Arterioles Of Rats Research Articles

Intraluminal pressure triggers myogenic response via activation of calcium spark and calcium-activated chloride channel in rat renal afferent arteriole.

Myogenic contraction of renal arterioles is an important regulatory mechanism for renal blood flow autoregulation. We have previously demonstrated that integrin-mediated mechanical force increases the occurrence of Ca2+ sparks in freshly isolated renal vascular smooth muscle cells (VSMCs). To further test whether the generation of Ca2+ sparks is a downstream signal of mechanotransduction in pressure-induced myogenic constriction, the relationship between Ca2+ sparks and transmural perfusion pressure was investigated in intact VSMCs of pressurized rat afferent arterioles. Spontaneous Ca2+ sparks were found in VSMCs when afferent arterioles were perfused at 80 mmHg. The spark frequency was significantly increased when perfusion pressure was increased to 120 mmHg. A similar increase of spark frequency was also observed in arterioles stimulated with β1-integrin-activating antibody. Moreover, spark frequency was significantly higher in arterioles of spontaneous hypertensive rats at 80 and 120 mmHg. Spontaneous membrane current recorded using whole cell perforated patch in renal VSMCs showed predominant activity of spontaneous transient inward currents instead of spontaneous transient outward currents when holding potential was set close to physiological resting membrane potential. Real-time PCR and immunohistochemistry confirmed the expression of Ca2+-activated Cl- channel (ClCa) TMEM16A in renal VSMCs. Inhibition of TMEM16A with T16Ainh-A01 impaired the pressure-induced myogenic contraction in perfused afferent arterioles. Our study, for the first time to our knowledge, detected Ca2+ sparks in VSMCs of intact afferent arterioles, and their frequencies were positively modulated by the perfusion pressure. Our results suggest that Ca2+ sparks may couple to ClCa channels and trigger pressure-induced myogenic constriction via membrane depolarization.

Inhibition of Shp2 ameliorates monocrotaline-induced pulmonary arterial hypertension in rats

BackgroundSrc homology 2 containing protein tyrosine phosphatase (PTP) 2 (Shp2) is a typical tyrosine phosphatase interacting with receptor tyrosine kinase to regulate multiple signaling pathways in diverse pathological processes. Here, we will investigate the effect of Shp2 inhibition on pulmonary arterial hypertension (PAH) in a rat model and its potential cellular and molecular mechanisms underlying.MethodsMonocrotaline (MCT)-induced PAH rat model was used in this study. Phps-1, a highly selective inhibitor for Shp2, was administered from 21 days to 35 days after MCT single-injection. Microcatheter method was applied to detected hemodynamic parameters. Histological methods were used to determine PVR changes in PAH rats. Moreover, cultured pulmonary artery smooth muscle cells (PASMCs) treated by platelet-derived growth factor (PDGF) with or without Phps-1 was used to investigate the potential cellular and molecular mechanisms underlying in vitro.ResultsInhibition of Shp2 significantly attenuated MCT-induced increases of mean pulmonary arterial pressure (mPAP), right ventricular systolic pressure (RVSP) and right ventricular hypertrophy (RVH) in rats. Shp2 inhibition effectively decreased thickening of pulmonary artery media and cardiomyocyte hypertrophy as well as perivascular and myocardial fibrosis in MCT-treated rats. Moreover, Shp2 inhibition ameliorated muscularization of pulmonary arterioles in MCT-induced PAH rats. Shp2 inhibition significantly reduced platelet-derived growth factor (PDGF)-triggered proliferation and migration of human pulmonary artery smooth muscle cells (PASMCs), which might be attributed to the inactivations of Akt and Stat3 pathways.ConclusionsShp2 contributes to the development of PAH in rats, which might be a potential target for the treatment of PAH.

Endothelium-dependent vasodilation in the cerebral arterioles of rats deteriorates during acute hyperglycemia and then is restored by reducing the glucose level.

