Mitogen-activated protein kinase (MAPK) signaling is strongly implicated in cardiovascular remodeling in pulmonary hypertension (PH) and right ventricle (RV) failure. The effects of a newly designed p38 inhibitor, LASSBio-1824, were investigated in experimentally induced PH. Male Wistar rats were exposed to hypoxia and SU5416 (SuHx), and normoxic rats were used as controls. Oral treatment was performed for 14 days with either vehicle or LASSBio-1824 (50 mg/kg). Pulmonary vascular resistance and RV structure and function were assessed by echocardiography and catheterization. Histological, immunohistochemical and Western blot analysis of lung and RV were performed to investigate cardiovascular remodeling and inflammation. Treatment with LASSBio-1824 normalized vascular resistance by attenuating vessel muscularization and endothelial dysfunction. In the heart, treatment decreased RV systolic pressure, hypertrophy and collagen content, improving cardiac function. Protein content of TNF-α, iNOS, phosphorylated p38 and caspase-3 were reduced both in lung vessels and RV tissues after treatment and a reduced activation of transcription factor c-fos was found in cardiomyocytes of treated SuHx rats. Therefore, LASSBio-1824 represents a potential candidate for remodeling-targeted treatment of PH.

Introduction: New therapeutic alternatives for pain relief include the use of phosphodiesterase-5 (PDE5) inhibitors, which could prevent the transmission of painful stimuli by neuron hyperpolarization via nitric oxide (NO)/cyclic 3',5'-guanosine monophosphate (cGMP) pathway. The present work investigated the antinociceptive activity of a new PDE5 inhibitor, lodenafil carbonate, in inflammatory and neuropathic pain models. Methods and Results: Although no effect was detected on neurogenic phase of formalin test in mice, oral administration of lodenafil carbonate dose-dependently reduced reactivity in the inflammatory phase (200.6 +/- 39.1 to 81.9 +/- 18.8 s at 10 nmol/kg, p= 0.0172) and this effect was totally blocked by NO synthase inhibitor, L-Nco-nitroarginine methyl ester (L-NAME). Lodenafil carbonate (10 mu mol/kg p.o.) significantly reduced nociceptive response as demonstrated by increased paw withdrawal latency to thermal stimulus (from 6.8 +/- 0.7 to 10.6 +/- 1.3 s, p= 0.0006) and paw withdrawal threshold to compressive force (from 188.0 +/- 14.0 to 252.5 +/- 5.3 g, p<0.0001) in carrageenan-induced paw inflammation model. In a spinal nerve ligation-induced neuropathic pain, oral lodenafil carbonate (10 nmol/kg) also reversed thermal hyperalgesia and mechanical allodynia by increasing paw withdrawal latency from 17.9 +/- 1.5 to 22.8 +/- 1.9 s (p= 0.0062) and paw withdrawal threshold from 26.0 +/- 2.8 to 41.4 +/- 2.9 g (p= 0.0196). These effects were reinforced by the reduced GFAP (3.4 +/- 0.5 to 1.4 +/- 0.3%, p= 0.0253) and TNF-alpha (1.1 +/- 0.1 to 0.4 +/- 0.1%, p= 0.0111) stained area densities as detected by immunofluorescence in ipsilateral dorsal horns. Conclusion: Lodenafil carbonate demonstrates important analgesic activity by promoting presynaptic hyperpolarization and preventing neuroplastic changes, which may perpetuate chronic pain, thus representing a potential treatment for neuropathic pain.

