Impact of myocardial structure and function postinfarction on diastolic strain measurements: implications for assessment of myocardial viability

Venular control of arteriolar perfusion has been the focus of several investigations in recent years. This study investigated 1) whether endogenous adenosine helps control venule-dependent arteriolar dilation and 2) whether venular leukocyte adherence limits this response via an oxidant-dependent mechanism in which nitric oxide (NO) levels are decreased. Intravital microscopy was used to assess changes in arteriolar diameters and NO levels in rat mesentery. The average resting diameter of arterioles (27.5 +/- 1.0 microm) paired with venules with minimal leukocyte adherence (2.1 +/- 0.3 per 100-microm length) was significantly larger than that of unpaired arterioles (24.5 +/- 0.8 microm) and arterioles (23.3 +/- 1.3 microm) paired with venules with higher leukocyte adherence (9.0 +/- 0.5 per 100-microm length). Local superfusion of adenosine deaminase (ADA) induced significant decreases in diameter and perivascular NO concentration in arterioles closely paired to venules with minimal leukocyte adherence. However, ADA had little effect on arterioles closely paired to venules with high leukocyte adherence or on unpaired arterioles. To determine whether the attenuated response to ADA for the high-adherence group was oxidant dependent, the responses were also observed in arterioles treated with 10(-4) M Tempol. In the high-adherence group, Tempol fully restored NO levels to those of the low-adherence group; however, the ADA-induced constriction remained attenuated, suggesting a possible role for an oxidant-independent vasoconstrictor released from the inflamed venules. These findings suggest that adenosine- and venule-dependent dilation of paired arterioles may be mediated, in part, by NO and inhibited by venular leukocyte adherence.     

Connexin expression and conducted vasodilation along arteriolar endothelium in mouse skeletal muscle.

Functional hyperemia requires the coordination of smooth muscle cell relaxation along and between branches of the arteriolar network. Vasodilation is conducted from cell to cell along the arteriolar wall through gap junction channels composed of connexin protein subunits. Within skeletal muscle, it is unclear whether arteriolar endothelium, smooth muscle, or both cell layers provide the cellular pathway for conduction. Furthermore, the constitutive profile of connexin expression within the microcirculation is unknown. We tested the hypothesis that conducted vasodilation and connexin expression are intrinsic to the endothelium of arterioles (17 +/- 1 microm diameter) that supply the skeletal muscle fibers in the cremaster of anesthetized C57BL/6 mice. ACh delivered to an arteriole (500 ms, 1-microA pulse; 1-microm micropipette) produced local dilation of 17 +/- 1 microm; conducted vasodilation observed 1 mm upstream was 9 +/- 1 microm (n = 5). After light-dye treatment to selectively disrupt endothelium (250-microm segment centered 500 microm upstream, confirmed by loss of local response to ACh while constriction to phenylephrine and dilation to sodium nitroprusside remained intact), we found that conducted vasodilation was nearly abolished (2 +/- 1 microm; P < 0.05). Whole-mount immunohistochemistry for connexins revealed punctate labeling at borders of arteriolar endothelial cells, with connexin40 and connexin37 in all branches and connexin43 only in the largest branches. Immunoreactivity for connexins was not apparent in smooth muscle or in capillary or venular endothelium, despite robust immunolabeling for alpha-actin and platelet endothelial cell adhesion molecule-1, respectively. We conclude that vasodilation is conducted along the endothelium of mouse skeletal muscle arterioles and that connexin40 and connexin37 are the primary connexins forming gap junction channels between arteriolar endothelial cells.     

Venular-arteriolar communication in the regulation of blood flow

Muscle blood flow is regulated to meet the metabolic needs of the tissue. With the vasculature arranged as a successive branching of arterioles and the larger, >50 microm, arterioles providing the major site of resistance, an increasing metabolic demand requires the vasodilation of the small arterioles first then the vasodilation of the more proximal, larger arterioles. The mechanism(s) for the coordination of this ascending vasodilation are not clear and may involve a conducted vasodilation and/or a flow-dependent response. The close arteriolar-venular pairing provides an additional mechanism by which the arteriolar diameter can be increased due to the diffusion of vasoactive substances from the venous blood. Evidence is presented that the venular endothelium releases a relaxing factor, a metabolite of arachidonic acid, that will vasodilate the adjacent arteriole. The stimulus for this release is not known, but it is hypothesized that hypoxia-induced ATP release from red blood cells may be responsible for the stimulation of arachidonic release from the venular endothelial cells. Thus the venous circulation is in an optimal position to monitor the overall metabolic state of the tissue and thus provide a feedback regulation of arteriolar diameter.     