Acute hyperglycemia in patients with traumatic brain injury correlates with a poor neurological outcome. We investigated the endothelium function of rat cerebral arterioles during acute hyperglycemia and after reducing the glucose levels using insulin. We also examined whether or not oxidative stress was involved in the cerebral arteriole response to acute hyperglycemia. In isoflurane-anesthetized, mechanically ventilated rats, we used closed cranial window preparation to measure the changes in the pial arteriolar diameter following the topical application of acetylcholine (ACh) or adenosine. We examined the pial arteriolar vasodilator response before hyperglycemia, during hyperglycemia, and after reducing the glucose level using insulin. After intravenous pretreatment with an NADPH oxidase inhibitor (apocynin or diphenylene iodonium), we reexamined the pial arteriolar vasodilator response following the topical application of ACh. Under control conditions, the topical application of ACh dose-dependently dilated the cerebral arterioles. The vasodilatory responses to topical ACh were impaired during hyperglycemia and improved after the administration of insulin. The vasodilatory responses to topical adenosine were not affected by the glucose levels. In the apocynin or diphenylene iodonium pretreatment group, the topical application of ACh dilated the cerebral arterioles during hyperglycemia. Acute hyperglycemia induces a dysfunction of the endothelium-dependent vasodilation of rat cerebral arterioles. The dysfunction can be reversed by improving the acute hyperglycemia and it can be prevented entirely by the administration of NADPH oxidase inhibitors. These results could suggest that controlling the glucose levels works protectivity to endothelium function of cerebral arterioles.

Daily muscle stretching enhances blood flow, endothelial function, capillarity, vascular volume and connectivity in aged skeletal muscle.

In aged rats, daily muscle stretching increases blood flow to skeletal muscle during exercise. Daily muscle stretching enhanced endothelium-dependent vasodilatation of skeletal muscle resistance arterioles of aged rats. Angiogenic markers and capillarity increased in response to daily stretching in muscles of aged rats. Muscle stretching performed with a splint could provide a feasible means of improving muscle blood flow and function in elderly patients who cannot perform regular aerobic exercise. Mechanical stretch stimuli alter the morphology and function of cultured endothelial cells; however, little is known about the effects of daily muscle stretching on adaptations of endothelial function and muscle blood flow. The present study aimed to determine the effects of daily muscle stretching on endothelium-dependent vasodilatation and muscle blood flow in aged rats. The lower hindlimb muscles of aged Fischer rats were passively stretched by placing an ankle dorsiflexion splint for 30minday-1 , 5daysweek-1 , for 4weeks. Blood flow to the stretched limb and the non-stretched contralateral limb was determined at rest and during treadmill exercise. Endothelium-dependent/independent vasodilatation was evaluated in soleus muscle arterioles. Levels of hypoxia-induced factor-1α, vascular endothelial growth factor A and neuronal nitric oxide synthase were determined in soleus muscle fibres. Levels of endothelial nitric oxide synthase and superoxide dismutase were determined in soleus muscle arterioles, and microvascular volume and capillarity were evaluated by microcomputed tomography and lectin staining, respectively. During exercise, blood flow to plantar flexor muscles was significantly higher in the stretched limb. Endothelium-dependent vasodilatation was enhanced in arterioles from the soleus muscle from the stretched limb. Microvascular volume, number of capillaries per muscle fibre, and levels of hypoxia-induced factor-1α, vascular endothelial growth factor and endothelial nitric oxide synthase were significantly higher in the stretched limb. These results indicate that daily passive stretching of muscle enhances endothelium-dependent vasodilatation and induces angiogenesis. These microvascular adaptations may contribute to increased muscle blood flow during exercise in muscles that have undergone daily passive stretch.

Activation of cannabinoid (CB2) receptors and reactivity of cerebral arterioles

While activation of cannabinoid (CB2) receptors has been shown to be neuroprotective, no studies have examined whether this neuroprotection is directed at cerebral arterioles and no studies have examined whether activation of CB2 receptors can rescue cerebrovascular dysfunction during a chronic disease states such as type 1 diabetes (T1D). Our goal was to test that hypothesis that administration of a CB2 agonist (JWH‐133) would improve impaired endothelial (eNOS)‐ and neuronal (nNOS)‐ dependent dilation of cerebral arterioles during T1D. In vivo diameter of cerebral arterioles in nondiabetic and diabetic rats was measured in response to an eNOS‐dependent agonist (adenosine 5′‐diphosphate; ADP), an nNOS‐dependent agonist (N‐methyl‐D‐aspartate; NMDA), and an NOS‐independent agonist (nitroglycerin) before and following JWH‐133 (1 mg/kg IP). Dilation of cerebral arterioles to ADP and NMDA was greater in nondiabetic than in diabetic rats. Treatment with JWH‐133 increased responses of cerebral arterioles to ADP and NMDA in both nondiabetic and diabetic rats. Responses of cerebral arterioles to nitroglycerin were similar between nondiabetic and diabetic rats, and JWH‐133 did not influence responses to nitroglycerin in either group. Thus, it appears that activation of CB2 receptors can influence vascular function of cerebral arterioles during physiologic and pathophysiologic states. We speculate that treatment with CB2 receptor agonists may have potential therapeutic benefits for the treatment of cerebral vascular disease via a mechanism that can increase cerebral blood flow.Support or Funding InformationThese studies were supported by funds from the Diabetes Action Research and Education FoundationThis abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Probucol prevents the attenuation of β2-adrenoceptor-mediated vasodilation of retinal arterioles in diabetic rats.