INTRODUCTION:Diabetic obese patients are susceptible to the development of cardiovascular disease, including hypertension and cardiac dysfunction culminating in diabetic cardiomyopathy (DC), which represents a life-threatening health problem with increased rates of morbidity and mortality. The aim of the study is to characterize the effects of a new benzofuran N-acylhydrazone compound, LASSBio-2090, on metabolic and cardiovascular alterations in Zucker diabetic fatty (ZDF) rats presenting DC.METHODS:Male non-diabetic lean Zucker rats (ZL) and ZDF rats treated with vehicle (dimethylsulfoxide) or LASSBio-2090 were used in this study. Metabolic parameters, cardiovascular function, left ventricle histology and inflammatory protein expression were analyzed in the experimental groups.RESULTS:LASSBio-2090 administration in ZDF rats reduced glucose levels to 85.0 ± 1.7 mg/dL (p < 0.05). LASSBio-2090 also lowered the cholesterol and triglyceride levels from 177.8 ± 31.2 to 104.8 ± 5.3 mg/dL and from 123.0 ± 11.4 to 90.9 ± 4.8 mg/dL, respectively, in obese diabetic rats (p < 0.05). LASSBio-2090 normalized plasma insulin, insulin sensitivity and endothelial function in aortas from diabetic animals (p < 0.05). It also enhanced systolic and diastolic left-ventricular function and reverted myocardial remodeling by blocking the threefold elevation of TNF-α levels in hearts from ZDF rats.CONCLUSION:LASSBio-2090 alleviates metabolic disturbance and cardiomyopathy in an obese and diabetic rat model, thus representing a novel strategy for the treatment of cardiovascular complications in obesity-associated type 2 diabetes mellitus.

Pulmonary arterial hypertension (PAH) is characterized by the remodeling of pulmonary arteries, with an increased pulmonary arterial pressure and right ventricle (RV) overload. This work investigated the benefit of the association of human umbilical cord mesenchymal stem cells (hMSCs) with lodenafil, a phosphodiesterase-5 inhibitor, in an animal model of PAH. Male Wistar rats were exposed to hypoxia (10% O2) for three weeks plus a weekly i.p. injection of a vascular endothelial growth factor receptor inhibitor (SU5416, 20 mg/kg, SuHx). After confirmation of PAH, animals received intravenous injection of 5.105 hMSCs or vehicle, followed by oral treatment with lodenafil carbonate (10 mg/kg/day) for 14 days. The ratio between pulmonary artery acceleration time and RV ejection time reduced from 0.42 ± 0.01 (control) to 0.24 ± 0.01 in the SuHx group, which was not altered by lodenafil alone but was recovered to 0.31 ± 0.01 when administered in association with hMSCs. RV afterload was confirmed in the SuHx group with an increased RV systolic pressure (mmHg) of 52.1 ± 8.8 normalized to 29.6 ± 2.2 after treatment with the association. Treatment with hMSCs + lodenafil reversed RV hypertrophy, fibrosis and interstitial cell infiltration in the SuHx group. Combined therapy of lodenafil and hMSCs may be a strategy for PAH treatment.

Aim: Investigate cardiac electrical and mechanical dysfunctions elicited by chronic anabolic steroid (AS) overdose. Methods: Male Wistar rats were treated with nandrolone decanoate (DECA) or vehicle (CTL) for 8 weeks. Electrocardiography and heart rate variability were assessed at weeks 2, 4, and 8. Cardiac reactivity to isoproterenol was investigated in isolated rat hearts. Action potential duration (APD) was measured from left ventricular (LV) muscle strips. L-type Ca2+ current (I-CaL), and transient outward potassium current (I-to) were recorded by whole-cell patch-clamp in LV cardiomyocytes. Sarcoplasmic reticulum (SR) Ca2+ mobilization and Ca2+-induced contractile response sensitivity were evaluated in skinned cardiac fibers. Muscarinic type 2 receptor (M2R), beta(1)-adrenergic receptor (beta(1)AR), sarcoplasmic Ca2+ ATPase (SERCA-2a), type 2 ryanodine receptor (RyR2), L-type Ca2+ channel (CACNA1), Kv4.2 (KCND2), and Kv4.3 (KCND3) mRNA expression levels were measured by quantitative RT-PCR. Results: Compared with CTL group, DECA group exhibited decreased high frequency band power density (HF) and increased low frequency power density (LF), Cardiac M2R mRNA level was decreased. QTc interval at 2nd, 4th, and 8th week as well as APD(30) and APD(90) were increased by DECA. 4, density was decreased, while I-CaL, density was increased by DECA. SR Ca2+ loading and release were decreased by DECA, while contractile sensitivity to Ca2+ was increased versus CTL group. Conclusion: DECA overdose induced cardiac rhythmic and mechanical abnormalities that can be associated with autonomic imbalance, up-regulated I-CaL and down-regulated I-to, abnormal SR Ca2+ mobilization, and increased contractile sensitivity to Ca2+.

