Angiotensin II (Ang II) is a natural mammalian hormone that has been described to exhibit antiplasmodial activity therefore constituting a promising alternative for the treatment of malaria. Despite its promise, the development of Ang II as an antimalarial is limited by its potent induction of vasoconstriction and its rapid degradation within minutes. Here, we used peptide design to perform targeted chemical modifications to Ang II to generate conformationally restricted (disulfide-crosslinked) peptide derivatives with suppressed vasoconstrictor activity and increased stability. Designed constrained peptides were synthesized chemically and then tested for antiplasmodial activity. Two lead constrained peptides were identified (i.e., peptides 1 and 2), each composed of 10 amino acid residues. These peptides exhibited very promising activity in both our Plasmodium gallinaceum (>80%) and Plasmodium falciparum (>40%) models, an activity that was equivalent to that of Ang II, and led to complete suppression of vasoconstriction. In addition, peptide 5 exhibited selective activity towards the pre-erythrocytic stage (98% of activity against P. gallinaceum), thus suggesting that it may be possible to design peptides that target specific stages of the malaria life cycle. The Ang II derived stable scaffolds presented here may provide the basis for development of a new generation of peptide-based drugs for the treatment of malaria.

Cerebrovascular lesions seen as white matter hyperintensity in MRI of elderly population caused due to micro-infracts and micro-bleeds contributes to vascular dementia. Such vascular insult caused by impairment in blood flow to specific area in brain involving small vessels can gradually worsen the pathology leading to cognitive deficits. In the present study we developed a transient model of vaso-constriction to study the impact of such pathology by bilateral injection of ET-1 (Endothelin-1; a 21 amino acid vasoconstricting peptide) into lateral ventricles of C57 mice. The impediment in cerebral blood flow decreased CD31 expression in endothelial cells lining the blood vessels around the hippocampal region, leading to memory deficits after 7 days. Activity dependent protein translation, critical for synaptic plasticity was absent in synaptoneurosomes prepared from hippocampal tissue. Further, Akt1- mTOR signaling cascade was downregulated indicating the possible cause for loss of activity dependent protein translation. However, these effects were reversed after 30 days indicating the ephemeral nature of deficits following a single vascular insult. Present study demonstrates that vasoconstriction leading to memory deficit and decline in activity dependent protein translation in hippocampus as a potential molecular mechanism impacting synaptic plasticity.

Non-compressible hemorrhage is the most common preventable cause of death on battlefield and in civilian traumatic injuries. We report the use of microsecond pulses of electric current to induce rapid constriction in femoral and mesenteric arteries and veins in rats. Electrically-induced vasoconstriction could be induced in seconds while blood vessels dilated back to their original size within minutes after stimulation. At higher settings, a blood clotting formed, leading to complete and permanent occlusion of the vessels. The latter regime dramatically decreased the bleeding rate in the injured femoral and mesenteric arteries, with a complete hemorrhage arrest achieved within seconds. The average blood loss from the treated femoral artery during the first minute after injury was about 7 times less than that of a non-treated control. This new treatment modality offers a promising approach to non-damaging control of bleeding during surgery, and to efficient hemorrhage arrest in trauma patients.

We recently reported exacerbated endotoxic signs of neuroinflammation and autonomic defects in offspring of preeclamptic (PE) dams. Here, we investigated whether PE programming similarly modifies hemodynamic and renal vasoconstrictor responsiveness to endotoxemia in PE offspring and whether this interaction is modulated by gestational angiotensin 1-7 (Ang1-7). Preeclampsia was induced by gestational treatment with L-NAME. Adult offspring was challenged with lipopolysaccharides (LPS, 5 mg/kg) and systolic blood pressure (SBP) and renal vasoconstrictions were assessed 4 h later. Male, but not female, offspring of PE rats exhibited SBP elevations that were blunted by LPS. Renal vasoconstrictions induced by angiotensin II (Ang II), but not phenylephrine, were intensified in perfused kidneys of either sex. LPS blunted the heightened Ang II responses in male, but not female, kidneys. While renal expressions of AT1-receptors and angiotensin converting enzyme (ACE) were increased in PE offspring of both sexes, ACE2 was upregulated in female offspring only. These molecular effects were diminished by LPS in male offspring. Gestational Ang1-7 caused sex-unrelated attenuation of phenylephrine vasoconstrictions and preferentially downregulated Ang II responses and AT1-receptor and nuclear factor-kB (NFkB) expressions in females. Together, endotoxemia and Ang1-7 offset in sexually-related manners imbalances in renal vasoconstriction and AT1/ACE/ACE2 signaling in PE offspring.