Probucol is an antihyperlipidemic drug with potent antioxidant properties. Oxidative stress plays an important role in the pathogenesis of diabetic retinopathy. In this study, we aimed to investigate the protective effects of probucol against diabetes-induced retinal vascular dysfunction in a rat model of diabetes. Diabetes was induced by a combination of streptozotocin treatment and D-glucose feeding, and retinal vasodilator responses were assessed by measuring the diameter of retinal arterioles. The vasodilator effect of salbutamol, a β2-adrenoceptor agonist, on retinal arterioles was significantly diminished 2weeks after the induction of diabetes. In non-diabetic rats, vasodilator responses to salbutamol were significantly reduced after an intravitreal injection of iberiotoxin, a blocker of large-conductance KCa (BKCa) channels. However, this effect was not observed in diabetic rats. Probucol had no significant effect on salbutamol-induced changes in diameter of retinal arterioles in non-diabetic rats, whereas it could prevent the attenuation of retinal vasodilator response to salbutamol in diabetic rats. These results suggest that the reduced function of BKCa channels is involved in the attenuation of β2-adrenoceptor-mediated retinal vasodilation in diabetic rats. Probucol preserves the BKCa channel function in retinal arterioles under diabetic conditions; therefore, it may show beneficial effects on diabetic retinopathy by preventing or slowing the impairment of the retinal circulation in patients with diabetes mellitus.

Therapeutic benefits of intravenous cardiosphere-derived cell therapy in rats with pulmonary hypertension.

Pulmonary arterial hypertension (PAH) is a progressive condition characterized by occlusive pulmonary arteriopathy, in which survival remains poor despite pharmacologic advances. The aim of this study was to evaluate the ability of cardiosphere-derived cells (CDCs), cardiac progenitor cells with potent anti-inflammatory and immunomodulatory properties, to attenuate hemodynamic and morphometric remodeling of the right ventricle (RV) and pulmonary arterioles in rats with established monocrotaline (MCT)-induced PAH. Animals were divided into 3 groups: 1) Control (CTL), 2) PAH in which CDCs were centrally infused (CDC) and 3) PAH in which saline was given (Sham). Significant increments in RV systolic pressure (RVSP) and RV hypertrophy were noted in Sham animals compared to CTL. In CDC rats at day 35, RSVP fell (- 38%; p< 0.001) and RV hypertrophy decreased (-26%; p< 0.01). TAPSE and cardiac output were preserved in all 3 groups at day 35. Pulmonary arteriolar wall thickness was greater in Sham rats compared to CTL, and reduced in CDC animals for vessels 20–50 μm (P<0.01; back to CTL levels) and 50–80μm (P<0.01) in diameter. The macrophage population was increased in Sham animals compared to CTL (P< 0.001), but markedly reduced in CDC rats. In conclusion, infusion of CDCs markedly attenuated several key pathophysiologic features of PAH. As adjunctive therapy to PAH-specific agents, CDCs have the potential to impact on the pathobiology of adverse pulmonary arteriolar remodeling, by acting on multiple mechanisms simultaneously.

Effects of Nuclear Factor-E2-related factor 2/Heme Oxygenase 1 on splanchnic hemodynamics in experimental cirrhosis with portal hypertension

ObjectiveWe explored the effects of Nuclear Factor-E2-related factor 2 (Nrf2) and Heme Oxygenase 1 (HO-1) on splanchnic hemodynamics in portal hypertensive rats. MethodsExperimental cirrhosis with portal hypertension was induced by intraperitoneal injection of carbon tetrachloride. The expression of proteins was examined by immunoblotting. Hemodynamic studies were performed by radioactive microspheres. The vascular perfusion system was used to measure the contractile response of mesentery arterioles in rats. ResultsNrf2 expression in the nucleus and HO-1 expression in cytoplasm was significantly enhanced in portal hypertensive rats. Portal pressure, as well as regional blood flow, increased significantly in portal hypertension and can be blocked by tin protoporphyrin IX. The expression of endogenous nitric oxide synthase and vascular endothelial growth factors increased significantly compared to normal rats, while HO-1 inhibition decreased the expression of these proteins significantly. The contractile response of mesenteric arteries decreased in portal hypertension, but can be partially recovered through tin protoporphyrin IX treatment. ConclusionsThe expression of Nrf2/HO-1 increased in mesenteric arteries of portal hypertensive rats, which was related to oxidative stress. HO-1was involved in increased portal pressure and anomaly splanchnic hemodynamics in portal hypertensive rats.

