.05). On the other hand, MDA levels were significantly reduced (P < 0.05) when Aged+Veh was compared to Aged+Without-Ex. CONCLUSION: These findings showed that genistein and exercise training could improve age-induced endothelial dysfunction and is related to the increased NO bioavailability.

Secretory phospholipase A2 inhibitor PX-18 preserves microvascular reactivity after cerebral ischemia in piglets.

Cerebral ischemia/reperfusion (I/R) results in cellular energy failure and dysfunction of the neurovascular unit that contribute to subsequent neuronal cell death in the neonate. PX-18 is a putative neuroprotective inhibitor of secretory phospholipase A(2) (sPLA(2)) but its in vivo testing has been limited by its poor solubility. Our purpose was to assess whether PX-18 preserved neuronal-vascular reactivity to I/R-sensitive endothelium-dependent (hypercapnia, bradykinin) and/or neuron-dependent (N-methyl-D-aspartate; NMDA) stimuli. To make the drug available for in vivo studies, PX-18 was formulated as a 3% nanosuspension applying high pressure homogenization. Newborn piglets (1-day old, n=40) were anesthetized and ventilated, and cerebrovascular reactivity to the above stimuli was determined by measuring changes in pial arteriolar diameters using the closed cranial window/intravital videomicroscopy technique. Intravenous infusion of PX-18 nanosuspension (6 mg/kg, 20 min) did not affect baseline arteriolar diameters, or hypercapnia-, bradykinin-, or NMDA-induced pial arteriolar vasodilation under normoxic conditions. Global cerebral ischemia (10 min) followed by 1 h of reperfusion significantly attenuated hypercapnia-, bradykinin-, and NMDA-induced vasodilation in untreated or vehicle-treated controls. However, PX-18 resulted in nearly full preservation of cerebrovascular reactivity to all these stimuli. In conclusion, inhibition of sPLA(2) by PX-18 improves neurovascular function both at the neuronal and the microvascular level following I/R. This effect of PX-18 likely contributes to its neuroprotective effect.

H-Wave induces arteriolar vasodilation in rat striated muscle via nitric oxide-mediated mechanisms.

H-Wave electrical device stimulation (HWDS) is used clinically to expedite recovery from soft tissue injuries. We hypothesized that HWDS induces arteriolar dilation, a mechanism involved in the healing process. Acute effects of HWDS on striated muscle arteriolar diameters were studied. Arteriolar diameters were measured in the cremaster muscle of 57 male anesthetized rats using intravital microscopy before and after HWDS or sham stimulation (SS) at 1 or 2 Hz for periods of 30-60 min. In a separate cohort, the role of nitric oxide (NO) in the response to HWDS was assessed by blocking NO synthase using topical L-NAME at 10(-5) M. Maximal arteriolar responses to stimulation were compared to prestimulation diameters. HWDS both at 1 and 2 Hz resulted in significant arteriolar vasodilation (p < 0.05). The arterioles in SS animals demonstrated no changes in diameter. Similarly, microvascular diameters did not change with HWDS following blockade of NO production. Because of Poiseuille's Law, the significant arteriolar dilation induced by HWDS would translate into increases in blood flow of 26-62%. In addition, lack of arteriolar dilation following HWDS with blockade of NO production suggests that NO plays a role in the microvascular response to HWDS. These studies suggest that arteriolar vasodilation accompanying HWDS may result in increased perfusion, contributing to the observed therapeutic effects of HWDS.

Microcirculatory responses to acupuncture stimulation and phototherapy.