Reversible cerebral vasoconstriction syndrome (RCVS) is one of the most important differential diagnosis in patients with thunderclap headache (TCH). We aimed to develop a new scoring system for RCVS in patients with TCH. We retrospectively analyzed 72 patients enrolled in the prospective study of TCH conducted in 2015-2016 (derivation set). We identified possible predictors for the diagnosis of RCVS and constructed a prediction model (RCVS-TCH score) using the multivariable logistic regression model. Diagnostic performance was validated to an independent validation set from our headache registry. The derivation set comprised 41 patients with RCVS and 31 with non-RCVS, and the validation set included 253 patients with TCH (165 with RCVS and 88 with non-RCVS). The RCVS-TCH score (range: 0-12) contained four predictors: recurrent TCHs, female sex, triggering factor for TCH (single or multi) and blood pressure surge. The C-index of RCVS-TCH score was 0.929 (95% CI = 0.874-0.984). The RCVS-TCH score ≥ 7 had a sensitivity of 80% and a specificity of 97% in discriminating RCVS from non-RCVS. In the validation set, RCVS-TCH score showed a C-index of 0.861 (95% CI = 0.815-0.908). In our study, the RCVS-TCH showed good performance, which may aid the diagnosis of RCVS among patients with TCH.

Cytochrome P450 (CYP) signalling pathway has been shown to play a vital role in the vasoreactivity of wild type mouse ophthalmic artery. In this study, we determined the expression, vascular responses and potential mechanisms of the CYP-derived arachidonic acid metabolites. The expression of murine CYP (Cyp2c44) and soluble epoxide hydrolase (sEH) in the wild type ophthalmic artery was determined with immunofluorescence, which showed predominant expression of Cyp2c44 in the vascular smooth muscle cells (VSMC), while sEH was found mainly in the endothelium of the wild type ophthalmic artery. Artery of Cyp2c44-/- and sEH-/- mice were used as negative controls. Targeted mass spectrometry-based lipidomics analysis of endogenous epoxide and diols of the wild type artery detected only 14, 15-EET. Vasorelaxant responses of isolated vessels in response to selective pharmacological blockers and agonist were analysed ex vivo. Direct antagonism of epoxyeicosatrienoic acids (EETs) with a selective inhibitor caused partial vasodilation, suggesting that EETs may behave as vasoconstrictors. Exogenous administration of synthetic EET regioisomers significantly constricted the vessels in a concentration-dependent manner, with the strongest responses elicited by 11, 12- and 14, 15-EETs. Our results provide the first experimental evidence that Cyp2c44-derived EETs in the VSMC mediate vasoconstriction of the ophthalmic artery.

Preeclampsia (PE) is a pregnancy syndrome characterized by hypertension and organ damage manifesting after 20 gestational weeks. The etiology is of multifactorial origin, where placental stress causes increased levels of placenta-derived extracellular vesicles (STBEVs) in the maternal circulation, shown to cause inflammation, endothelial activation, vasoconstriction, and anti-angiogenic activity. General endothelial dysfunction is believed to be initiated by endothelial insult during pregnancy that alters vascular function resulting in increased arterial stiffness, cardiac dysfunction, and increased risk of cardiovascular disease later in life. We compared the effect of normal and PE derived STBEVs in vitro on vascular contractility of human subcutaneous arteries using wire myography. Cellular structures of exposed vessels were investigated by transmission electron microscopy. We explored strategies to pharmacologically block the effects of the STBEVs on human vessels. The PE STBEVs caused significantly stronger angiotensin II-mediated contractions and extended structural damage to human subcutaneous arteries compared to normal STBEVs. These negative effects could be reduced by blocking vesicle uptake by endothelial cells, using chlorpromazine or specific antibodies towards the LOX-1 receptor. The therapeutic potential of blocking vesicle uptake should be further explored, to reduce the permanent damage caused on the vasculature during PE pregnancy to prevent future cardiovascular risk.