Reactive oxygen species mediate arachidonic acid-induced dilation in porcine coronary microvessels

Reactive oxygen species (ROS) have been proposed to mediate vasodilation in the microcirculation. We investigated the role of ROS in arachidonic acid (AA)-induced coronary microvascular dilation. Porcine epicardial coronary arterioles (110 +/- 4 microm diameter) were mounted onto pipettes in oxygenated Krebs buffer. Vessels were incubated with vehicle or 1 mM Tiron (a nonselective ROS scavenger), 250 U/ml polyethylene-glycolated (PEG)-superoxide dismutase (SOD; an O2- scavenger), 250 U/ml PEG-catalase (a H2O2 scavenger), or the cyclooxygenase (COX) inhibitors indomethacin (10 microM) or diclofenac (10 microM) for 30 min. After endothelin constriction (30-60% of resting diameter), cumulative concentrations of AA (10(-10)-10(-5)M) were added and internal diameters measured by video microscopy. AA (10-7 M) produced 37 +/- 6% dilation, which was eliminated by the administration of indomethacin (4 +/- 7%, P < 0.05) or diclofenac (-8 +/- 8%, P < 0.05), as well as by Tiron (-4 +/- 5%, P < 0.05), PEG-SOD (-10 +/- 6%, P < 0.05), or PEG-catalase (1 +/- 4%, P < 0.05). Incubation of small coronary arteries with [3H]AA resulted in the formation of prostaglandins, which was blocked by indomethacin. In separate studies in microvessels, AA induced concentration-dependent increases in fluorescence of the oxidant-sensitive probe dichlorodihydrofluorescein diacetate, which was inhibited by pretreatment with indomethacin or by SOD + catalase. We conclude that in porcine coronary microvessels, COX-derived ROS contribute to AA-induced vasodilation.     

TNF-alpha activation of arterioles and venules alters distribution and levels of ICAM-1 and affects leukocyte-endothelial cell interactions

The observation that leukocyte-endothelial cell (EC) interactions are localized to specific regions on the microvessel wall suggests that adhesion molecule distribution is not uniform. We investigated ICAM-1 distribution and leukocyte-EC interactions in blood-perfused microvessels (<80 mum) in cremaster muscle of anesthetized mice, using intravital confocal microscopy and immunofluorescent labeling. Variability of ICAM-1 expression directly determines leukocyte adhesion distribution within the venular microcirculation and contributes to leukocyte rolling in arterioles during inflammation. The number of rolling interactions increased with ICAM-1 intensity (r(2) = 0.69, P < 0.05), and rolling velocity was lower in regions of higher ICAM-1 intensity. In controls, venular ICAM-1 expression was approximately twofold higher than in arterioles. After TNF-alpha treatment, ICAM-1 expression was significantly increased, 2.8 +/- 0.2-fold in arterioles and 1.7 +/- 0.2-fold in venules (P < 0.05). ICAM-1 expression on activated arteriolar ECs only reached the level of control venular ICAM-1. Arteriolar but not venular ECs underwent redistribution of ICAM-1 among cells; some cells increased and some decreased ICAM-1 expression, magnifying the variability of ICAM-1. TNF-alpha treatment increased the length of bright fluorescent regions per unit vessel length (42%, control; 70%, TNF-alpha) along the arteriolar wall, whereas no significant change was observed in venules (60%, control; 63%, TNF-alpha). The spatial distribution and expression levels of adhesion molecules in the microcirculation determine the timing and placement of leukocyte interactions and hence significantly impact the inflammatory response. That arteriolar ECs respond to TNF-alpha by upregulation of ICAM-1, although in a different way compared with venules, suggests an explicit role for arterioles in inflammatory responses.     