This study was aimed to clarify differences in how specific agonists of the 3 estrogen receptors (ERs) influence diastolic function and the renin-angiotensin system (RAS) after ovariectomy (OVX) in 24 female spontaneously hypertensive rat (SHR) undergoing bilateral OVX at 12 weeks of age. Eight weeks after surgery, rats were randomized (n = 6/group) to receive equipotent, daily treatments of one of the ER agonists (ERα agonist, propyl pyrazole trisphenol 94 μg/kg; ERβ agonist, diarylpropionitrile 58 μg/kg; G-protein-coupled estrogen receptor [GPER] agonist, G1 100 μg/kg), or vehicle (peanut oil). After 4 weeks of treatment, left ventricular function/structure and systemic/intracardiac pressure measurements were obtained by echocardiography and a fluid-filled catheter attached to a pressure transducer, respectively. Selective ER agonist treatment with G1 or propyl pyrazole trisphenol led to improvements in diastolic function after estrogen loss when compared with vehicle-treated OVX rats. Although mean arterial blood pressure was not overtly different among groups, chronic G1, but not the other ER ligands, enhanced the in vitro vasorelaxant responsiveness to acetylcholine in aortic rings. These favorable effects of G1 were further linked to reductions in cardiac angiotensin-converting enzyme activity, AT1R protein expression, and Ang II immunoreactivity. Activation of ERβ had no effect on cardiac function and did not alter components of the canonical cardiac RAS in comparison with vehicle-treated OVX SHR. These data imply that of the 3 ERs, GPER has a unique role in preserving diastolic function and favorably modulating the cardiac RAS independent of arterial pressure. Specifically, if GPER is pharmacologically activated, it could provide a therapeutic opportunity to limit the development and/or progression of diastolic dysfunction in hypertensive women after estrogen loss.

Increased mortality due to type 2 diabetes mellitus (T2DM) has been associated with renal and/or cardiovascular dysfunction. Dipeptidyl dipeptidase-4 inhibitors (iDPP-4s) may exert cardioprotective effects through their pleiotropic actions via glucagon-like peptide 1-dependent mechanisms. In this study, the pharmacological profile of a new iDPP-4 (LASSBio-2124) was investigated in rats with cardiac and renal dysfunction induced by T2DM. T2DM was induced in rats by 2 weeks of a high-fat diet followed by intravenous injection of streptozotocin. Metabolic disturbance and cardiac, vascular, and renal dysfunction were analyzed in the experimental groups. Sitagliptin and LASSBio-2124 administration after T2DM induction reduced elevated glucose levels to 319.8 ±13.2 and 279.7 ± 17.8 mg/dL, respectively (p<0.05). LASSBio-2124 also lowered the cholesterol and triglyceride levels from 76.8 ±8.0 to 42.7 ± 3.2 mg/dL and from 229.7 ±25.4 to 100.7 ± 17.1 mg/dL, in diabetic rats. Sitagliptin and LASSBio-2124 reversed the reduction of the plasma insulin level. LASSBio-2124 recovered the increased urinary flow in diabetic animals and reduced 24-h proteinuria from 23.7 ±1.5 to 13.3 ±2.8 mg (p<0.05). It also reduced systolic and diastolic left-ventricular dysfunction in hearts from diabetic rats. The effects of LASSBio-2124 were superior to those of sitagliptin in the cardiovascular systems of T2DM rats. This new prototype showed promise for the avoidance of comorbidities in a T2DM experimental model, and thus may constitute an innovative therapeutic agent for the treatment of these conditions in the clinical field in future.