Vascular dysfunction is an important phenomenon in hypertension. We hypothesized that angiotensin II (AngII) affects transcriptome in the vasculature in a region-specific manner, which may help to identify genes related to vascular dysfunction in AngII-induced hypertension. Mesenteric artery and aortic transcriptome was profiled using Illumina WG-6v2.0 chip in control and AngII infused (490 ng/kg/min) hypertensive mice. Gene set enrichment and leading edge analyses identified Sphingosine kinase 1 (Sphk1) in the highest number of pathways affected by AngII. Sphk1 mRNA, protein and activity were up-regulated in the hypertensive vasculature. Chronic sphingosine-1-phosphate (S1P) infusion resulted in a development of significantly increased vasoconstriction and endothelial dysfunction. AngII-induced hypertension was blunted in Sphk1-/- mice (systolic BP 167 ± 4.2 vs. 180 ± 3.3 mmHg, p < 0.05), which was associated with decreased aortic and mesenteric vasoconstriction in hypertensive Sphk1-/- mice. Pharmacological inhibition of S1P synthesis reduced vasoconstriction of mesenteric arteries. While Sphk1 is important in mediating vasoconstriction in hypertension, Sphk1-/- mice were characterized by enhanced endothelial dysfunction, suggesting a local protective role of Sphk1 in the endothelium. S1P serum level in humans was correlated with endothelial function (arterial tonometry). Thus, vascular transcriptome analysis shows that S1P pathway is critical in the regulation of vascular function in AngII-induced hypertension, although Sphk1 may have opposing roles in the regulation of vasoconstriction and endothelium-dependent vasorelaxation.

Compromised microvascular reactivity underlies many conditions and injuries, but its assessment remains difficult, particularly in low perfusion tissues. In this paper, we develop a new mathematical model for the assessment of vasomodulation in low perfusion settings. A first-order model was developed to approximate changes in T1 relaxation times as a result of vasomodulation. Healthy adult rats (N = 6) were imaged on a 3-Tesla clinical MRI scanner, and vasoactive response was probed on gadofosveset using hypercapnic gases at 20% and 5% CO2 to induce vasoconstriction and vasodilation, respectively. MRI included dynamic 3D T1 mapping and T1-weighted images during gas challenge; heart rate was continuously monitored. Laser Doppler perfusion measurements were performed to corroborate MRI findings. The model was able to identify hypercapnia-mediated vasoconstriction and vasodilation through the partial derivative [Formula: see text]. MRI on animals revealed gradual vasoconstriction in the skeletal muscle bed in response to 20% CO2 followed by gradual vasodilation on transitioning to 5% CO2. These trends were confirmed on laser Doppler perfusion measurements. Our new mathematical model has the potential for detecting microvascular dysfunction that manifests in the early stages across multiple metabolic and ischemic pathologies.

Intravital microscopy of orthotopic lung tissue is technically demanding, especially for repeated investigations. Therefore, we have established a novel approach, which allows non-invasive repetitive in vivo microscopy of ectopic lung tissue in dorsal skinfold chambers. Syngeneic subpleural peripheral lung tissue and autologous endometrium (control) were transplanted onto the striated muscle within dorsal skinfold chambers of C57BL/6 mice. Grafts were analysed by intravital fluorescence microscopy over 14 days. Angiogenesis occurred in the grafts on day 3, as indicated by sinusoidal microvessels on the grafts' edges with very slow blood flow, perifocal oedema, and haemorrhage. By day 10, lung transplants were completely revascularized, exhibited a dense network of microvessels with irregular diameters, chaotic angioarchitecture, and high blood flow. Compared to lung tissue, endometrial grafts contained a structured, glomerulus-like vessel architecture with lower blood flow. Despite missing ventilation, hypoxic vasoconstriction of the lung tissue arterioles occurred. In contrast, endometrium tissue arterioles dilated during hypoxia and constricted in hyperoxia. This demonstrates that ectopic lung grafts keep their ability for organ-specific hypoxic vasoconstriction. These findings indicate that our approach is suitable for repetitive in vivo pulmonary microcirculation analyses. The high blood flow and hypoxia-induced vasoconstriction in lung grafts suggest a physiological intrinsic vasoregulation independent of the recipient tissue.

