Relaxation In Mesenteric Arteries Research Articles

Endothelial PAR2 activation evokes resistance artery relaxation.

Protease-activated receptor-1 & -2 (PAR1 and PAR2) are expressed widely in cardiovascular tissues including endothelial and smooth muscle cells. PAR1 and PAR2 may regulate blood pressure via changes in vascular contraction or relaxation mediated by endothelial Ca2+ signaling, but the mechanisms are incompletely understood. By using single-cell Ca2+ imaging across hundreds of endothelial cells in intact blood vessels, we explored PAR-mediated regulation of blood vessel function using PAR1 and PAR2 activators. We show that PAR2 activation evoked multicellular Ca2+ waves that propagated across the endothelium. The PAR2-evoked Ca2+ waves were temporally distinct from those generated by muscarinic receptor activation. PAR2 activated distinct clusters of endothelial cells, and these cells were different from those activated by muscarinic receptor stimulation. These results indicate that distinct cell clusters facilitate spatial segregation of endothelial signal processing. We also demonstrate that PAR2 is a phospholipase C-coupled receptor that evokes Ca2+ release from the IP3 -sensitive store in endothelial cells. A physiological consequence of this PAR2 signaling system is endothelium-dependent relaxation. Conversely, PAR1 activation did not trigger endothelial cell Ca2+ signalingnor relax or contract mesenteric arteries. Neither did PAR1 activators alter the response to PAR2 or muscarinic receptor activation. Collectively, these results suggest that endothelial PAR2 but not PAR1 evokes mesenteric artery relaxation by evoking IP3 -mediated Ca2+ release from the internal store. Sensing mediated by PAR2 receptors is distributed to spatially separated clusters of endothelial cells.

Vanillin Induces Relaxation in Rat Mesenteric Resistance Arteries by Inhibiting Extracellular Ca2+ Influx.

Vanillin is a phenolic aldehyde, which is found in plant species of the Vanilla genus. Although recent studies have suggested that vanillin has various beneficial properties, the effect of vanillin on blood vessels has not been studied well. In the present study, we investigated whether vanillin has vascular effects in rat mesenteric resistance arteries. To examine the vascular effect of vanillin, we measured the isometric tension of arteries using a multi-wire myograph system. After the arteries were pre-contracted with high K+ (70 mM) or phenylephrine (5 µM), vanillin was administered. Vanillin induced concentration-dependent vasodilation. Endothelial denudation or treatment of eNOS inhibitor (L-NNA, 300 μM) did not affect the vasodilation induced by vanillin. Treatment of K+ channel inhibitor (TEA, 10 mM) or sGC inhibitor (ODQ, 10 μM) or COX-2 inhibitor (indomethacin, 10 μM) did not affect the vanillin-induced vasodilation either. The treatment of vanillin decreased the contractile responses induced by Ca2+ addition. Furthermore, vanillin significantly reduced vascular contraction induced by BAY K 8644 (30 nM). Vanillin induced concentration-dependent vascular relaxation in rat mesenteric resistance arteries, which was endothelium-independent. Inhibition of extracellular Ca2+ influx was involved in vanillin-induced vasodilation. Treatment of vanillin reduced phopsho-MLC20 in vascular smooth muscle cells. These results suggest the possibility of vanillin as a potent vasodilatory molecule.

Dynein Coordinates β2-Adrenoceptor-Mediated Relaxation in Normotensive and Hypertensive Rat Mesenteric Arteries.

The voltage-gated potassium channel (Kv)7.4 and Kv7.5 channels contribute to the β-adrenoceptor-mediated vasodilatation. In arteries from hypertensive rodents, the Kv7.4 channel is downregulated and function attenuated, which contributes to the reduced β-adrenoceptor-mediated vasodilatation observed in these arteries. Recently, we showed that disruption of the microtubule network, with colchicine, or inhibition of the microtubule motor protein, dynein, with ciliobrevin D, enhanced the membrane abundance and function of Kv7.4 channels in rat mesenteric arteries. This study aimed to determine whether these pharmacological compounds can improve Kv7.4 function in third-order mesenteric arteries from the spontaneously hypertensive rat, thereby restoring the β-adrenoceptor-mediated vasodilatation. Wire and intravital myography was performed on normotensive and hypertensive male rat mesenteric arteries and immunostaining was performed on isolated smooth muscle cells from the same arteries. Using wire and intravital microscopy, we show that ciliobrevin D enhanced the β-adrenoceptor-mediated vasodilatation by isoprenaline. This effect was inhibited partially by the Kv7 channel blocker linopirdine and was dependent on an increased functional contribution of the β2-adrenoceptor to the isoprenaline-mediated relaxation. In mesenteric arteries from the spontaneously hypertensive rat, ciliobrevin D and colchicine both improved the isoprenaline-mediated vasorelaxation and relaxation to the Kv7.2 -7.5 activator, ML213. Immunostaining confirmed ciliobrevin D enhanced the membrane abundance of Kv7.4. As well as an increase in the function of Kv7.4, the functional changes were associated with an increase in the contribution of β2-adrenoceptor following isoprenaline treatment. Immunostaining experiments showed ciliobrevin D prevented isoprenaline-mediated internalizationof the β2-adrenoceptor. Overall, these data show that colchicine and ciliobrevin D can induce a β2-adrenoceptor-mediated vasodilatation in arteries from the spontaneously hypertensive rat as well as reinstating Kv7.4 channel function.