BACKGROUND: Acupuncture stimulation and phototherapy have been reported to have analgesic effects and improve the microcirculation. However, few studies have directly examined changes in peripheral blood vessels, either quantitatively or objectively. We assessed the responses of arteriolar blood flow to acupuncture stimulation and phototherapy under direct vision to examine the effects of these treatments. METHODS: We used 40 rabbits with a rabbit ear chamber attached to the auricle. The rabbit ear chamber was fixed to the auricle under a dissecting microscope. Arterioles were selected and observed with the use of a microscope video camera. Pentobarbital was injected IV. The trachea was intubated and spontaneous respiration was maintained. Rabbits were randomly assigned to receive acupuncture stimulation (acupuncture group, n = 10), near-infrared lamp irradiation (lamp group, n = 10), near-infrared low-powered laser irradiation (laser group, n = 10), or no irradiation (control group, n = 10). In the acupuncture group, an acupuncture needle was placed in the auricle for 20 min. The lamp group repeatedly received 1 s of near infrared irradiation (1540 mW) followed by 4 s of treatment cessation. The laser group continuously received 60 mW of laser irradiation. In the lamp and laser groups, the auricle (same site as that of the acupuncture needles in the acupuncture group) was irradiated for 10 min with a contact probe. Arteriolar diameter and blood flow velocity were measured at baseline and for 60 min after acupuncture or irradiation treatment. Blood flow rate was calculated by multiplying the blood flow velocity by the cross-sectional area of the vessels. RESULTS: Arteriolar diameter significantly increased to 131% +/- 14% in the acupuncture group (P < 0.005), 129% +/- 19% in the lamp group (P < 0.005), and 128% +/- 11% in the laser group (P < 0.005) when compared with the pretreatment value (100%). Maximum values were reached 20 min after the end of the acupuncture stimulation, and 10 min after the end of lamp and laser irradiation. The three groups showed significant increases in arteriolar diameter when compared with the control group (P < 0.005). Blood flow velocity and blood flow rate showed similar trends to arteriolar diameter. Treatment effect persisted for 40-50 min after the end of stimulation and irradiation. CONCLUSIONS: Acupuncture stimulation and phototherapy were directly confirmed to increase the diameter and blood flow velocity of the peripheral arterioles. Acupuncture stimulation and phototherapy, associated with minimal systemic and local side effects, can enhance the microcirculation and may be a useful supportive treatment for diseases caused by poor peripheral blood flow.

Propagated endothelial Ca2+ waves and arteriolar dilation in vivo: measurements in Cx40BAC GCaMP2 transgenic mice.

To study endothelial cell (EC)- specific Ca(2+) signaling in vivo we engineered transgenic mice in which the Ca(2+) sensor GCaMP2 is placed under control of endogenous connexin40 (Cx40) transcription regulatory elements within a bacterial artificial chromosome (BAC), resulting in high sensor expression in arterial ECs, atrial myocytes, and cardiac Purkinje fibers. High signal/noise Ca(2+) signals were obtained in Cx40(BAC)-GCaMP2 mice within the ventricular Purkinje cell network in vitro and in ECs of cremaster muscle arterioles in vivo. Microiontophoresis of acetylcholine (ACh) onto arterioles triggered a transient increase in EC Ca(2+) fluorescence that propagated along the arteriole with an initial velocity of approximately 116 microm/s (n=28) and decayed over distances up to 974 microm. The local rise in EC Ca(2+) was followed (delay, 830+/-60 ms; n=8) by vasodilation that conducted rapidly (mm/s), bidirectionally, and into branches for distances exceeding 1 mm. At intermediate distances (300 to 600 microm), rapidly-conducted vasodilation occurred without changing EC Ca(2+), and additional dilation occurred after arrival of a Ca(2+) wave. In contrast, focal delivery of sodium nitroprusside evoked similar local dilations without Ca(2+) signaling or conduction. We conclude that in vivo responses to ACh in arterioles consists of 2 phases: (1) a rapidly-conducted vasodilation initiated by a local rise in EC Ca(2+) but independent of EC Ca(2+) signaling at remote sites; and (2) a slower complementary dilation associated with a Ca(2+) wave that propagates along the endothelium.

Effects of thermal stimulation, applied to the hindpaw via a hot water bath, upon ovarian blood flow in anesthetized nonpregnant rats.