Vascular dysfunction predisposes to cardiovascular complications of metabolic syndrome (MetS). The current study investigated the mechanism(s) of curcumin's (CUR) protective effect against vascular reactivity irregularities in MetS. MetS was induced by feeding rats on high fructose high salt diet. Tension studies were undertaken in aortic rings to assess the influence of CUR on vasoconstrictor or vasorelaxant responses. The effect on advanced glycation endproducts (AGEs) was studied by incubating aortic tissues with methylglyoxal, the AGEs precursor, in the absence and presence of CUR. In addition, CUR effects on in-vitro generation of AGEs and diphenyl-2-picrylhydrazyl (DPPH) free radicals were studied. The incubation with CUR for 1 hr produced significant and concentration-dependent alleviation of the exaggerated vasoconstriction observed in aortas isolated from MetS, however failed to improve the concomitant attenuation of vasodilatory responses to ACh in PE-precontracted aortas. By contrast, CUR caused direct concentration-dependent vasodilations of precontracted aortas, effects that were blunted after nitric oxide synthase inhibition by L-NAME. Similar to its effects in MetS aortas, CUR alleviated exaggerated PE vasoconstriction but did not affect impaired ACh vasodilations in AGEs-exposed aortas. In addition, CUR showed significant dose-dependent DPPH free radicals scavenging activity and inhibited both MG and fructose induced AGEs formation at the level of protein oxidation step as evident from the effect on dityrosine and N-formylkyramine. CUR alleviates exaggerated vasoconstriction in MetS through interfering with AGEs formation and AGEs-induced vascular injury. Free radical scavenging and direct vasodilatory activities could also participate in the advantageous vascular actions of CUR.

Hutchinson-Gilford Progeria Syndrome (HGPS) is a rare, fatal genetic disease that accelerates atherosclerosis. With a limited pool of HGPS patients, clinical trials face unique challenges and require reliable preclinical testing. We previously reported a 3D tissue engineered blood vessel (TEBV) microphysiological system fabricated with iPSC-derived vascular cells from HGPS patients. HGPS TEBVs exhibit features of HGPS atherosclerosis including loss of smooth muscle cells, reduced vasoactivity, excess extracellular matrix (ECM) deposition, inflammatory marker expression, and calcification. We tested the effects of HGPS therapeutics Lonafarnib and Everolimus separately and together, currently in Phase I/II clinical trial, on HGPS TEBVs. Everolimus decreased reactive oxygen species levels, increased proliferation, reduced DNA damage in HGPS vascular cells, and improved vasoconstriction in HGPS TEBVs. Lonafarnib improved shear stress response of HGPS iPSC-derived endothelial cells (viECs) and reduced ECM deposition, inflammation, and calcification in HGPS TEBVs. Combination treatment with Lonafarnib and Everolimus produced additional benefits such as improved endothelial and smooth muscle marker expression and reduced apoptosis, as well as increased TEBV vasoconstriction and vasodilation. These results suggest that a combined trial of both drugs may provide cardiovascular benefits beyond Lonafarnib, if the Everolimus dose can be tolerated.

Capillary leakage syndrome, vasomotor disturbances and gut atony are common clinical problems in intensive care medicine. Various inflammatory mediators and signalling pathways are involved in these pathophysiological alterations among them platelet-activating factor (PAF). The related signalling mechanisms of the PAF-induced dysfunctions are only poorly understood. Here we used the model of the isolated perfused rat small intestine to analyse the role of calcium (using calcium deprivation, IP-receptor blockade (2-APB)), cAMP (PDE-inhibition plus AC activator), myosin light chain kinase (inhibitor ML-7) and Rho-kinase (inhibitor Y27632) in the following PAF-induced malfunctions: vasoconstriction, capillary and mucosal leakage, oedema formation, malabsorption and atony. Among these, the PAF-induced vasoconstriction and hyperpermeability appear to be governed by similar mechanisms that involve IP3 receptors, extracellular calcium and the Rho-kinase. Our findings further suggest that cAMP-elevating treatments - while effective against hypertension and oedema - bear the risk of dysmotility and reduced nutrient uptake. Agents such as 2-APB or Y27632, on the other hand, showed no negative side effects and improved most of the PAF-induced malfunctions suggesting that their therapeutic usefulness should be explored.

Vasoconstriction during tachyarrhythmia contributes to maintenance of arterial pressure (AP) by decreasing peripheral blood flow. This cross-sectional observational study aimed to ascertain whether the relationship between peripheral blood flow and AP recovery occurs during both paroxysmal supraventricular (PSVT, n = 19) and ventricular tachycardias (VT, n = 17). Peripheral blood flow was evaluated using forearm tissue oxygen index (TOI), and mean AP (MAP) was measured using a catheter inserted in the brachial or femoral artery during an electrophysiological study. PSVT and VT rapidly decreased MAP with a comparable heart rate (P = 0.194). MAP recovered to the baseline level at 40 s from PSVT onset, but not VT. The forearm TOI decreased during both tachyarrhythmias (P ≤ 0.029). The TOI response was correlated with MAPrecovery (i.e., MAP recovery from the initial rapid decrease) at 20-60 s from PSVT onset (r = -- 0.652 to - 0.814, P ≤ 0.0298); however, this association was not observed during VT. These findings persisted even after excluding patients who had taken vasoactive drugs. Thus, restricting peripheral blood flow was associated with MAP recovery during PSVT, but not VT. This indicates that AP recovery depends on the type of tachyarrhythmia: different cardiac output and/or vasoconstriction ability during tachyarrhythmia.