Governance of arteriolar oscillation by ryanodine receptors

To investigate the role of ryanodine receptors in glomerular arterioles, experiments were performed using an isolated perfused hydronephrotic kidney model. In the first series of studies, BAYK-8644 (300 nM), a calcium agonist, constricted afferent (19.6 +/- 0.6 to 17.6 +/- 0.5 microm, n = 6, P < 0.01) but not efferent arterioles. Furthermore, BAYK-8644 elicited afferent arteriolar oscillatory movements. Subsequent administration of nifedipine (1 microM) inhibited both afferent arteriolar oscillation and constriction by BAYK-8644 (to 19.4 +/- 0.5 microm). In the second group, although BAYK-8644 constricted afferent arterioles treated with 1 microM of thapsigargin (19.7 +/- 0.6 to 16.8 +/- 0.6 microm, n = 5, P < 0.05), it failed to induce rhythmic contraction. Removal of extracellular calcium with EGTA (2 mM) reversed BAYK-8644-induced afferent arteriolar constriction (to 20.0 +/- 0.5 microm). In the third series of investigations, ryanodine (10 microM) but not 2-aminoethoxyphenyl borate (100 microM) abolished afferent arteriolar vasomotion by BAYK-8644. In the fourth series of experiments, in the presence of caffeine (1 mM), the stronger activation of voltage-dependent calcium channels by higher potassium media resulted in greater afferent arteriolar constriction and faster oscillation. Our results indicate that L-type calcium channels are rich in preglomerular but not postglomerular microvessels. Furthermore, the present findings suggest that either prolonged calcium influx through voltage-dependent calcium channels (BAYK-8644) or sensitized ryanodine receptors (caffeine) is required to trigger periodic calcium release through ryanodine receptors in afferent arterioles.     

Chronic exposure to nicotine alters endothelium-dependent arteriolar dilatation: effect of superoxide dismutase.

The first goal of this study was to determine whether chronic injection of nicotine alters endothelium-dependent arteriolar dilatation. We measured the diameter of cheek pouch resistance arterioles (approximately 50 microm in diameter) in response to endothelium-dependent (acetylcholine and ADP) and -independent (nitroglycerin) agonists in control hamsters and hamsters treated with nicotine (2 microg. kg-1. day-1 for 2-3 wk). In control hamsters, acetylcholine (0.1 and 1.0 microM) dilated arterioles by 13 +/- 2 and 31 +/- 3%, respectively, and ADP (1.0 and 10 microM) dilated arterioles by 18 +/- 1 and 30 +/- 1%, respectively. In contrast, acetylcholine (0.1 and 1.0 microM) dilated arterioles by only 5 +/- 2 and 12 +/- 3%, respectively, and ADP (1.0 and 10 microM) dilated arterioles by only 7 +/- 2 and 13 +/- 3%, respectively, in animals treated with nicotine (P < 0.05 vs. response in control hamsters). Nitroglycerin produced similar dose-related dilatation of cheek pouch arterioles in control and nicotine-treated hamsters. Our second goal was to examine a possible mechanism for impaired endothelium-dependent arteriolar dilatation during chronic treatment with nicotine. We found that superfusion of the cheek pouch microcirculation with superoxide dismutase (150 U/ml) restored impaired endothelium-dependent, but did not alter endothelium-independent, arteriolar dilatation in hamsters treated with nicotine. Superfusion with superoxide dismutase did not alter endothelium-dependent or -independent arteriolar dilatation in control hamsters. We suggest that chronic exposure to nicotine produces selective impairment of endothelium-dependent arteriolar dilatation via a mechanism related to the synthesis/release of oxygen-derived free radicals.     

Arteriolar smooth muscle Ca2+ dynamics during blood flow control in hamster cheek pouch.