Objective: To demonstrate the antinociceptive and antihypersensitivity mechanisms of Cris-104 (1-{2-[5-(4-fluorophenyl)–1H-pyrazol-4-yl]ethyl}piperidine), a novel selective α 4 β 2 * nicotinic acetylcholine receptor (nAChR) agonist, in rodent acute/inflammatory and chronic pain models. Materials and methods: Hot-plate and formalin tests in mice were used to examine Cris-104-induced antinociceptive effects on thermal/inflammatory pain. Cris-104 effects on hypersensitivity, norepinephrine (NE) release in the spinal dorsal horn, and neuronal activity in the locus coeruleus (LC) were examined in rats with lumbar spinal nerve ligation using behavioral, microdialysis, and extracellular recording methods. Cris-104 effects on spontaneous locomotion were examined in an open-field test. Results: Cris-104 induced dose-dependent antinociception effects in hot-plate and formalin tests, and these effects were blocked by the general nAChR antagonist mecamylamine, the selective α 4 β 2 * nAChR antagonist dihydro-beta-erythroidine, and the α 2 -adrenoceptor antagonist yohimbine, but not by the α 1 -adrenoceptor antagonist prazosin. Systemic and spinally perfused Cris-104 increased NE concentrations in microdialysates from the spinal cord in both normal and SNL rats. Systemic Cris-104 increased neuronal activity in the LC of normal rats. Mecamylamine blocked the effects of Cris-104 on spinal NE release and LC neuronal activity. Systemic Cris-104 did not affect locomotor activity significantly. Conclusion: The α 4 β 2 neuronal nAChR agonist, Cris-104, was effective for treatment of pain via descending noradrenergic inhibition of pain signaling. Keywords: pain, nicotinic receptor, Cris-104, epibatidine

Introduction: Pulmonary arterial hypertension (PAH) is a disease characterized by pulmonary vascular remodeling with subsequent right ventricular (RV) failure. Hypothesis: The p38-alpha mitogen-activated protein kinase (p38-α MAPK) plays a pathogenic role in PAH. This work investigated the effects of a selective p38-alpha MAPK inhibitor, named LASSBio-1824, on hypoxia + SU5416 -induced PAH. Methods: Male Wistar rats (180-220 g) were exposed to 3 weeks of hypoxia and i.p. injection of VEGFR antagonist (SU5416; 20 mg/kg/week). Experimental groups were: Control + vehicle (DMSO), SuHx + vehicle (PAH group) and SuHx + LASSBio-1824 (20 mg/kg/day for 2 weeks). Results: Pulmonary acceleration time (ms) was reduced from 44.7 ± 1.1 (control) to 20.6 ± 1.7 in SuHx + vehicle group (P < 0.05) and restored to 38.9 ± 2.9 in SuHx + LASSBio-1824 group (P< 0.05). RV systolic pressures (mmHg) were increased from 30.1 ± 3.2 (control) to 109.5 ± 7.5 in SuHx group and reduced to 47.5 ± 5.3 after treatment with LASSBio-1824 (P < 0.05). The medial wall thickness of distal pulmonary arterioles (< 50 μm) was measured by immunohistochemistry for alpha-SMA expression which was increased from 40.7 ± 3.5 % (control) to 64.1 ± 3.1 % in PAH animals (P < 0.05) and was reduced in the treated group (52.2 ± 2.9). Acetylcholine-induced maximal relaxation (% ) in pulmonary arteries was reduced from 77.9 ± 5.2 to 17.2 ± 3.5 in PAH group. LASSBio-1824 increased that response to 67.8 ± 5.6 % (P < 0.05), indicating the ame- lioration of endothelial function. RV hypertrophy was detected because its wall thickness (mm) increased from 0.56 ± 0.02 (control) to 1.3 ± 0.03 in SuHx + vehicle group (P < 0.05) and the ratio between RV and left ventricle + septum increased from 1.50 ± 0.03 to 2.0 ± 0.2. Treatment with LASSBio-1824 significantly reduced the RV hypertrophy (ratio of 1.7 ± 0.07). Conclusion: The inhibition of p38-alpha MAPK isoform by LASSBio-1824 represents an impor- tant approach for the treatment of PAH in the future, which improves the underlying remodeling and inflammation processes.