Abstract 003: Treatment With Ligilactobacillus Improves Gut Permeability And Hypertension In Spontaneously Hypertensive Rats

Emerging evidence provide that a microbial imbalance (dysbiosis) is linked to several diseases including cardiovascular diseases. One of the features in hypertension is increased gut permeability. Supporting this, plasma endotoxin level is increased in hypertension and our previous study indicated that the plasma level of exotoxin from Streptococcus was increased in spontaneously hypertensive rats (SHR). Thus, we hypothesized that improvement of gut permeability in hypertension might be expected to prevent development of hypertension. In this study, we performed a 16S metagenome analysis using a synthetic long read sequencing approach by LoopSeq in feces from 10- to 15-week old SHR and Wistar-Kyoto rat (WKY). The analysis revealed that two difference species of anti-inflammatory microbiome were strongly decreased in SHR compared with WKY (n=4). We focus on the decreased Ligilactobacillus sp. (LAB) in SHR and oral administration of LAB to SHR for 2 weeks. SHR (10-15-week-old) exhibited increased gut permeability in proximal colon (P<0.05, n=4), which was improved by treatment with LAB for 2 weeks (P<0.05, n=4). The expression level of tight junction protein Claudin4 was decreased in SHR proximal colon (SHR vs WKY, 1.0±0.1 vs 1.7±0.2-fold increase to SHR, P<0.05, n=8), which was significantly increased by LAB treatment (SHR vs SHR+LAB, 1.0±0.1 vs 2.8±0.5-fold increase to SHR, P<0.05, n=10). Proximal colon in SHR increased the expression level of TNF-α mRNA (1.0±0.2 vs 0.2±0.1, P<0.05, n=8), which was reversed by LAB (1.0±0.2 vs 0.3±0.1, P<0.05, n=10). The level of mucin 2 was lower in SHR proximal colon (1.0±0.2 vs 4.3±0.8, P<0.05, n=8), which was not reversed by LAB (1.0±0.1 vs 0.8±0.3, n=10). We observed that high blood pressure in SHR was attenuated by treatment with LAB for 2 weeks (194.0±5.9 vs 149.7±4.8 mmHg, P<0.05, n=10). LAB also improved decrease in acetylcholine-induced relaxation in SHR mesenteric artery (P<0.05, n=5-8). We conclude that treatment of SHR with the decreased anti-inflammatory microbiota, LAB improves gut permeability in proximal colon possibly through inhibition of inflammation, resulting in lowering blood pressure. These results suggest that anti-inflammatory microbiome might be a new therapeutic target for hypertension.

Effects of 3-methyladenine, an autophagy inhibitor, on the elevated blood pressure and arterial dysfunction of angiotensin II-induced hypertensive mice

Autophagy is an intracellular degradation system that disassembles cytoplasmic components through autophagosomes fused with lysosomes. Recently, it has been reported that autophagy is associated with cardiovascular diseases, including pulmonary hypertension, atherosclerosis, and myocardial ischemia. However, the involvement of autophagy in hypertension is not well understood. In the present study, we hypothesized that excessive autophagy contributes to the dysfunction of mesenteric arteries in angiotensin II (Ang II)-induced hypertensive mice. Treatment of an autophagy inhibitor, 3-methyladenine (3-MA), reduced the elevated blood pressure and wall thickness, and improved endothelium-dependent relaxation in mesenteric arteries of Ang II-treated mice. The expression levels of autophagy markers, beclin1 and LC3 II, were significantly increased by Ang II infusion, which was reduced by treatment of 3-MA. Furthermore, treatment of 3-MA induced vasodilation in the mesenteric resistance arteries pre-contracted with U46619 or phenylephrine, which was dependent on endothelium. Interestingly, nitric oxide production and phosphorylated endothelial nitric oxide synthase (p-eNOS) at S1177 in the mesenteric arteries of Ang II-treated mice were increased by treatment with 3-MA. In HUVECs, p-eNOS was reduced by Ang II, which was increased by treatment of 3-MA. 3-MA had direct vasodilatory effect on the pre-contracted mesenteric arteries. In cultured vascular smooth muscle cells (VSMCs), Ang II induced increase in beclin1 and LC3 II and decrease in p62, which was reversed by treatment of 3-MA. These results suggest that autophagy inhibition exerts beneficial effects on the dysfunction of mesenteric arteries in hypertension.

Vasorelaxant Effect of Trachelospermi caulis Extract on Rat Mesenteric Resistance Arteries.