Juvenile growth reduces the influence of epithelial sodium channels on myogenic tone in skeletal muscle arterioles.

Previous studies have documented that rapid juvenile growth is accompanied by functional changes in the arteriolar endothelium, but much less is known about functional changes in arteriolar smooth muscle over this period. In this study, we investigate the possible contribution of epithelial sodium channels (ENaC) to the myogenic behaviour of arterioles at two stages of juvenile growth. The effects of the ENaC inhibitor benzamil on different levels of myogenic tone were studied in isolated gracilis muscle arterioles from rats aged 21-28days ("weanlings") and 42-49days ("juveniles"). ENaC subunit expression in the arteriolar wall was also determined, and the interaction between ENaC and nitric oxide (NO) in regulating vascular tone was explored by combined use of benzamil and NG -monomethyl-l-arginine (l-NMMA). At physiological pressures, both steady-state myogenic tone and the dynamic adjustments in this tone triggered by acute pressure changes were less in juvenile arterioles than in weanling arterioles. α, β and γ ENaC protein was present in arterioles at both ages, but benzamil only had an effect on myogenic tone in weanling arterioles. In these vessels, benzamil increased, rather than decreased, myogenic tone, and this effect was prevented by l-NMMA or endothelial removal. These findings suggest that although ENaC is present in gracilis muscle arterioles of both weanling and juvenile rats, it is not obligatory for the genesis of myogenic activity in these vessels at either age. However, ENaC activity can significantly modulate the level of myogenic tone through stimulation of endothelial NO release at an early stage of growth.

Tetrahydrobiopterin rescues impaired responses of cerebral resistance arterioles during type 1 diabetes.

Our goal was to test the hypothesis that administration of tetrahydrobiopterin (BH4) would improve impaired endothelial nitric oxide synthase-dependent dilation of cerebral arterioles during type 1 diabetes. In addition, we examined the influence of BH4 on levels of superoxide in brain tissue. In vivo diameter of cerebral arterioles in nondiabetic and diabetic rats was measured in response to endothelial nitric oxide synthase-dependent agonists (acetylcholine and adenosine 5'-diphosphate) and an endothelial nitric oxide synthase-independent agonist (nitroglycerine) before and during application of BH4 (1.0 µM). We also measured levels of superoxide from cortex tissue in nondiabetic and diabetic rats under basal states and during BH4 Acetylcholine and adenosine 5'-diphosphate dilated cerebral arterioles in nondiabetic rats, but this vasodilation was significantly impaired in diabetic rats. In contrast, nitroglycerine produced similar vasodilation in nondiabetic and diabetic rats. Application of BH4 did not enhance vasodilation in nondiabetic rats but improved impaired cerebral vasodilation in diabetic rats. Basal superoxide levels were increased in cortex tissue from diabetic rats, and BH4 reduced these levels to that found in nondiabetic rats. Thus, BH4 is an important mediator of endothelial nitric oxide synthase-dependent responses of cerebral arterioles in diabetes and may have therapeutic potential for the treatment of cerebral vascular disease.

Gα11 expression and effect of sildenafil in muscularization of non-muscular pulmonary arterioles in rat with pulmonary arterial hypertension