IntroductionLeft ventricular diastolic dysfunction (LVDD) increases the risk of developing heart failure with preserved ejection fraction (HFpEF), the predominant type of heart failure seen in women. Emerging epidemiologic evidence suggests that early menopause is positively associated with incident HF, which suggests potential protective roles of estrogens in LVDD development and progression. There are three estrogen receptors (ER) expressed in the heart, ER‐α, ER‐β, and G‐protein coupled ER (GPER). We previously found that GPER plays greater protective roles in cultured myocyte and mast cell lines than ER‐α or ER‐β, by inhibiting Ang II‐induced cell hypertrophy and serum‐induced cell proliferation, respectively. Moreover, estrogen loss‐induced LVDD in the hypertensive mRen2. Lewis rat was linked to increases in cardiac mast cell number, chymase, and Ang II. The aim of this study was to investigate how different specific agonists of estrogen receptors (ER‐α agonist PPT; ER‐β agonist DPN; and GPER agonist G‐1) influence diastolic function and cardiac renin angiotensin system (RAS) components.MethodsTwenty‐four SHR females underwent bilateral ovariectomy (OVX) at 12 weeks of age; 8 weeks post‐surgery, rats were randomized (n=6/group) to receive equipotent, daily treatments (0.24 μMol/kg/day, s.c.) of one of the above ER agonists (PPT‐ 94 μg/kg; DPN‐ 58 μg/kg; G‐1‐100 μg/kg), or vehicle (peanut oil). Following four weeks of treatment, rats were anesthetized (isoflurane) for direct blood pressure and Doppler echo measures of diastolic function, defined by e′ and E/e′. LV tissue for RAS activity and expression were evaluated. Data were analyzed by one‐way ANOVA. P<0.05 was significant.ResultsAs expected, the doses chosen for the specific ER agonists did not overtly affect blood pressure. Of the ER agonists, G‐1 had the most profound and significant impact on diastolic function, increasing myocardial relaxation (e′) and decreasing Doppler‐derived LV filling pressures (E/e′) by 53% and 24% versus control, respectively. Neither DPN nor PPT modified the altered cardiac function induced by OVX. GPER activation by G‐1 had a significant inhibitory effect on cardiac ACE activity when compared to vehicle, while the other Ang II‐forming pathway in the heart, Ang‐1–12/chymase, was not overtly altered by ER activation. All three agonists suppressed cardiac ACE2 activity. AT1R gene expression was increased in OVX‐vehicle by 47 to 61% when compared to hearts from OVX‐ERs (P=0.051) (Figure).ConclusionThese data imply that of the three estrogen receptors, GPER has a unique role in the preservation of diastolic function and modulation of the cardiac RAS, which would contribute to the protection in LVDD disease progression after estrogen loss, if pharmacologically activated.Support or Funding InformationNIH AG033727 (LG) and NIH HL 051952 (CMF and LG) INCT‐INOFAR ‐ Proc. 465249/2014‐0 (GZS and RTS) Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) (JS) Conselho Nacional de Desenvolvimento Cientifico e Tecnológico (CNPq) (GZS and RTS)This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Introduction: Prevention of cardiac remodeling induced by myocardial infarction (MI) is of great importance because it is one of the factors for the development of heart failure (HF). Thus, LASSBio-1027 was tested in acute model of MI due to its vasodilatory and anti-proliferative profile through the activation of adenosine A2A and A3 receptors.