Intracellular calcium concentration ([Ca2+]i) governs the contractile status of arteriolar smooth muscle cells (SMC). Although studied in vitro, little is known of SMC [Ca2+]i dynamics during the local control of blood flow. We tested the hypothesis that the rise and fall of SMC [Ca2+]i underlies arteriolar constriction and dilation in vivo. Aparenchymal segments of second-order arterioles (diameter 35 +/- 2 microm) were prepared in the superfused cheek pouch of anesthetized hamsters (n = 18) and perifused with the ratiometric dye fura PE-3 (AM) to load SMC (1 microM, 20 min). Resting SMC [Ca2+]i was 406 +/- 37 nM. Elevating superfusate O2 from 0 to 21% produced constriction (11 +/- 2 microm) that was unaffected by dye loading; [Ca2+]i increased by 108 +/- 53 nM (n = 6, P < 0.05). Cycling of [Ca2+]i during vasomotion (amplitude, 150 +/- 53 nM; n = 4) preceded corresponding diameter changes (7 +/- 1 microm) by approximately 2 s. Microiontophoresis (1 microm pipette tip; 1 microA, 1 s) of phenylephrine (PE) transiently increased [Ca2+]i by 479 +/- 64 nM (n = 8, P < 0.05) with constriction (26 +/- 3 microm). Flushing blood from the lumen with saline increased fluorescence at 510 nm by approximately 45% during excitation at both 340 and 380 nm with no difference in resting [Ca2+]i, diameter or respective responses to PE (n = 7). Acetylcholine microiontophoresis (1 microA, 1 s) transiently reduced resting SMC [Ca2+]i by 131 +/- 21 nM (n = 6, P < 0.05) with vasodilation (17 +/- 1 microm). Superfusion of sodium nitroprusside (10 microM) transiently reduced SMC [Ca2+]i by 124 +/- 18 nM (n = 6, P < 0.05), whereas dilation (23 +/- 5 microm) was sustained. Resolution of arteriolar SMC [Ca2+]i in vivo discriminates key signaling events that govern the local control of tissue blood flow.     

Effects of trigeminal neurotransmitters on piglet pial arterioles.

We investigated effects of calcitonin gene-related peptide (CGRP), substance P (SP), and neurokinin A (NKA) on pial arterioles in newborn pigs. Pial arteriolar diameter was determined using a closed cranial window and intravital microscopy. Initial diameters were approximately 100 microns. Calcitonin-gene related peptide dilated pial arterioles by 22 +/- 8% at 10(-9)M and by 34 +/- 6% at 10(-8)M (n = 8), and this response was not significantly altered by prior administration of indomethacin (5mg/kg, iv) (n = 6) or administration of NG-methyl-L-arginine (5mg/kg, iv, and 10(-3)M in CSF) (n = 10). Substance P dilated arterioles at 10(-10)M through 10(-5)M (maximal response = 23 +/- 3%) (n = 6), and this response was unaffected by indomethacin administration (n = 6). In contrast, NG-methyl-L-arginine blocked much of the pial arteriolar dilation to SP. Unlike the other two peptides, NKA did not change pial arteriolar diameter. Radioimmunoassay determinations indicated that cerebrospinal fluid levels of 6-keto-prostaglandin F1 and prostaglandin E2 did not change appreciably during application of CGRP or SP. We conclude that CGRP and SP but not NKA are dilator stimuli in the piglet pial circulation. Dilation by CGRP probably involves direct activation of receptors on vascular smooth muscle, while SP probably partially dilates pial arterioles via release of an endothelium-dependent relaxing factor.     

Resveratrol reduction of infarct size in Long-Evans rats subjected to focal cerebral ischemia

Although vasomotion has been considered a feature of the microvascular bed under physiological conditions, it has also been observed following hypotension in several tissues. In this work, 158 mesenteric microvessels of 36 rats were investigated quantitatively in normovolemic and hemorrhaged animals, focussing on diameter changes, particularly vasomotion incidence and characteristics. The femoral arteries of Wistar rats (body weight BW = 188 +/- 23 g, mean +/- SD) anesthetized with pentobarbital were cannulated for arterial pressure (AP) monitoring and blood withdrawal. The protocol consisted of 15 min control and 30 min of hemorrhagic hypotension (AP = 52 +/- 5 mmHg, hemorrhaged vol. = 17 +/- 4 ml/kg BW). During control normovolemic conditions, analysis of mesenteric microcirculation using intravital videomicroscopy revealed neither arteriolar nor venular vasomotion. During hemorrhagic hypotension (HH) microvascular blood flow reduced to 25% of control. While venules did not show diameter changes during HH, arterioles contracted to 85 +/- 20% of control and arteriolar vasomotion appeared in 42% of the animals and 27% of the arterioles. The amplitude of arteriolar diameter change during HH relative to mean diameter and to control diameter averaged 65 +/- 24% (range: 32-129%) and 41 +/- 10% (range: 25-62%), respectively. Vasomotion analysis showed two major frequency components: 1.7 +/- 0.8 and 7.0 +/- 5.2 cycles/min. Arterioles showing vasomotion had a mean control diameter larger than the remaining arterioles and showed the largest constriction during HH. We conclude that hemorrhagic hypotension does not change venular diameter but induces arteriolar constriction and vasomotion in rat mesentery. This activity is expressed as slow waves with high amplitude and fast waves with low amplitude, and is dependent on vessel size.     