To investigate expression changes and role of Gα11 protein in the processes of muscularization of non-muscular pulmonary arterioles and effect of sildenafil intervention in rats with pulmonary arterial hypertension (PAH). Thirty SD rats were randomly divided into three groups, including normal control group, monocrotaline (MCT) group and sildenafil group; PAH model was prepared with 50 mg/kg MCT treatment for 4 weeks in the MCT group, and these rats were treated by 25 mg/kg sildenafil for 2 weeks after PAH formation in the sildenafil group, and the normal control group were treated with the equal amounts of physiological saline instead of monocrotaline; pulmonary artery pressure was measured with jugular vein catheterization; hematoxylin and eosin (HE) staining method was used to detect the pulmonary arteriolar morphology and vascular tissue parameters; expression of the target Gα11 protein, vascular smooth muscle marker osteopontin (OPN) and proliferation marker proliferating cell nuclear antigen (PCNA) was detected by Western blot. Pulmonary artery mean pressure (mPAP), non-muscular pulmonary arterioles wall thickness index (TI) and area index (AI) of the MCT group were higher than those of the normal control group[(27.43±3.97) vs (11.93±1.52) mmHg (1 mmHg=0.133 kPa), 0.49±0.07 vs 0.31±0.09 and 0.74±0.05 vs 0.45±0.10](all P<0.05), and meanwhile the expression levels of Gα11 and the related proteins including OPN and PCNA were significantly enhanced. mPAP, TI and AI[(18.59±1.44) mmHg, 0.39±0.09 and 0.56±0.04]of the sildenafil group were all lower than those of the MCT group (all P<0.05), and furthermore, expressions of Gα11, OPN and PCNA also reduced in line with these changes. Gα11 protein plays a role in the development of PAH and pulmonary non-muscular arteriole muscularization, and sildenafil effectively suppresses PAH and pulmonary vascular remodeling by inhibiting Gα11 expression.

Sanguinate's effect on pial arterioles in healthy rats and cerebral oxygen tension after controlled cortical impact

Sanguinate, a polyethylene glycol-conjugated carboxyhemoglobin, was investigated for cerebral vasoactivity in healthy male Sprague-Dawley rats (Study 1) and for its ability to increase brain tissue oxygen pressure (PbtO2) after controlled cortical impact (CCI) — traumatic brain injury (TBI) (Study 2). In both studies ketamine-acepromazine anesthetized rats were ventilated with 40% O2. In Study 1, a cranial window was used to measure the diameters of medium — (50–100μm) and small-sized (<50μm) pial arterioles before and after four serial infusions of Sanguinate (8mL/kg/h, cumulative 16mL/kg IV), volume-matched Hextend, or normal saline. In Study 2, PbtO2 was measured using a phosphorescence quenching method before TBI, 15min after TBI (T15) and then every 10min thereafter for 155min. At T15, rats received either 8mL/kg IV Sanguinate (40mL/kg/h) or no treatment (saline, 4mL/kg/h). Results showed: 1) in healthy rats, percentage changes in pial arteriole diameter were the same among the groups, 2) in TBI rats, PbtO2 decreased from 36.5±3.9mmHg to 19.8±3.0mmHg at T15 in both groups after TBI and did not recover in either group for the rest of the study, and 3) MAP increased 16±4mmHg and 36±5mmHg after Sanguinate in healthy and TBI rats, respectively, while MAP was unchanged in control groups. In conclusion, Sanguinate did not cause vasoconstriction in the cerebral pial arterioles of healthy rats but it also did not acutely increase PbtO2 when administered after TBI. Sanguinate was associated with an increase in MAP in both studies.

Decreased Uterine Vascularization and Uterine Arteriolar Expansive Remodeling by Matrix Metalloproteinases in Hypertensive Pregnancy

Normal pregnancy is associated with extensive remodeling of spiral arteries in order to maintain nutrients and blood supply to the growing fetus. Inadequate placentation and invasion of the uterine wall could lead to uteroplacental ischemia and hypertensive pregnancy (HTN‐Preg); however, the extent of uterine vascularization and the mechanisms involved are unclear. We hypothesized that HTN‐Preg is associated with changes in uterine vascularization and arterial remodeling by matrix metalloproteinases (MMPs). Blood pressure (BP) and fetal parameters were assessed on day‐19 in normal pregnant rats (Preg) and pregnant rats with reduced uterine perfusion pressure (RUPP), and the uteri were isolated for measurement of tissue vascularity and MMPs expression/activity. BP was greater and pups number and weight, and uterus weight were reduced in RUPP vs. Preg rats (BP 128.2±1.7 vs. 91.60±2.8mmHg; pups number 8.8±1.1 vs. 13.00±0.7; pup weight 1.48±0.059 vs. 2.02±0.02g; uterus weight 3.82±0.37 vs. 4.92±0.27g). Histological analysis of uterine tissue sections showed reduced total number and whole area of uterine arterioles in RUPP vs. Preg rats (number 5.75±0.95 vs. 11.50±0.87; area 0.05±0.01 vs. 0.12±0.02mm2). Immunohistochemistry with the endothelial cell marker CD31 showed localization in the uterine arterial intima and confirmed reduction in the number and size of uterine arterioles in RUPP vs. Preg rats. Further analysis of the uterine arterial wall showed reduction in the area of all three layers of the uterine arterial wall in RUPP vs. Preg rats (intima 0.015±0.003 vs. 0.031±0.005; media 0.012±0.002 vs. 0.038±0.005; adventitia 0.021±0.003 vs. 0.048±0.007mm2). Western blots and gelatin zymography showed a decrease in expression and activity of MMP‐2 and ‐9 in uterus and uterine arteries of RUPP vs. Preg rats. Immunohistochemistry showed decreased total amount of MMP‐2 and ‐9 in the uterus, and reduction in MMP distribution in the intima and media of uterine arteries of RUPP vs. Preg rats. Thus, in HTN‐Preg rats there is reduction in the number and size of uterine arteries and decreased expression and activity of MMP‐2 and ‐9. These results suggest that reduced uterine vascularization and uterine arteriolar expansive remodeling by MMPs could be a factor in the uteroplacental ischemia associated with HTN‐Preg and preeclampsia.Support or Funding InformationThis work was supported by grants from National Heart, Lung, and Blood Institute (HL‐65998,HL‐111775).