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Aims: Pulmonary arterial hypertension (PAH) is a disease characterized by an increase in pulmonary vascular resistance and right ventricular (RV) failure. We aimed to determine the effects of human mesenchymal stem cell (hMSC) therapy in a SU5416/hypoxia (SuH) mice model of PAH.Methods and Results: C57BL/6 mice (20–25 g) were exposure to 4 weeks of hypoxia combined vascular endothelial growth factor receptor antagonism (20 mg/kg SU5416; weekly s.c. injections; PAH mice). Control mice were housed in room air. Following 2 weeks of SuH exposure, we injected 5 × 105 hMSCs cells suspended in 50 μL of vehicle (0.6 U/mL DNaseI in PBS) through intravenous injection in the caudal vein. PAH mice were treated only with vehicle. Ratio between pulmonary artery acceleration time and RV ejection time (PAAT/RVET), measure by echocardiography, was significantly reduced in the PAH mice, compared with controls, and therapy with hMSCs normalized this. Significant muscularization of the PA was observed in the PAH mice and hMSC reduced the number of fully muscularized vessels. RV free wall thickness was higher in PAH animals than in the controls, and a single injection of hMSCs reversed RV hypertrophy. Levels of markers of exacerbated apoptosis, tissue inflammation and damage, cell proliferation and oxidative stress were significantly greater in both lungs and RV tissues from PAH group, compared to controls. hMSC injection in PAH animals normalized the expression of these molecules which are involved with PAH and RV dysfunction development and the state of chronicity.Conclusion: These results indicate that hMSCs therapy represents a novel strategy for the treatment of PAH in the future.

Pulmonary hypertension (PH) is a disease of women (female-to-male ratio 4:1), and is associated with cardiac and skeletal muscle dysfunction. Herein, the activation of a new estrogen receptor (GPER) by the agonist G1 was evaluated in oophorectomized rats with monocrotaline (MCT)-induced PH. Depletion of estrogen was induced by bilateral oophorectomy (OVX) in Wistar rats. Experimental groups included SHAM or OVX rats that received a single intraperitoneal injection of MCT (60 mg/kg) for PH induction. Animals received s.c. injection of either vehicle or G1, a GPER agonist, (400 µg/kg/day) for 14 days after the onset of disease. Rats with PH exhibited exercise intolerance and cardiopulmonary alterations, including reduced pulmonary artery flow, biventricular remodeling, and left ventricular systolic and diastolic dysfunction. The magnitude of these PH-induced changes was significantly greater in OVX versus SHAM rats. G1 treatment reversed both cardiac and skeletal muscle functional aberrations caused by PH in OVX rats. G1 reversed PH-related cardiopulmonary dysfunction and exercise intolerance in female rats, a finding that may have important implications for the ongoing clinical evaluation of new drugs for the treatment of the disease in females after the loss of endogenous estrogens.

Background: Activation of adenosine receptors might mediate antinociceptive and anti-inflammatory effects. Molecular docking studies and binding assays demonstrated that 3,4-methylenedioxybenzoyl-2-thienylhydrazone (LASSBio-1027) is a ligand to adenosine receptors A2A and A3. This work evaluated the antinociceptive effect of LASSBio-1027 in murine acute and chronic pain/inflammatory models. Methods: Protocols were approved by the Animal Care and Use Committee at Universidade Federal Rio de Janeiro (Protocol 113/14). Formalin injection (20 uL i.pl.) was used to induce acute/inflammation pain in male Swiss mice (25-30 g). The antinociceptive effect was tested after oral administration (gavage) of vehicle, LASSBio-1027 (25, 50 and 100 mg/kg), morphine (30 mg/kg) or acetyl salicylic acid (300 mg/kg). Chronic inflammatory pain was induced by subcutaneous injection of Complete Freund Adjuvant (CFA) around the tibiotarsal joint in male Swiss mice under 2% sevoflurane anesthesia. Thermal and mechanical hyperalgesia and paw edema were measured in the animals after 13 days of treatment with vehicle, LASSBio-1027 (25, 50 and 100 mg/kg) or acetylsalicylic acid (300 mg/kg). At the end of treatment, protein expression of TNF-alpha, iNOS, phosphorylated p-38 and c-fos in the paw and spinal cord were evaluated. Toluidine blue staining examined the histology features of the tibiotarsal joint. Results: Treatment with LASSBio-1027 reduced in a dose-dependent manner the early and later phases of formalin-induced reactivity. At 100 mg/kg, the animal reactivity reduced from 39.7 ± 5.1 (vehicle) to 18.4 ± 1.6 s (p<0.01) and from 249.2 ± 20.8 s (vehicle) to 117.2 ± 17.9 s (p<0.01) in the early and late phase, respectively. Pre-treatment with MRE 3008F20 or ZM 241385, adenosine A3 and A2A antagonists reduced both responses. LASSBio-1027 also reduced CFA-induced thermal and mechanical hyperalgesia, paw edema and normalized overexpressed TNF-alpha, iNOS, phosphorylated p-38 (paw) and c-fos (spinal cord). The joint damage and inflammation were ameliorated with LASSBio-1027 treatment. Conclusions: A novel agonist of A3 and A2A receptors, named LASSBio-1027, could be a candidate for drug development to treat inflammatory-induced acute or chronic disease.