Background: Trachelospermi caulis (T. caulis) has been used as a traditional herbal medicine in Asian countries. Although it is well known that T. caulis has beneficial effects, no sufficient research data are available on the cardiovascular effect of T. caulis. We investigated whether T. caulis extract has vascular effects in rat resistance arteries in this study. Methods: To examine whether T. caulis extract affects vascular reactivity, we measured isometric tension of rat mesenteric resistance arteries using a multi-wire myograph system. T. caulis extract was administered after arteries were pre-contracted with high K+ (70 mM) or phenylephrine (5 µM). Vanillin, a single active component of T. caulis, was used to treat mesenteric arteries. Results: T. caulis extract caused vascular relaxation in a concentration-dependent manner, which was endothelium-independent. To further identify the mechanism, we incubated the arteries in Ca2+-free solution containing high K+, followed by a cumulative administration of CaCl2 (0.01–2.0 mM) with or without T. caulis extract (250 µg/mL). The treatment of T. caulis extract decreased contractile responses induced by the addition of Ca2+, which suggested that the extracellular Ca2+ influx was inhibited by the T. caulis extract. Moreover, an active compound of T. caulis extract, vanillin, also induced vasodilation in mesenteric resistance arteries. Conclusion: T. caulis extract and its active compound, vanillin, concentration-dependently induced vascular relaxation in mesenteric resistance arteries. These results suggest that the administration of T. caulis extract could help decrease blood pressure.

Moderate-Intensity Exercise Improves Endothelial Function by Altering Gut Microbiome Composition in Rats Fed a High-Fat Diet.

Obesity changes gut microbial ecology and is related to endothelial dysfunction. Although the correlation between gut microbial ecology and endothelial dysfunction has been studied in obese persons, the underlying mechanisms by which exercise enhances endothelial function in this group remain unclear. This study investigated whether exercise improves endothelial function and alters gut microbiome composition in rats fed a high-fat diet (HFD). Obesity was induced by an HFD for 11 weeks. Whole-body composition and endothelium-dependent relaxation of mesenteric arteries were measured. Blood biochemical tests were performed, and gut microbiomes were characterized by 16S rRNA gene sequencing on an Illumina HiSeq platform. Exercise training for 8 weeks improved body composition in HFD-fed rats. Furthermore, compared with the untrained/HFD group, aerobic exercise significantly increased acetylcholine-induced, endothelium-dependent relaxation in mesenteric arteries (P < 0.05) and circulating vascular endothelial growth factor levels (P < 0.01) and decreased circulating C-reactive protein levels (P < 0.05). In addition, exercise and HFD resulted in alterations in the composition of the gut microbiome; exercise reduced the relative abundance of Clostridiales and Romboutsia. Moreover, 12 species of bacteria, including Romboutsia, were significantly associated with parameters of endothelial function in the overall sample. These results suggest that aerobic exercise enhances endothelial function in HFD-fed rats by altering the composition of the gut microbiota. These findings provide new insights on the application of physical exercise for improving endothelial function in obese persons.

Modulation of Vasomotor Function by Perivascular Adipose Tissue of Renal Artery Depends on Severity of Arterial Dysfunction to Nitric Oxide and Severity of Metabolic Parameters.

Perivascular adipose tissue (PVAT) enhances vascular relaxation of mesenteric arteries in SHRSP.Z-Leprfa/IzmDmcr rats (SPZF), a metabolic syndrome model. We investigated and compared the effects of PVAT on the renal artery in SPZF with those on SHR/NDmcr-cp rats (CP). Renal arteries with and without PVAT were isolated from 23-week-old SPZF and CP. The effects of PVAT on acetylcholine- and nitroprusside-induced relaxation were examined using bioassays with phenylephrine-contracted arterial rings. Acetylcholine-induced relaxations without PVAT in SPZF and CP were 0.7- and 0.5-times lower in females than in males, respectively. In the presence of PVAT, acetylcholine-induced relaxations increased 1.4- and 2-times in male and female CP, respectively, but did not differ in SPZF. Nitroprusside-induced relaxation with and without PVAT was 0.7-times lower in female than in male SPZF but did not differ in CP. Angiotensin-II type-1 receptor (AT1R)/AT1R-associated protein mRNA ratios were lower in CP than in the SPZF and negatively correlated with the difference in arterial relaxation with and without PVAT. The effects of renal artery PVAT differed between the SPZF and CP groups. Higher levels of enhanced AT1R activity in SPZF PVAT may drive these differences by impairing the vascular smooth muscle responses to nitric oxide.

17β-Estradiol Treatment Improves Acetylcholine-Induced Relaxation of Mesenteric Arteries in Ovariectomized UC Davis Type 2 Diabetes Mellitus Rats in Prediabetic State.