Abstract P090: Agonistic Autoantibodies to the Angiotensin II Type 1 Receptor Enhances ANG II Induced Renal Vascular Sensitivity and Reduces Renal Function During Pregnancy

Preeclamptic women produce agonistic autoantibodies to the Angiotensin II type 1 receptor (AT1-AA) and exhibit increased blood pressure (BP) and vascular sensitivity to angiotensin II (ANG II). Although, together AT1-AAs and ANGII increase the BP, renal artery resistant index, and vasoconstriction of renal afferent arterioles in pregnant rats; the renal hemodynamics in the presence of the AT1-AAs during pregnancy has not been examined. Thus the objective of this study was to examine the changes in the glomerular filtration rate (GFR) and renal blood flow (RBF) during pregnancy in the presence of AT1-AAs and/or ANGII. Methods: Pregnant Sprague Dawley rats were divided into 4 groups: Normal Pregnant (NP, n=6), Pregnant + ANG II (Preg + ANG II, n=6), Pregnant + AT1-AA (Preg + AT1-AA, n=8), and Pregnant + ANG II + AT1-AA (Preg + ANGII + AT1-AA, n= 6). On day 13 of pregnancy, rats were implanted with mini-pumps infusing ANG II (50 ng/kg/min) and/or AT1-AA (1:40 dilution). On day 19 of pregnancy, rats were subjected to terminal renal function surgeries using FITC labeled Inulin. During the surgery, the BP was recorded and a transonic flowmeter probe was placed on the left renal artery to measure RBF. Results: BP was elevated in all pregnant rats administered ANG II and/or the AT1-AA. Although GFR was reduced, it was not significant between Preg + ANG II and Preg + AT1-AA vs. NP rats (1.5 ±0.24, 1.60 ±0.17 vs. 1.90 ±0.16 ml/min). However, the GFR was further decreased in Preg + ANGII + AT1-AA rats (1.20 ±0.08). No difference was observed with the RBF between Preg + ANG II and Preg + AT1-AA vs NP rats (14.4 ±2.96, 14.4 ±1.48 vs. 15.4 ±1.75 ml/min). RBF was decreased in Preg + ANGII + AT1-AA vs NP rats (7.4 ±1.09 vs. 15.4 ±1.75 ml/min). No change in RVR between Preg + ANG II and Preg + AT1-AA vs. NP rats (9.7 ±2.69, 8.3 ±0.58 vs. 6.4 ±0.77). However, the RVR was drastically increased between Preg + ANGII + AT1-AA vs NP rats (18.4 ±2.91 vs. 6.4 ±0.77). Conclusion: Together ANG II and AT1-AA drastically decreases renal function by 37%, RBF by 50%, and caused a 3 fold increase in RVR vs NP rats. These data indicate the importance of AT1-AAs to drastically enhance ANG II induced renal vascular sensitivity and reduce renal function during preeclampsia. Research Supported by T32HL105324 and RO1HD067541

The zinc transporter ZIP12 regulates the pulmonary vascular response to chronic hypoxia.