Hypertensive individuals are at greater risk for developing chronic kidney disease (CKD). Reducing proteinuria has been suggested as a possible therapeutic approach to treat CKD. However, the mechanisms underlying the development of proteinuria in hypertensive conditions are incompletely understood. Cardiac and vascular dysfunction is associated with changes in the O-GlcNAcylation pathway in hypertensive models. We hypothesized that O-GlcNAcylation is also involved in renal damage, especially development of proteinuria, associated with hypertension. Using the spontaneously hypertensive rat (SHR) model, we observed higher renal cortex O-GlcNAcylation, glutamine-fructose aminotransferase (GFAT), and O-GlcNAc transferase (OGT) protein expression, which positively correlated with proteinuria. Interestingly, this was observed in hypertensive, but not pre-hypertensive, rats. Pharmacological inhibition of GFAT decreased renal cortex O-GlcNAcylation, proteinuria, and albuminuria in SHR. Using a proximal tubule cell line, we observed that increased O-GlcNAcylation reduced megalin surface expression and albumin endocytosis in vitro, and the effects were correlated in vivo. Moreover, megalin is O-GlcNAcylated both in vitro and in vivo. In conclusion, our results demonstrate a new mechanism involved in hypertension-associated proteinuria.

Objective: To demonstrate the antinociceptive and antihypersensitivity mechanisms of Cris-104 (1- {2-[5-(4-fluoropheny1)-1H-pyrazol-4-yl]ethyl}piperidine), a novel selective alpha(4)beta(2)* nicotinic acetylcholine receptor (nAChR) agonist, in rodent acute/inflammatory and chronic pain models. Materials and methods: Hot-plate and formalin tests in mice were used to examine Cris-104-induced antinociceptive effects on thermal/inflammatory pain. Cris-104 effects on hypersensitivity, norepinephrine (NE) release in the spinal dorsal horn, and neuronal activity in the locus coeruleus (LC) were examined in rats with lumbar spinal nerve ligation using behavioral, microdialysis, and extracellular recording methods. Cris-104 effects on spontaneous locomotion were examined in an open-field test. Results: Cris-104 induced dose-dependent antinociception effects in hot-plate and formalin tests, and these effects were blocked by the general nAChR antagonist mecamylamine, the selective alpha(4)beta(2)* nAChR antagonist dihydro-beta-erythroidine, and the alpha(2)-adrenoceptor antagonist yohimbine, but not by the alpha(2)-adrenoceptor antagonist prazosin. Systemic and spinally perfused Cris-104 increased NE concentrations in microdialysates from the spinal cord in both normal and SNL, rats. Systemic Cris-104 increased neuronal activity in the LC of normal rats. Mecamylamine blocked the effects of Cris-104 on spinal NE release and LC neuronal activity. Systemic Cris-104 did not affect locomotor activity significantly. Conclusion: The alpha(4)beta(2) neuronal nAChR agonist, Cris-104, was effective for treatment of pain via descending noradrenergic inhibition of pain signaling.