Auricular vasomotor responses are considered to be signs of clinical conditions including migraine. The mechanisms of auricular vasomotor control are still debatable. This study aimed at investigating perivascular co-transmitters of vasomotor control in the auricle. Another aim was to provide three-dimensional arterial maps of the auricle, as a proxy of periarterial autonomic innervation. Twelve paired human auricles were used to visualize the arteries following Spalteholz clearing and μ-CT-based reconstruction. Perivascular innervation staining was conducted using anti-tyrosine hydroxylase (TH), anti-neuropeptide Y (NPY), anti-vasoactive intestinal peptide (VIP) and anti-choline acetyl transferase (ChAT). The combined Spalteholz technique and μ-CT revealed a highly consistent arrangement of the auricular vasculature. The superficial temporal (STA) and posterior auricular artery (PAA) supply the helical rim arcade and arcade, with the STA mainly forming the superior and the PAA forming the middle and inferior auricular artery. Co-existence of sympathetic NPY+ and TH+ terminals mediating vasoconstriction, and VIP+ and ACh+ indicating cholinergic vasodilatation, was found in the perivascular zone. The presence of both sympathetic vasoconstriction and cholinergic co-innervation for active vasodilatation was shown in the perivascular auricular zone. Assuming that the highly-consistent vasculature gives way to these terminals, this periarterial innervation may be found spread out across the helix.

Although delayed cerebral vasospasm (DCV) following subarachnoid hemorrhage (SAH) is closely related to the progression of brain damage, little is known about the molecular mechanism underlying its development. High mobility group box-1 (HMGB1) plays an important role as an initial inflammatory mediator in SAH. In this study, an SAH rat model was employed to evaluate the effects of anti-HMGB1 monoclonal antibody (mAb) on DCV after SAH. A vasoconstriction of the basilar artery (BA) associated with a reduction of nuclear HMGB1 and its translocation in vascular smooth muscle cells were observed in SAH rats, and anti-HMGB1 mAb administration significantly suppressed these effects. Up-regulations of inflammation-related molecules and vasoconstriction-mediating receptors in the BA of SAH rats were inhibited by anti-HMGB1 mAb treatment. Anti-HMGB1 mAb attenuated the enhanced vasocontractile response to thrombin of the isolated BA from SAH rats and prevented activation of cerebrocortical microglia. Moreover, locomotor activity and weight loss recovery were also enhanced by anti-HMGB1 mAb administration. The vasocontractile response of the BA under SAH may be induced by events that are downstream of responses to HMGB1-induced inflammation and inhibited by anti-HMGB1 mAb. Anti-HMGB1 mAb treatment may provide a novel therapeutic strategy for DCV and early brain injury after SAH.

Curcumin exerts beneficial effects on cardiovascular diseases, including hypertension. However, its mechanisms are unknown. We propose that curcumin prevents the development of hypertension by regulating AT1 receptor (AT1R) expression in arteries. The present study examined how curcumin regulates AT1R expression in vascular smooth muscle cells and investigated the physiological significance of this regulation in angiotensin (Ang) II-induced hypertension. The results showed that curcumin decreased AT1R expression in a concentration- and time-dependent manner in vascular smooth muscle cells. Using luciferase reporters with an entire AT1 or a mutant AT1R in A10 cells, the AT1R promoter activity was inhibited by 10(-6 )M curcumin, and the proximal element (from -61 to +25 bp) of the AT1R promoter was crucial for curcumin-induced AT1R down-regulation. An electrophoretic mobility shift assay showed that curcumin decreased specificity protein 1 (SP1) binding with the AT1R promoter in A10 cells. Curcumin treatment reduced Ang II-induced hypertension in C57Bl/6J mice, which was accompanied by lower AT1R expression in the arteries and decreased Ang II-mediated vasoconstriction in the mesenteric artery. These findings indicate that curcumin down-regulates AT1R expression in A10 cells by affecting SP1/AT1R DNA binding, thus reducing AT1R-mediated vasoconstriction and subsequently prevents the development of hypertension in an Ang II-induced hypertensive model.