The typical response of the adult mammalian pulmonary circulation to a low oxygen environment is vasoconstriction and structural remodelling of pulmonary arterioles, leading to chronic elevation of pulmonary artery pressure (pulmonary hypertension) and right ventricular hypertrophy. Some mammals, however, exhibit genetic resistance to hypoxia-induced pulmonary hypertension1-3. We used a congenic breeding program and comparative genomics to exploit this variation in the rat and identified the gene, Slc39a12, as a major regulator of hypoxia-induced pulmonary vascular remodelling. Slc39a12 encodes the zinc transporter, ZIP12. We report that ZIP12 expression is increased in many cell types, including endothelial, smooth muscle and interstitial cells, in the remodelled pulmonary arterioles of rats, cows and humans susceptible to hypoxia-induced pulmonary hypertension. We show that ZIP12 expression in pulmonary vascular smooth muscle cells is hypoxia-dependent and that targeted inhibition of ZIP12 inhibits the rise in intracellular labile zinc in hypoxia-exposed pulmonary vascular smooth muscle cells and their proliferation in culture. We demonstrate that genetic disruption of ZIP12 expression attenuates the development of pulmonary hypertension in rats housed in a hypoxic atmosphere. This entirely novel and unexpected insight into the fundamental role of a zinc transporter in mammalian pulmonary vascular homeostasis suggests a new drug target for the pharmacological management of pulmonary hypertension.

Exercise training causes differential changes in gene expression in diaphragm arteries and 2A arterioles of obese rats.

We employed next-generation, transcriptome-wide RNA sequencing (RNA-Seq) technology to assess the effects of two different exercise training protocols on transcriptional profiles in diaphragm second-order arterioles (D2a) and in the diaphragm feed artery (DFA) from Otsuka Long Evans Tokushima Fatty (OLETF) rats. Arterioles were isolated from the diaphragm of OLETF rats that underwent an endurance exercise training program (EX; n = 13), interval sprint training program (SPRINT; n = 14), or remained sedentary (Sed; n = 12). Our hypothesis was that exercise training would have similar effects on gene expression in the diaphragm and soleus muscle arterioles because diaphragm blood flow increases during exercise to a similar extent as in soleus. Results reveal that several canonical pathways that were significantly altered by exercise in limb skeletal muscles were not among the pathways significantly changed in the diaphragm arterioles including actin cytoskeleton signaling, role of NFAT in regulation of immune response, protein kinase A signaling, and protein ubiquitination pathway. EX training altered the expression of a smaller number of genes than did SPRINT in the DFA but induced a larger number of genes with altered expression in the D2a than did SPRINT. In fact, FDR differential expression analysis (FDR, 10%) indicated that only two genes exhibited altered expression in D2a of SPRINT rats. Very few of the genes that exhibited altered expression in the DFA or D2a were also altered in limb muscle arterioles. Finally, results indicate that the 2a arterioles of soleus muscle (S2a) from endurance-trained animals and the DFA of SPRINT animals exhibited the largest number of genes with altered expression.

The molecular pathway of ATP-sensitive potassium channel in endothelial cells for mediating arteriole relaxation.

The endothelial molecular pathway of a new ATP-sensitive potassium channel (KATP) opener natakalim was investigated in mesenteric arterioles of rats. A DMT wire myograph was used to evaluate the vasorelaxation effects of natakalim. Ca(2+) responses of endothelial cells induced by natakalim were measured by laser confocal fluorescence microscopy. NO assay kits and Western blotting were used. The new KATP opener natakalim significantly produced endothelium-dependent arteriolar dilation and increased endothelial cell intracellular calcium concentration ([Ca(2+)]i) as well as NO release, which could be inhibited by SB366791 and capsazepine, the specific TRPV1 blockers. Additionally, down-regulation of endothelial TRPV1 by RNA interference inhibited the Ca(2+) influx induced by natakalim. These results suggest that endothelial KATP mediated natakalim-induced vasorelaxation through increasing [Ca(2+)]i and NO production. Activation of endothelial TRPV1 channels and subsequent Ca(2+) entry, and NO release at least partly contribute to endothelium-dependent vasorelaxation induced by natakalim.

Involvement of prostaglandin I2 in nitric oxide-induced vasodilation of retinal arterioles in rats