The effects of thermal stimulation, applied to the hindpaw via a hot bath set to either 40 degrees C (non-noxious) or 49 degrees C (noxious), upon ovarian blood flow were examined in nonpregnant anesthetized rats. Ovarian blood flow was measured using a laser Doppler flowmeter. Blood pressure was markedly increased following 49 degrees C stimulation. Ovarian blood flow, however, showed no obvious change during stimulation, although a small increase was observed after stimulation. Ovarian blood flow and blood pressure responses to 49 degrees C stimulation were abolished after hindlimb somatic nerves proximal to the stimuli were cut. Heat stimulation (49 degrees C) resulted in remarkable increases in both ovarian blood flow and blood pressure in rats in which the sympathetic nerves supplying the ovary were cut but the hindlimb somatic nerves remained intact. The efferent activity of the ovarian plexus nerve was increased during stimulation at 49 degrees C. Stimulation at 40 degrees C had no effect upon ovarian blood flow, blood pressure or ovarian plexus nerve activity. Electrical stimulation of the distal part of the severed ovarian plexus nerve resulted in a decrease in both the diameter of ovarian arterioles, observed using a digital video microscope, and ovarian blood flow.The present results demonstrate that noxious heat, but not non-noxious warm, stimulation of the hindpaw skin in anesthetized rats influences ovarian blood flow in a manner that is attributed to reflex responses in ovarian sympathetic nerve activity and blood pressure.

Chronic static magnetic field exposure alters microvessel enlargement resulting from surgical intervention.

Magnetic field therapy has recently become a widely used complementary/alternative medicine for the treatment of vascular, as well as other musculoskeletal pathologies, including soft tissue injuries. Recent studies in our laboratory and others have suggested that acute static magnetic field (SMF) exposure can have a modulatory influence on the microvasculature, acting to normalize vascular function; however, the effect of chronic SMF exposure has not been investigated. This study aimed to measure, for the first time, the adaptive microvascular response to a chronic 7-day continuous magnetic field exposure. Murine dorsal skinfold chambers were applied on day 0, and neodymium static magnets (or size and weight-matched shams) were affixed to the chambers at day 0, where they remained until day 7. Separate analysis of arteriolar and venular diameters revealed that chronic SMF application significantly abrogated the luminal diameter expansion observed in sham-treated networks. Magnet-treated venular diameters were significantly reduced at day 4 and day 7 (34.3 and 54.4%, respectively) compared with sham-treated vessels. Arteriolar diameters were also significantly reduced by magnet treatment at day 7 (50%), but not significantly at day 4 (31.6%), although the same trend was evident. Venular functional length density was also significantly reduced (60%) by chronic field application. These results suggest that chronic SMF exposure can alter the adaptive microvascular remodeling response to mechanical injury, thus supporting the further study of chronic application of SMFs for the treatment of vascular pathologies involving the dysregulation of microvascular structure.

Role of 20-HETE in the pial arteriolar constrictor response to decreased hematocrit after exchange transfusion of cell-free polymeric hemoglobin.

The cerebrovascular response to decreases in hematocrit and viscosity depends on accompanying changes in arterial O2 content. This study examines whether 1) the arteriolar dilation seen after exchange transfusion with a 5% albumin solution can be reduced by the K(ATP) channel antagonist glibenclamide (known to inhibit hypoxic dilation), and 2) the arteriolar constriction seen after exchange transfusion with a cell-free hemoglobin polymer to improve O2-carrying capacity can be blocked by inhibitors of the synthesis or vasoconstrictor actions of 20-HETE. In anesthetized rats, decreasing hematocrit by one-third with albumin exchange transfusion dilated pial arterioles (14 +/- 2%; SD), whereas superfusion of the surface of the brain with 10 muM glibenclamide blocked this response (-10 +/- 7%). Exchange transfusion with polymeric hemoglobin decreased the diameter of pial arterioles by 20 +/- 3% without altering arterial pressure. This constrictor response was attenuated by superfusing the surface of the brain with a 20-HETE antagonist, WIT-002 (10 microM; -5 +/- 1%), and was blocked by two chemically dissimilar selective inhibitors of the synthesis of 20-HETE, DDMS (50 microM; 0 +/- 4%) and HET-0016 (1 microM; +6 +/- 4%). The constrictor response to hemoglobin transfusion was not blocked by an inhibitor of nitric oxide (NO) synthase, and the inhibition of the constrictor response by DDMS was not altered by coadministration of the NO synthase inhibitor. We conclude 1) that activation of K(ATP) channels contributes to pial arteriolar dilation during anemia, whereas 2) constriction to polymeric hemoglobin transfusion at reduced hematocrit represents a regulatory response that limits increased O2 transport and that is mediated by increased formation of 20-HETE, rather than by NO scavenging.