We recently reported sex differences in mesenteric arterial function of the UC Davis type-2 diabetes mellitus (UCD-T2DM) rats as early as the prediabetic state. We reported that mesenteric arteries (MA) from prediabetic male rats exhibited a greater impairment compared to that in prediabetic females. However, when females became diabetic, they exhibited a greater vascular dysfunction than males. Thus, the aim of this study was to investigate whether the female sex hormone, estrogen preserves mesenteric arterial vasorelaxation in UCD-T2DM female rats at an early prediabetic state. Age-matched female Sprague Dawley and prediabetic (PD) UCD-T2DM rats were ovariectomized (OVX) and subcutaneously implanted with either placebo or 17β-estradiol (E2, 1.5 mg) pellets for 45 days. We assessed the contribution of endothelium-derived relaxing factors (EDRF) to acetylcholine (ACh)-induced vasorelaxation, using pharmacological inhibitors. Responses to sodium nitroprusside (SNP) and phenylephrine (PE) were also measured. Additionally, metabolic parameters and expression of some targets associated with vascular and insulin signaling were determined. We demonstrated that the responses to ACh and SNP were severely impaired in the prediabetic state (PD OVX) rats, while E2 treatment restored vasorelaxation in the PD OVX + E2. Moreover, the responses to PE was significantly enhanced in MA of PD OVX groups, regardless of placebo or E2 treatment. Overall, our data suggest that 1) the impairment of ACh responses in PD OVX rats may, in part, result from the elevated contractile responses to PE, loss of contribution of endothelium-dependent hyperpolarization (EDH) to vasorelaxation, and a decreased sensitivity of MA to nitric oxide (NO), and 2) the basis for the protective effects of E2 may be partly attributed to the elevation of the NO contribution to vasorelaxation and its interaction with MA as well as potential improvement of insulin signaling. Here, we provide the first evidence of the role of E2 in protecting MA from early vascular dysfunction in prediabetic female rats.

Small extracellular vesicles from plasma of women with preeclampsia increase myogenic tone and decrease endothelium-dependent relaxation of mouse mesenteric arteries.

Preeclampsia (PE) is a common syndrome of pregnancy, characterized by new-onset hypertension and proteinuria after gestational week 20, or new onset of hypertension and significant end-organ dysfunction. In the worst cases, it can threaten the survival of both mother and baby. Extracellular vesicles (EVs) are lipid-bilayer nanoparticles released from cells. They are involved in cell-cell communication and transport of diverse cargo molecules. Small extracellular vesicles (sEVs, exosomes) are defined by their size and biogenesis within the endocytic compartment of the cell or reverse budding of the plasma membrane. The function of circulating gestational EVs, released from maternal organs or the placenta, remains to be explored. Here, we focused on sEVs that circulate in the maternal blood in the third trimester of human pregnancy and hypothesized that sEVs from pregnant women with PE play a role in regulation of vessel tone. When compared to sEVs from women with uncomplicated pregnancies, ex vivo exposure of isolated mouse mesenteric arteries to sEVs purified from the plasma of pregnant women with PE led to constriction in response to intraluminal pressure. This effect was not observed using microvesicles from the plasma of women with PE or using PE plasma that was depleted of EVs. Blood vessels exposed to sEVs from women with PE were also more resistant to methacholine-stimulated relaxation. Immunofluorescence microscopy confirmed the presence of sEVs within the vessel wall. Together, these data support the notion that circulating sEVs from pregnant women play a role in the regulation of arterial tone.

Raspberry Supplementation Attenuates Blood Pressure and Improves Mesenteric Artery Relaxation in a Nitric Oxide-Independent Mechanism in Angiotensin II-Treated Rats

ObjectivesBerries are rich in polyphenols with strong antioxidant capacity. They have been suggested to exert antihypertensive properties, which may be due to an increase in the levels of endothelial nitric oxide (NO), a potent vasodilator. Thus, the objective of this study was to evaluate the effects of raspberry supplementation on vascular function in angiotensin (Ang) II-treated rats. MethodsEight-week-old Sprague-Dawley rats (n = 8/group) were fed an AIN-93M diet (control, Ang II, and Losartan (LOS; 20 mg/kg/BW) groups) or AIN-93M diet supplemented with 10% w/w freeze-dried raspberry (RB + Ang II) for 7 weeks. At week 5, rats were implanted with subcutaneous osmotic minipumps that delivered 0.9% saline (CON) or Ang II (270 ng/kg BW/min) for an additional 3 weeks. Blood pressure (BP) was measured using tail-cuff plethysmography (n = 7–8/group). First order mesenteric arteries were isolated for vascular relaxation measurements following constriction by U46619 (1 μM) in a wire myograph (n = 3–8/group). Data were found to be normal and analyzed using ANOVA or mixed-model where appropriate. ResultsAng II group showed a significant increase in systolic BP (SBP) (+26.8%, p = 0.002) 2 weeks after implantation as compared to baseline, while CON, RB + Ang II, and LOS groups showed no differences. Final SBP was significantly lower in the CON (–26.4%, p = 0.01), LOS (–28.3%, p = 0.0005), and RB + Ang II (–20.8%, p = 0.03) groups compared to Ang II group. Relaxation was significantly inhibited in the Ang II group in the presence of acetylcholine after incubation with L-NAME (100 μM), a NO synthase inhibitor, when compared to LOS (p = 0.0003) and RB + Ang II (p = 0.0004) groups. Similarly, relaxation in the Ang II group was significantly inhibited after incubation with ODQ (10 μM), a sGC inhibitor, when compared to CON (p = 0.0002), LOS (p = 0.0004), and RB + Ang II (p = 0.0001) group. No significant changes in vessel relaxation were observed after incubation with LY294002 (20 μM), a PI3K inhibitor. ConclusionsWe demonstrate that RB supplementation attenuated Ang II-induced hypertension and abolished the inhibitory effects of L-NAME and ODQ. This suggests that RB may exert its vasodilatory effects in a NO-independent manner. Funding SourcesThe Agriculture and Food Research Initiative (grant no. 2019–67,017-29,257/project accession no. 1,018,642) from the USDA National Institute of Food and Agriculture.