Reduced perivascular PO2 increases nitric oxide release from endothelial cells

Many studies have suggested that endothelial cells can act as "oxygen sensors" to large reductions in oxygen availability by increasing nitric oxide (NO) production. This study determined whether small reductions in oxygen availability enhanced NO production from in vivo intestinal arterioles, venules, and parenchymal cells. In vivo measurements of perivascular NO concentration ([NO]) were made with NO-sensitive microelectrodes during normoxic and reduced oxygen availability. During normoxia, intestinal first-order arteriolar [NO] was 397 +/- 26 nM (n = 5), paired venular [NO] was 298 +/- 34 nM (n = 5), and parenchymal cell [NO] was 138 +/- 36 nM (n = 3). During reduced oxygen availability, arteriolar and venular [NO] significantly increased to 695 +/- 79 nM (n = 5) and 534 +/- 66 nM (n = 5), respectively, whereas parenchymal [NO] remained unchanged at 144 +/- 34 nM (n = 4). During reduced oxygenation, arteriolar and venular diameters increased by 15 +/- 3% and 14 +/- 5%, respectively: NG-nitro-L-arginine methyl ester strongly suppressed the dilation to lower periarteriolar Po2. Micropipette injection of a CO2 embolus into arterioles significantly attenuated arteriolar dilation and suppressed NO release in response to reduced oxygen availability. These results indicated that in rat intestine, reduced oxygen availability increased both arteriolar and venular NO and that the main site of NO release under these conditions was from endothelial cells.     

Abolition of arteriolar dilation but not constriction to histamine in cremaster muscle of eNOS-/- mice

Histamine increases the permeability of capillaries and venules but little is known of its precapillary actions on the control of tissue perfusion. Using gene ablation and pharmacological interventions, we tested whether histamine could increase muscle blood flow through stimulating nitric oxide (NO) release from microvascular endothelium. Vasomotor responses to topical histamine were investigated in second-order arterioles in the superfused cremaster muscle of anesthetized C57BL6 mice and null platelet endothelial cell adhesion molecule-1 (PECAM-1-/-) and null endothelial NO synthase (eNOS-/-) mice aged 8-12 wk. Neither resting (17 +/- 1 microm) nor maximum diameters (36 +/- 2 microm) were different between groups, nor was the constrictor response (approximately 5 +/- 1 microm) to elevating superfusate oxygen from 0 to 21%. For arterioles of C57BL6 and PECAM-1-/- mice, cumulative addition of histamine to the superfusate produced vasodilation (1 nM-1 microM; peak response, 9 +/- 1 microm) and then vasoconstriction (10-100 microM; peak response, 12 +/- 2 microm). In eNOS-/- mice, histamine produced only vasoconstriction. In C57BL6 and PECAM-1-/- mice, vasodilation was abolished with Nomega-nitro-l-arginine (30 microM); in all mice, vasoconstriction was abolished with nifedipine (1 microM). Vasomotor responses were eliminated with pyrilamine (1 microM; H1 receptor antagonist) yet remained intact with cimetidine (1 microM; H2 receptor antagonist). These findings illustrate that the biphasic vasomotor response of mouse cremaster arterioles to histamine is mediated through H1 receptors on endothelium (NO-dependent vasodilation) as well as smooth muscle (Ca2+ entry and constriction). Thus histamine can increase as well as decrease muscle blood flow, according to local concentration. However, when NO production is compromised, only vasoconstriction and flow reduction occur.     

Diameter changes in skeletal muscle venules during arterial pressure reduction

Previous studies in skeletal muscle have shown a substantial (>100%) increase in venous vascular resistance with arterial pressure reduction to 40 mmHg, but a microcirculatory study showed no significant venular diameter changes in the horizontal direction during this procedure. To examine the possibility of venular collapse in the vertical direction, a microscope was placed horizontally to view a vertically mounted rat spinotrapezius muscle preparation. We monitored the diameters of venules (mean diameter 73. 8 +/- 37.0 microm, range 13-185 microm) oriented horizontally and vertically with a video system during acute arterial pressure reduction by hemorrhage. Our analysis showed small but significant (P < 0.0001) diameter reductions of 1.0 +/- 2.5 microm and 1.8 +/- 3. 1 microm in horizontally and vertically oriented venules, respectively, upon reduction of arterial pressure from 115.0 +/- 26. 3 to 39.8 +/- 12.3 mmHg. The venular responses were not different after red blood cell aggregation was induced by Dextran 500 infusion. We conclude that diameter changes in venules over this range of arterial pressure reduction are isotropic and would likely increase venous resistance by <10%.     