The soluble guanylyl cyclase/cGMP system plays an important role in the vasodilator response to nitric oxide (NO) in various vascular beds. However, in rat retinal arterioles, the cyclooxygenase-1/cAMP-mediated pathway contributes to the vasodilator effects of NO, although the specific prostanoid involved remains to be elucidated. In the present study, we investigated the role of prostaglandin I2 and its receptor (prostanoid IP receptor) system in NO-induced vasodilation of rat retinal arterioles in vivo. Fundus images were captured using a digital camera that was equipped with a special objective lens. Changes in diameter of retinal arterioles were assessed. The NO donor (±)-(E)-4-ethyl-2-[(E)-hydroxyimino]-5-nitro-3-hexenamide (NOR3) increased the diameter of retinal arterioles but decreased systemic blood pressure in a dose-dependent manner. Treatment of rats with indomethacin, a non-selective cyclooxygenase inhibitor, markedly attenuated the retinal vasodilator, but not depressor responses to NOR3. The prostanoid IP receptor antagonist 4,5-dihydro-N-[4-[[4-(1-methylethoxy)phenyl]methyl]phenyl]-1H-imadazol-2-amine (CAY10441), and the prostaglandin I2 synthase inhibitor 9α,11α-azoprosta-5Z,13E-dien-1-oic acid (U-51605), both showed similar preventive effects against the NOR3-induced retinal vasodilator response. Neither CAY10441 nor U-51605 showed any significant effects on the depressor response to NOR3. NOR3 enhanced the release of prostaglandin I2 from cultured human retinal microvascular endothelial cells and the NOR3-induced prostaglandin I2 release was almost completely abolished by the cyclooxygenase-1 inhibitor SC-560, but not by the cyclooxygenase-2 inhibitor NS-398. However, NOR3 did not increase the release of prostaglandin I2 from human intestinal microvascular endothelial cells. These results suggest that NO exerts its dilatory effect via cyclooxygenase-1/prostaglandin I2/prostanoid IP receptor signaling mechanisms in the retinal vasculature.

Xiaokening stimulates endothelial nitric oxide release in diabetic rats.

Diabetes mellitus induces microangiopathic changes that lead to endothelial dysfunction. This study investigated the effect of Xiaokening, a type of Chinese compound medicine, on the mesenteric arteriolar endothelial cell function of diabetic rats and its underlying mechanism. Diabetes mellitus was induced in rat models via intraperitoneal injection of 60 mg/kg streptozotocin and observed over three weeks. Mesenteric arterioles, which were isolated in a cannulated and pressurised state, were incubated with intravascular injections of 1, 3 or 5 g/L Xiaokening for 24, 48 or 72 hours. The effects of Xiaokening on the release of nitric oxide (NO) on the mesenteric arterioles were detected under shear stress of 1, 10 and 20 dyn/cm(2). Biochemical methods were used to determine the activities of superoxide dismutase (SOD) and xanthine oxidase (XO). The expressions of endothelial NO synthase (eNOS), SOD and XO in the mesenteric arterioles were assessed using Western blot. Compared to normal rat arterioles, less NO was released in the mesenteric arterioles of diabetic rats. Xiaokening was found to have a concentration- and time-dependent effect on NO release; when the shear stress was increased, there was a gradual increase in the release of NO. Compared to normal arterioles, the expression of eNOS in the mesenteric arterioles of diabetic rats was lower. Incubation with Xiaokening increased SOD activity and expression, and decreased XO activity and expression in the mesenteric arterioles of the diabetic rats. Xiaokening was able to significantly increase NO release and improve the endothelial function of mesenteric arterioles through antioxidative mechanisms.

Effects of Perfluorocarbon Dodecafluoropentane (NVX-108) on Cerebral Microvasculature in the Healthy Rat

NVX-108, a dodecafluoropentane-based perfluorocarbon (PFC) emulsion, has therapeutic potential as an oxygen- carrying fluid for emergency medical treatment for traumatic brain injury (TBI) and hemorrhagic shock. Potential cerebral vasoactive properties were assessed by directly measuring pial arteriolar vessel diameters before and after a 30 minute intravenous (IV) infusion of 1.0 ml/kg (high dose [H]) or 0.25 ml/kg (low dose [L]) NVX-108 compared to 2.0 ml/kg Saline (control) in healthy anesthetized rats (N = 6/group). Results showed that post-infusion vessel diameters for small (< 50 µm) and medium (50-100 µm)-sized pial arterioles were significantly (p < 0.05) narrower after only the NVX- 108 H infusion although this vasoconstriction was not statistically significant when analyzed as a percentage change in these vessels. Pial arteriolar vessel diameters were not significantly different for mean value or percentage change after either NVX-108 L or Saline infusions. There were no significant post-infusion changes from baseline in systolic, mean or diastolic blood pressures after any of the treatments although post-infusion blood pressure was statistically higher in the NVX-108 L group compared to NVX-108 H and Saline groups. Arterial blood gases, methemoglob in and lactate were not different from baseline or among groups. No adverse events were observed at either dose of NVX-108. In conclusion, neither 0.25 nor 1.0 ml/kg NVX-108 caused vasoconstriction in cerebral pial arterioles of healthy rats nor resulted in blood pressure changes; the compound should be considered for further investigation for TBI therapy.