Endothelial Cell Mineralocorticoid Receptor (ECMR) deletion improves fetal growth and vascular function in the RUPP mouse model of preeclampsia

Preeclampsia (PE), a hypertensive disorder of pregnancy, induces adverse pregnancy outcomes including fetal growth restriction (FGR) and afflicts ~5‐10% of pregnancies. Compelling clinical data indicate that PE risk and severity in pregnant women is strongly associated with endothelial dysfunction. Previous work by our group shows that deletion of endothelial cell mineralocorticoid receptors (ECMR) improves endothelial function in premenopausal hypertensive female mice, however whether ECMR promotes endothelial dysfunction in PE is unknown. We tested the hypothesis that ECMR deletion improves FGR and endothelial function in a mouse model of PE, the reduced uterine perfusion pressure (RUPP) mouse. Pregnant ECMR‐intact (WT) and ECMR‐deficient (KO) mice were randomized to RUPP or sham surgery on GD13 and sacrificed on GD18 (WT Sham=6, WT RUPP=5, KO Sham=7, KO RUPP=6). Vascular function was measured via wire myography on 2nd order mesenteric arteries. Conscious blood pressure was measured by radiotelemetry throughout pregnancy in a subset of mice. RUPP surgery did not decrease maternal weights in either WT or KO mice (P&gt;0.05) at GD18. RUPP surgery significantly decreased GD18 pup weight, a measure of FGR, (726±20mg WT+Sham vs 660±11mg WT+RUPP, *P&lt;0.05) and trended to decrease placental efficiency (pup/placenta weight) (8.23±0.23 WT+Sham vs 7.45±0.22 WT+RUPP P=0.13) in WT pregnant mice. ECMR deletion protected pregnant mice both from RUPP‐induced reductions in pup weight (710±20mg KO+Sham vs 690±20mg KO+RUPP) and decreases in placental efficiency (7.63±0.19 KO+Sham vs 7.76±0.31 KO+RUPP). RUPP reduced acetylcholine‐mediated relaxation (10‐9‐3x10‐5 M) in mesenteric arteries of WT pregnant mice but not KO pregnant mice (*P&lt;0.05, 2‐way ANOVA, repeated measures), indicating RUPP induced endothelial dysfunction in WT mice only. Preincubation with nitric oxide (NO) synthase inhibitor LNAME ablated differences in Ach‐mediated relaxation between WT+sham and WT+RUPP mice (P&gt;0.05), indicating that reductions in endothelial function in WT+RUPP mice were mediated by reduced NO. However, ECMR deletion as a variable increased Ach‐mediated relaxation in the presence of LNAME in both Sham and RUPP mice (*P&lt;0.05). Therefore, ECMR deletion may have protected pregnant mice from endothelial dysfunction via a non‐NO mechanism. RUPP did not increase a‐receptor agonist phenylephrine (Phe)‐induced constriction in WT mice, however, ECMR deletion as a variable decreased Phe‐mediated constriction independent of RUPP surgery indicating that ECMR deletion reduces smooth muscle‐mediated vasoconstriction in pregnant sham and RUPP mice. These data indicate that specific deletion of ECMR improves vascular function to increase vasodilation and prevent constriction in PE pregnancy. Preliminary data further indicates that systolic blood pressure increased from GD8‐18 (pre‐post RUPP) in WT+RUPP but decreased in KO+RUPP pregnant mice (2.7 vs ‐8.6 DmmHg, respectively, N=1). Lastly RUPP surgery trended to increase plasma levels of the ECMR agonist corticosterone in WT pregnant mice (2.03±0.4pg/ml WT+Sham vs 3.33±0.95pg/ml WT+RUPP). Collectively, these data indicate that ECMR activation promotes FGR, vascular dysfunction and hypertension in the RUPP model of PE which may lead to clinical studies for the consideration of specific MR antagonists as therapeutic options for PE pregnancies.

Sex‐Dependent Changes in Vascular Function in Adult Rats Following Prenatal Exposure to Methamphetamine