Dilation of rat diaphragmatic arterioles by flow and hypoxia: roles of nitric oxide and prostaglandins.

The in vitro responses to ACh, flow, and hypoxia were studied in arterioles isolated from the diaphragms of rats. The endothelium was removed in some vessels by low-pressure air perfusion. In endothelium-intact arterioles, pressurized to 70 mmHg in the absence of luminal flow, ACh (10(-5) M) elicited dilation (from 103 +/- 10 to 156 +/- 13 microm). The response to ACh was eliminated by endothelial ablation and by the nitric oxide synthase antagonists NG-nitro-L-arginine (L-NNA; 10(-5) M) and NG-nitro-L-arginine methyl ester (L-NAME, 10(-5) M) but not by indomethacin (10(-5) M). Increases in luminal flow (5-35 microl/min in 5 microl/min steps) at constant distending pressure (70 mmHg) elicited dilation (from 98 +/- 8 to 159 +/- 12 microm) in endothelium-intact arterioles. The response to flow was partially inhibited by L-NNA, L-NAME, and indomethacin and eliminated by endothelial ablation and by concurrent treatment with L-NAME and indomethacin. The response to hypoxia was determined by reducing the periarteriolar PO2 from 100 to 25-30 Torr by changing the composition of the gas used to bubble the superfusing solution. Hypoxia elicited dilation (from 110 +/- 9 to 165 +/- 12 microm) in endothelium-intact arterioles but not in arterioles from which the endothelium had been removed. Hypoxic vasodilation was eliminated by treatment with indomethacin and was not affected by L-NAME or L-NNA. In rat diaphragmatic arterioles, the response to ACh is dependent on endothelial nitric oxide release, whereas the response to hypoxia is mediated by endothelium-derived prostaglandins. Flow-dilation requires that both nitric oxide and cyclooxygenase pathways be intact.     

Myogenic responses and compliance of mesenteric and splenic vasculature in the rat

In the rat, the spleen is a major site of fluid efflux out of the blood. By contrast, the mesenteric vasculature serves as a blood reservoir. We proposed that the compliance and myogenic responses of these vascular beds would reflect their different functional demands. Mesenteric and splenic arterioles ( approximately 150-200 microm) and venules (<250 microm) from rats anesthetized with pentobarbital sodium were mounted in a pressurized myograph. Mesenteric arterial diameter decreased from 146 +/- 6 to 133 +/- 6 microm on raising intraluminal pressures from 80 to 120 mmHg. This response was enhanced in the presence of N(omega)-nitro-l-arginine methyl ester (l-NAME; 139 +/- 6 to 112 +/- 7 microm). There was no such myogenic response in the splenic arterioles, except in the presence of l-NAME (194 +/- 4 to 164 +/- 4.2 microm). We propose that, whereas mesenteric arterioles exhibit myogenic responses, this is normally masked by NO-mediated dilation in the splenic vessels. The mesenteric venules were highly distensible (active, 184 +/- 15 to 320 +/- 30.9 microm; passive in Ca(2+)-free media, 209 +/- 31 to 344 +/- 27 microm; 4-8 mmHg) compared with the splenic vessels (active, 169 +/- 11 to 184 +/- 16 microm; passive, 187 +/- 12 to 207 +/- 17 microm). We conclude that, in response to an increase in perfusion pressure, mesenteric arterial diameter would decrease to limit the changes in flow and microvascular pressure. In addition, mesenteric venous capacitance would increase. By contrast, splenic arterial diameter would increase, while there would be little change in venous diameter. This would enhance the increase in intrasplenic microvascular pressure and increase fluid extravasation.     

Hindlimb unweighting alters endothelium-dependent vasodilation and ecNOS expression in soleus arterioles.