Carbon monoxide from heme oxygenase-2 Is a tonic regulator against NO-dependent vasodilatation in the adult rat cerebral microcirculation.

Although the brain generates NO and carbon monoxide (CO), it is unknown how these gases and their enzyme systems interact with each other to regulate cerebrovascular function. We examined whether CO produced by heme oxygenase (HO) modulates generation and action of constitutive NO in the rat pial microcirculation. Immunohistochemical analyses indicated that HO-2 occurred in neurons and arachnoid trabecular cells, where NO synthase 1 (NOS1) was detectable, and also in vascular endothelium-expressing NOS3, suggesting colocalization of CO- and NO-generating sites. Intravital microscopy using a closed cranial window preparation revealed that blockade of the HO activity by zinc protoporphyrin IX significantly dilates arterioles. This vasodilatation depended on local NOS activities and was abolished by CO supplementation, suggesting that the gas derived from HO-2 tonically regulates NO-mediated vasodilatory response. Bioimaging of NO by laser-confocal microfluorography of diaminofluorescein indicated detectable amounts of NO at the microvascular wall, the subdural mesothelial cells, and arachnoid trabecular cells, which express NOS in and around the pial microvasculature. On CO inhibition by the HO inhibitor, regional NO formation was augmented in these cells. Such a pattern of accelerated NO formation depended on NOS activities and was again attenuated by the local CO supplementation. Studies using cultured porcine aortic endothelial cells suggested that the inhibitory action of CO on NOS could result from the photo-reversible gas binding to the prosthetic heme. Collectively, CO derived from HO-2 appears to serve as a tonic vasoregulator antagonizing NO-mediated vasodilatation in the rat cerebral microcirculation.

Upregulation of vascular arginase in hypertension decreases nitric oxide-mediated dilation of coronary arterioles.

One characteristic of hypertension is a decreased endothelium-dependent nitric oxide (NO)-mediated vasodilation; however, the underlying mechanism is complex. In endothelial cells (ECs), L-arginine is the substrate for both NO synthase (NOS) and arginase. Because arginase has recently been shown to modulate NO-mediated dilation of coronary arterioles by reducing l-arginine availability, we hypothesized that upregulation of vascular arginase in hypertension contributes to decreased NO-mediated vasodilation. To test this hypothesis, hypertension (mean arterial blood pressure >150 mm Hg) was maintained for 8 weeks in pigs by aortic coarctation. Coronary arterioles from normotensive (NT) and hypertensive (HT) pigs were isolated and pressurized for in vitro study. NT vessels dilated dose-dependently to adenosine (partially mediated by endothelial release of NO) and sodium nitroprusside (endothelium-independent vasodilator). Conversely, HT vessels exhibited reduced dilation to adenosine but dilated normally to sodium nitroprusside. Adenosine-stimulated NO release was increased approximately 3-fold in NT vessels but was reduced in HT vessels. Moreover, arginase activity was 2-fold higher in HT vessels. Inhibition of arginase activity by N(omega)-hydroxy-nor-l-arginine or incubation with l-arginine partially restored NO release and dilation to adenosine in HT vessels. Immunohistochemistry showed that arginase expression was increased but NOS expression was decreased in arteriolar ECs of HT vessels. These results suggest that NO-mediated dilation of coronary arterioles is inhibited in hypertension by an increase in arginase activity in EC, which limits l-arginine availability to NOS for NO production. The inability of arginase blockade or l-arginine supplementation to completely restore vasodilation may be related to downregulation of endothelial NOS expression.