BackgroundPrenatal exposure to central nervous system stimulants such as cocaine, nicotine, and caffeine alters cardiovascular function in adult offspring. Methamphetamine use during pregnancy is associated with negative consequences in the offspring. However, most studies of methamphetamine use during pregnancy have focused on behavioral and neurological outcomes. Relatively little is known regarding the impact of prenatal methamphetamine exposure on the adult cardiovascular system. This study examined the hypothesis that methamphetamine use during pregnancy alters contractile function of the vasculature in the adult offspring. We also investigated the role of perivascular adipose tissue (PVAT) in methamphetamine‐induced changes in vascular function.MethodsPregnant female rats received daily injections of saline or methamphetamine (5 mg/kg) throughout gestation. Agonist‐induced contraction and relaxation responses were measured in aortas of 5 month old offspring in the presence and absence of perivascular adipose tissue.ResultsPrenatal methamphetamine significantly attenuated acetylcholine‐induced relaxation in male (but not female) aortas when PVAT remained intact. However, prenatal methamphetamine had no impact on acetylcholine‐induced relaxation when PVAT was removed. Nitroprusside‐induced relaxation was unaffected by prenatal methamphetamine. Prenatal exposure to methamphetamine had no impact on acetylcholine‐induced relaxation in third order mesenteric arteries of male or female offspring regardless of the presence of PVAT. Angiotensin II‐induced contractile responses were significantly potentiated in male (but not female) aortas regardless of the presence of PVAT. This effect was abolished by L‐Nitro arginine methyl ester (L‐NAME). Contractile responses to phenylephrine and serotonin were unaffected in both male and female aortas. Basal blood pressure in adult male and female offspring was also unaffected by prenatal exposure to methamphetamine.ConclusionsPrenatal exposure to methamphetamine sex‐dependently alters PVAT function in the aorta. Methamphetamine also enhances angiotensin II‐induced contraction through a mechanism that involves suppression of nitric oxide signaling. These data provide evidence that methamphetamine use during pregnancy induces sex‐dependent changes in the vasculature that may increase the risk of cardiovascular disease in adult offspring.

Perivascular Adipose Tissue Compensation for Endothelial Dysfunction in the Superior Mesenteric Artery of Female SHRSP.Z-Leprfa/IzmDmcr Rats

Regulation of arterial tone by perivascular adipose tissue (PVAT) differs between sexes. In male SHRSP.Z-Lepr^fa/IzmDmcr rats (SHRSP.ZF), PVAT exerts a compensatory relaxation effect for the loss of endothelium-mediated vasorelaxation, which occurs during the early stages of metabolic syndrome. However, this effect deteriorates by 23 weeks of age. Here, therefore, we compared the effects of PVAT in female and male SHRSP.ZF. Acetylcholine-induced relaxation in superior mesenteric artery without PVAT did not differ between 23-week-old females and males. However, the presence of PVAT enhanced relaxation in 23-week-old females, but not in males. The mRNA levels of angiotensin II type 1 receptor (AT1R) in PVAT did not differ between sexes, but AT1R-associated protein (ATRAP) and apelin levels were higher in females than in males. We observed a positive relationship between differences in artery relaxation with and without PVAT and ATRAP or apelin mRNA levels. In 30-week-old females, PVAT-enhanced relaxation disappeared, and mRNA levels of AT1R increased, while apelin levels decreased compared to 23-week-old females. These results demonstrated that in SHRSP.ZF, PVAT compensation for endothelium dysfunction extended to older ages in females than in males. Apelin and AT1R/ATRAP expression in PVAT may be predictors of favorable effects.

High Salt Upregulates Ca2+-Sensing Receptor Expression and Ca2+-Induced Relaxation of Contracted Mesenteric Arteries from Dahl Salt-Sensitive Rats.

High Ca2+ lowers blood pressure in hypertension, but the mechanism is not clear. The missing link may be the perivascular sensory nerve Ca2+-sensing receptor (CaSR) that mediates a vasodilator system after activation by interstitial Ca2+ Our results show that high salt increased CaSR expression in mesenteric arteries as well as Ca2+ relaxation of contracted mesenteric arteries from salt-sensitive (SS) rats. The CaSR was expressed as a doublet (≈120-150 kDa) in arteries from animals fed a high-salt diet for 1-4 weeks. The higher molecular weight glycosylated protein increased in arteries from SS animals; however, expression of the low molecular mass high-mannose protein decreased over 4 weeks of feeding the diet. In tissues from salt-resistant (SR) rats, the diet decreased CaSR expression after 4 weeks. Ca2+ relaxation of mesenteric arteries under phenylephrine tone increased in SS rats but decreased in arteries from SR rats fed the high-salt diet. Ca2+-activated K+ channels have a larger role in Ca2+ relaxation of arteries in SR than SS rats. The data suggest that high salt epigenetically regulates the receptor at the translational level in vivo and that the in vitro effect of Ca2+ is on receptor trafficking and signaling. In conclusion, upregulated expression of the CaSR in salt sensitivity increased receptor-mediated vascular relaxation. These findings show that CaSR signaling may compensate for changes in the vasculature in salt-sensitive hypertension. SIGNIFICANCE STATEMENT: The perivascular sensory nerve Ca2+-sensing receptor (CaSR) mediates Ca2+ relaxation of isolated mesenteric arteries under tension. This receptor may therefore play a significant role in relaxation of resistance arteries in vivo, thus explaining the blood pressure-lowering effect of dietary Ca2+. The present studies describe the effect of high salt-induced upregulation of the CaSR in salt-sensitive rats and the roles played by Ca2+-activated K+ channels and nitric oxide in Ca2+ responses.

Prenatal Exposure to Methamphetamine Causes Vascular Dysfunction in Adult Male Rat Offspring.