The purpose of this study was to test the hypothesis that endothelium-dependent dilation is impaired in soleus resistance arteries from hindlimb-unweighted (HLU) rats. Male Sprague-Dawley rats (300-350 g) were exposed to HLU (n = 14) or weight-bearing control (Con, n = 14) conditions for 14 days. After the 14-day treatment period, soleus first-order (1A) arterioles were isolated and cannulated with micropipettes to assess vasodilator responses to an endothelium-dependent dilator, ACh (10(-9)-10(-4) M), and an endothelium-independent dilator, sodium nitroprusside (SNP, 10(-9)-10(-4) M). Arterioles from HLU rats were smaller than Con arterioles (maximal passive diameter = 140 +/- 4 and 121 +/- 4 microm in Con and HLU, respectively) but developed similar spontaneous myogenic tone (43 +/- 3 and 45 +/- 3% in Con and HLU, respectively). Arteries from Con and HLU rats dilated in response to increasing doses of ACh, but dilation was impaired in arterioles from HLU rats (P = 0.03), as was maximal dilation to ACh (85 +/- 4 and 65 +/- 4% possible dilation in Con and HLU, respectively). Inhibition of nitric oxide (NO) synthase (NOS) with N(omega)-nitro-L-arginine (300 microM) reduced ACh dilation by approximately 40% in arterioles from Con rats and eliminated dilation in arterioles from HLU rats. The cyclooxygenase inhibitor indomethacin (50 microM) did not significantly alter dilation to ACh in either group. Treatment with N(omega)-nitro-L-arginine + indomethacin eliminated all ACh dilation in Con and HLU rats. Dilation to sodium nitroprusside was not different between groups (P = 0.98). To determine whether HLU decreased expression of endothelial cell NOS (ecNOS), mRNA and protein levels were measured in single arterioles with RT-PCR and immunoblot analysis. The ecNOS mRNA and protein expression was significantly lower in arterioles from HLU rats than in Con arterioles (20 and 65%, respectively). Collectively, these data indicate that HLU impairs ACh dilation in soleus 1A arterioles, in part because of alterations in the NO pathway.     

N-methyl-D-aspartate-induced vasodilation is mediated by endothelium-independent nitric oxide release in piglets

N-methyl-D-aspartate (NMDA) elicits pial arteriolar dilation that has been associated with neuronal nitric oxide (NO) production. However, endothelial factors or glial P-450 epoxygenase products may play a role. We tested whether NMDA-induced pial vasodilation 1) primarily involves NO diffusion from the parenchyma to the surface arterioles, 2) involves intact endothelial function, and 3) involves a miconazole-sensitive component. Arteriolar diameters were determined using closed cranial window-intravital microscopy in anesthetized piglets. NMDA (10-100 microM) elicited virtually identical dose-dependent dilations in paired arterioles (r = 0.94, n = 15). However, NMDA- but not bradykinin (BK)-induced dilations of arteriolar sections over large veins were reduced by 31 +/- 1% (means +/- SE, P < 0.05, n = 4) compared with adjacent sections on the cortical surface. Also, 100 microM NMDA increased cerebrospinal fluid levels of NO metabolites from 3.7 +/- 1.0 to 5.3 +/- 1.2 microM (P < 0.05, n = 6). Endothelial stunning by intracarotid injection of phorbol 12,13-dibutyrate did not affect NMDA-induced vasodilation but attenuated vascular responses to hypercapnia and BK by approximately 70% (n = 7). Finally, miconazole (n = 6, 20 microM) pretreatment and coapplication with NMDA did not alter vascular responses to NMDA. In conclusion, NMDA appears to dilate pial arterioles exclusively through release and diffusion of NO from neurons to the pial surface in piglets.     

High-fat diet-induced reduction in nitric oxide-dependent arteriolar dilation in rats: role of xanthine oxidase-derived superoxide anion