An emerging challenge in tissue engineering biomimetic models is recapitulating the physiological complexity associated with real tissues. Recently, our laboratory introduced the rat mesentery culture model as an ex vivo experimental platform for investigating the multi-cellular dynamics involved in angiogenesis within an intact microvascular network using time-lapse imaging. A critical question remains whether the vessels maintain their functionality. The objective of this study was to determine whether vascular smooth muscle cells in cultured microvascular networks maintain the ability to constrict. Adult rat mesenteric tissues were harvested and cultured for three days in either MEM or MEM plus 10% serum. On Day 0 and Day 3 live microvascular networks were visualized with FITC conjugated BSI-lectin labeling and arteriole diameters were compared before and five minutes after topical exposure to vasoconstrictors (50 mM KCl and 20 nM Endothelin-1). Arterioles displayed a vasoconstriction response to KCl and endothelin for each experimental group. However, the Day 3 serum cultured networks were angiogenic, characterized by increased vessel density, and displayed a decreased vasoconstriction response compared to Day 0 networks. The results support the physiological relevance of the rat mesentery culture model as a biomimetic tool for investigating microvascular growth and function ex vivo.

Renal dysfunction is one of the most common complications of septic injury. One critical contributor to septic injury-induced renal dysfunction is renal vascular dysfunction. Protein palmitoylation serves as a novel regulator of vascular function. Here, we examined whether palmitoyl acyltransferase (PAT)-DHHC21 contributes to septic injury-induced renal dysfunction through regulating renal hemodynamics. Multispectral optoacoustic imaging showed that cecal ligation and puncture (CLP)-induced septic injury caused impaired renal excretion, which was improved in DHHC21 functional deficient (Zdhhc21dep/dep) mice. DHHC21 deficiency attenuated CLP-induced renal pathology, characterized by tissue structural damage and circulating injury markers. Importantly, DHHC21 loss-of-function led to better-preserved renal perfusion and oxygen saturation after CLP. The CLP-caused reduction in renal blood flow was also ameliorated in Zdhhc21dep/dep mice. Next, CLP promoted the palmitoylation of vascular α1-adrenergic receptor (α1AR) and the activation of its downstream effector ERK, which were blunted in Zdhhc21dep/dep mice. Vasoreactivity analysis revealed that renal arteries from Zdhhc21dep/dep mice displayed reduced constriction response to α1AR agonist phenylephrine compared to those from wild-type mice. Consistently, inhibiting PATs with 2-bromopalmitate caused a blunted vasoconstriction response to phenylephrine in small arteries isolated from human kidneys. Therefore, DHHC21 contributes to impaired renal perfusion and function during septic injury via promoting α1AR palmitoylation-associated vasoconstriction.

Acute-on-chronic liver disease is a clinical syndrome characterized by decompensated liver fibrosis, portal hypertension and splanchnic hyperdynamic circulation. We aimed to determine whether the alpha-1 agonist phenylephrine (Phe) facilitates endothelial nitric oxide (NO) release by mesenteric resistance arteries (MRA) in rats subjected to an experimental microsurgical obstructive liver cholestasis model (LC). Sham-operated (SO) and LC rats were maintained for eight postoperative weeks. Phe-induced vasoconstriction (in the presence/absence of the NO synthase -NOS- inhibitor L-NAME) and vasodilator response to NO donor DEA-NO were analysed. Phe-induced NO release was determined in the presence/absence of either H89 (protein kinase -PK- A inhibitor) or LY 294002 (PI3K inhibitor). PKA and PKG activities, alpha-1 adrenoceptor, endothelial NOS (eNOS), PI3K, AKT and soluble guanylate cyclase (sGC) subunit expressions, as well as eNOS and AKT phosphorylation, were determined. The results show that LC blunted Phe-induced vasoconstriction, and enhanced DEA-NO-induced vasodilation. L-NAME increased the Phe-induced contraction largely in LC animals. The Phe-induced NO release was greater in MRA from LC animals. Both H89 and LY 294002 reduced NO release in LC. Alpha-1 adrenoceptor, eNOS, PI3K and AKT expressions were unchanged, but sGC subunit expression, eNOS and AKT phosphorylation and the activities of PKA and PKG were higher in MRA from LC animals. In summary, these mechanisms may help maintaining splanchnic vasodilation and hypotension observed in decompensated LC.