Methamphetamine use during pregnancy can have negative consequences on the offspring. However, most studies investigating the impact of prenatal exposure to methamphetamine have focused on behavioral and neurological outcomes. Relatively little is known regarding the impact of prenatal methamphetamine on the adult cardiovascular system. This study investigated the impact of chronic fetal exposure to methamphetamine on vascular function in adult offspring. Pregnant female rats received daily saline or methamphetamine (5 mg/kg) injections starting on gestational day 1 and continuing until the pups were born. Vascular function was assessed in 5 month old offspring. Prenatal methamphetamine significantly decreased both the efficacy and potency of acetylcholine-induced relaxation in isolated male (but not female) aortas when perivascular adipose tissue (PVAT) remained intact. However, prenatal methamphetamine had no impact on acetylcholine-induced relaxation when PVAT was removed. Nitroprusside-induced relaxation of the aorta was unaffected by prenatal methamphetamine. Angiotensin II-induced contractile responses were significantly potentiated in male (but not female) aortas regardless of the presence of PVAT. This effect was reversed by L-nitro arginine methyl ester (L-NAME). Serotonin- and phenylephrine-induced contraction were unaffected by prenatal methamphetamine. Prenatal methamphetamine had no impact on acetylcholine-induced relaxation of third order mesenteric arteries and no effect on basal blood pressure. These data provide evidence that prenatal exposure to methamphetamine sex-dependently alters vasomotor function in the vasculature and may increase the risk of developing vascular disorders later in adult life.

Abstract 12763: High Innate Aerobic Capacity Protects Against Angiotensin II-Induced Hypertension and Vascular Remodeling

Low aerobic capacity is associated with increased cardiovascular morbidity and mortality. A heterogeneous rat population selectively bred to segregate genes that sustain innate aerobic exercise capacity, low capacity runners (LCR) show impaired cardiac function in old age and premature death versus high capacity runners (HCR). Here we investigated whether high innate aerobic capacity offers early protection against cardiovascular disease by examining the hypertensive response of 7-month-old LCR and HCR rats to angiotensin II (Ang II, 150 μg/kg/min for 14 days). LCR and HCR rats showed no difference in basal blood pressure (105 mmHg) or vessel wall morphology, but in LCRs the mean arterial blood pressure increased to 145 mmHg versus 125 mmHg in HCRs. Congruently, LCR rats had increased aortic medial thickness and fibrosis. Aortic cytosolic and mitochondrial superoxide levels were also significantly increased in LCRs. Vascular reactivity experiments revealed impaired endothelium-dependent and -independent relaxation of mesenteric arteries of LCRs. Ang II infusion significantly increased the infiltration of macrophages into the aortic adventitia of LCRs as determined by CD68 and Mac-3 staining. Gene expression analysis by qPCR showed that markers of classically-activated macrophages CD86 and TNF-α were unchanged in rat strains but markers of alternatively-activated macrophages CD163 and Arg-1 were upregulated only in LCRs. Plasma IL-6, RANTES, CCL2 and Ang II and aortic VCAM1 levels were elevated only in LCRs whereas no difference in eNOS levels was observed between the rat strains. Aortic sections of LCRs showed increased senescence in adventitia whereas vascular smooth muscle cells (VSMC) isolated from LCR rats were more susceptible to Ang II-induced senescence. LCRs also showed increased cardiac hypertrophy and fibrosis. These data support the “outside-in” hypothesis, in which vascular inflammation initiated in adventitia progresses inwards to media and intima, causing VSMC and endothelial dysfunction, and that fibroblasts play a critical role in Ang II-induced hypertension and vascular remodeling under low innate aerobic capacity conditions, whereas high innate aerobic capacity is potentially protective against hypertension.

Abstract 03: Ablation Of Endothelial Dysfunction Improves Blood Pressure And Fetal Growth Restriction In A Mouse Model Of Preeclampsia-like Hyperleptinemia