Obesity frequently leads to the development of hypertension. We hypothesized that high-fat diet (HFD)-induced obesity impairs the endothelium-dependent dilation of arterioles. Male Wistar rats were fed with normal (control) or HFD (60% of saturated fat, for 10 wk). In rats with HFD, body weight, mean arterial blood pressure, and serum insulin, cholesterol, and glucose were elevated. In isolated gracilis muscle arterioles (diameter: approximately 160 microm) of HFD, rat dilations to ACh (at 1 microM, maximum: 83 +/- 3%) and histamine (at 10 microM, maximum: 16 +/- 4%) were significantly (P < 0.05) decreased compared with those of control responses (maximum: 90 +/- 2 and 46 +/- 4%, respectively). Dilations to the NO donor sodium nitroprusside were similar in the two groups. Inhibition of NO synthesis by N(omega)-nitro-l-arginine methyl ester reduced ACh- and histamine-induced dilations in control arterioles but had no effect on microvessels of HFD rats. The superoxide dismutase mimetic Tiron or xanthine oxidase inhibitor allopurinol enhanced ACh (maximum: 90 +/- 2 and 93 +/- 2%, respectively)- and histamine (maximum: 30 +/- 7 and 37 +/- 8%, respectively)-induced dilations in HFD arterioles, whereas the NAD(P)H oxidase inhibitor apocynin had no significant effect. Correspondingly, in carotid arteries of HFD rats, an enhanced superoxide production was shown by lucigenin-enhanced chemiluminescence, in association with an increased xanthine oxidase, but not NAD(P)H oxidase activity. In addition, a marked xanthine oxidase immunostaining was detected in the endothelial layer of the gracilis arterioles of HFD, but not in control rats. These findings suggest that, in obese rats, NO mediation of endothelium-dependent dilation of skeletal muscle arterioles is reduced because of an enhanced xanthine oxidase-derived superoxide production. These alterations demonstrate substantial dysregulation of arteriolar tone by the endothelium in HFD-induced obesity, which may contribute to disturbed tissue blood flow and development of increased peripheral resistance.     

Modulation of vascular reactivity to serotonin in the dog lung

Experiments were conducted to compare the effects of cyclooxygenase inhibition (COI) on vascular reactivity to serotonin (5-HT) in the isolated blood-perfused canine left lower lung lobe (LLL) and in isolated canine intrapulmonary lobar artery rings with and without a functional endothelium. LLLs (n = 6), perfused at constant blood flow, were challenged with bolus doses of 50, 100, and 250 micrograms 5-HT before COI, after COI with 45 microM meclofenamate, and after infusion of prostacyclin (PGI2) during COI. Lobar vascular resistance was segmentally partitioned by venous occlusion. Pulmonary arterial pressure increased from 13.5 +/- 1.0 to 16.3 +/- 0.8 cmH2O (P less than 0.01) after COI but declined to 13.1 +/- 1.1 cmH2O (P less than 0.01) subsequent to PGI2 infusion (91.3 +/- 14.5 ng.min-1.g LLL-1). The pulmonary arterial pressure changes were related to changes in postcapillary resistance. The dose-dependent pressor response to 5-HT was potentiated by COI (P less than 0.01) but reversibly attenuated (P less than 0.05) by PGI2 infusion. Isolated intrapulmonary artery rings (2-4 mm diam) exhibited a dose-related increase in contractile tension to 5-HT. The response to 5-HT was enhanced (P less than 0.05) in rings devoid of a functional endothelium. However, COI (10 microM indomethacin) did not alter (P greater than 0.05) the dose-related increase in contractile tension to 5-HT in rings with an intact endothelium. Our results suggest that both PGI2 and endothelium-derived relaxing factors modulate pulmonary vascular reactivity to 5-HT.(ABSTRACT TRUNCATED AT 250 WORDS)     

Microvascular pressure measurement reveals a coronary vascular waterfall in arterioles larger than 110 microm

Pressure-flow relationships at the entrance of the coronary circulation in the diastolic myocardium exhibit a zero-flow pressure intercept (P(int)). We tested whether this intercept is the same throughout the vascular bed. Microvascular pressure-flow relationships were therefore measured in vessels of various sizes of the maximally dilated vasculature of perfused unstimulated papillary muscle using the servo-null technique. From these relationships, P(int) were calculated with nonlinear regression. The P(int) at the level of the septal artery (diameter, 150-250 microm) was 23.2 +/- 4.4 cmH2O (n = 12). In arterioles with a diameter range between 24 and 110 microm, P(int) was 1.7 +/- 0.5 cmH2O (n = 6, P < 0.01), significantly lower than in the septal artery but significantly higher than zero, and not dependent on vessel size. In venules with the same diameters, P(int) was 1.1 +/- 1.1 cmH2O (n = 4), which was not different from zero. We conclude that, in the dilated vascular bed of the papillary muscle, two vascular waterfalls are found. The first waterfall is located in arterioles between 150 and 110 microm. The second waterfall is probably located in the small postcapillary venules.