Placental ischemia, an initiating event of preeclampsia (PE), increases plasma leptin levels. We recently developed a model of midgestation (gestation day (GD)11-18) leptin infusion mimicking the midgestation rise in leptin levels of PE patients. Our previous work demonstrates that deletion of endothelial mineralocorticoid receptors (ECMR) improves markers of vascular dysfunction in leptin-infused female mice. We hypothesized vascular function improvement with ECMR deletion ablates hypertension and fetal growth restriction in pregnant leptin-infused mice. Pregnant ECMR +/+ (WT) and ECMR -/- (KO) mice were infused with vehicle or leptin by osmotic pump (lep, 0.9mg/kg/day, s.c.) on GD11-18 while implanted with radiotelemeters for conscious blood pressure (BP) measurement and wire myography of thoracic aorta and 2 nd order mesenteric arteries at GD18 (*=P&lt;0.05). Leptin infusion did not decrease maternal weight in any groups. Leptin decreased pup weight (0.86±0.04g WT vs 0.52±0.11 WT+lep*) and placental efficiency (pup/placenta ratio) (9.7±0.7 WT vs 7.9±0.6 WT+lep*) in WT pregnant mice. ECMR deletion prevented leptin-mediated decreases in pup weight (0.91±0.06g KO vs 1.0±0.07 KO+lep) and placental efficiency (9.6±0.5 KO vs 9.4±1.2 KO+lep). Mean arterial pressure (BP) increased in leptin-infused WT (102±3mmHg WT vs 120±12 WT+lep*), but not KO (107±2 KO vs 106±8 KO+lep), mice from GD11-18. Leptin infusion reduced acetylcholine-mediated relaxation in both aorta and mesenteric arteries of WT* and constriction to KCl in mesenteric arteries in WT*, but not KO, pregnant mice (2-way ANOVA, repeated measures). Leptin increased plasma endothelin-1 (ET-1, 1.1±0.3 pg/ml WT vs 4.4±1.8 WT+lep*), placental mRNA expression of prepro-ET-1 (1.9±0.3-fold change from WT*) and ET-1 converting enzyme-1 (1.6±0.3-fold change from WT*) in pregnant WT mice. Leptin infusion also increased adrenal aldosterone-synthase protein (1.4±0.4 WT ratio/β actin vs 3.2±0.3 WT+lep*) and angiotensin II type 1 receptor b (3.5±0.8-fold change from WT*) mRNA in pregnant mice. Collectively, these data indicate that leptin infusion induces hypertension and fetal growth restriction in pregnant mice due to vascular dysfunction and increased ECMR activation in pregnant mice.

Vasodilation and blood pressure-lowering effect mediated by 5,6-EEQ lactone in 5/6 nephrectomy hypertensive rats

Microvascular dysfunction is a key contributor to vascular hypertension, one of the most common chronic diseases in the world. Microvascular dysfunction leads to the loss of nitric oxide-mediated endothelial dilation and the subsequent compensatory function of endothelium-derived hyperpolarizing (EDH) factors in the regulation of vascular tone. Previously, we showed that lactone metabolite derived from arachidonic acid induces endothelial-dependent vasodilation in isolated human microvessels. Based on structural similarities, we hypothesize that additional lactone metabolites formed from eicosapentaenoic fatty acid (EPA) may bear EDH properties. AimTo elucidate the vasodilatory and blood pressure (BP)-reducing characteristics of the 5,6-EEQ (5,6-epoxyeicosatetraenoic acids) lactone (EPA-L) in hypertensive 5/6 nephrectomy (5/6Nx) rats. Methods5/6Nx hypertensive rats intravenously administrated with EPA-L for five days. BP, blood and urine chemistry, and kidney function were detected and analyzed. Vascular dilation was detected using a pressure myograph with or without Ca2+ − activated K+ (KCa) endothelial channel inhibitors. KCNN3 and KCNN4 gene expression (mRNA) detected in mesenteric arteries from 5/6Nx and NT rats. ResultsEPA-L administration to 5/6Nx rats significantly (p < 0.05) reduced BP and heart rate without affecting kidney function. 5/6Nx rat mesenteric arterioles exhibited a lower dilation response to acetylcholine (10-7 mol/l) than normotensive (NT) vessels, while EPA-L administration restored the vessel relaxation response. The EPA-L-driven relaxation of mesenteric arteries was significantly reduced by pretreatment with TRAM-34 and apamin. However, KCa channel expression did not significantly differ between 5/6Nx and NT mesenteric arteries. ConclusionEPA-L reduces BP by improving microvessel dilation involving calcium-dependent potassium endothelial channels.

Down-regulation of AMPK/PPARδ signalling promotes endoplasmic reticulum stress-induced endothelial dysfunction in adult rat offspring exposed to maternal diabetes.

Exposure to maternal diabetes is associated with increased prevalence of hypertension in the offspring. The mechanisms underlying the prenatal programming of hypertension remain unclear. Because endoplasmic reticulum (ER) stress plays a key role in vascular endothelial dysfunction in hypertension, we investigated whether aberrant ER stress causes endothelial dysfunction and high blood pressure in the offspring of dams with diabetes. Pregnant Sprague-Dawley rats were intraperitoneally injected with streptozotocin (35 mg/kg) or citrate buffer at Day 0 of gestation. Compared with control mother offspring (CMO), the diabetic mother offspring (DMO) had higher blood pressure and impaired endothelium-dependent relaxation in mesenteric arteries, accompanied by decreased AMPK phosphorylation and PPARδ expression, increased ER stress markers, and reactive oxygen species (ROS) levels. The inhibition of ER stress reversed these aberrant changes in DMO. Ex vivo treatment of mesenteric arteries with an AMPK agonist (A769662) or a PPARδ agonist (GW1516) improved the impaired EDR in DMO and reversed the tunicamycin-induced ER stress, ROS production, and EDR impairment in mesenteric arteries from CMO. The effects of A769662 were abolished by co-treatment with GSK0660 (PPARδ antagonist), whereas the effects of GW1516 were unaffected by Compound C (AMPK inhibitor). These results suggest an abnormal foetal programming of vascular endothelial function in offspring of rats with maternal diabetes that is associated with increased ER stress, which can be ascribed to down-regulation of AMPK/PPARδ signalling cascade.