2-((2-(4-Iodo-2,5-dimethoxyphenyl)ethylamino)methyl)phenol (25I-NBOH) and 2-(((2-(4-chloro-2,5-dimethoxyphenyl)ethyl)amino)methyl)phenol (25C-NBOH) induce adverse effects on the cardiovascular system.

Two new psychoactive substances (NPSs) classified as phenethylamines, namely 2-((2-(4-Iodo-2,5-dimethoxyphenyl)ethylamino)methyl)phenol (25I-NBOH) and 2-(((2-(4-chloro-2,5-dimethoxyphenyl)ethyl)amino)methyl)phenol (25C-NBOH), are being abused by people seeking recreational hallucinogens. These NPSs may cause serious health problems as their adverse effects are not known in most cases. Therefore, in the present study, we evaluated the cardiotoxicity of 25I-NBOH and 25C-NBOH using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, rat electrocardiography (ECG), Langendorff test, and human ether-a-go-go-related gene (hERG) assay. Furthermore, we analyzed the expression levels of p21 CDC42/RAC1-activated kinase 1 (PAK1), which is known to play various roles in the cardiovascular system. In the MTT assay, treatment with 25I-NBOH or 25C-NBOH dramatically decreased viability of H9c2 cardiomyocytes. Meanwhile, these two compounds significantly increased QT intervals and RR intervals in the rat ECG measurement. 25I-NBOH down-regulated the PAK1 protein expression in rat primary cardiomyocytes as well as H9c2 cells. However, 25C-NBOH had no effect on the PAK1 expression in H9c2 cells. In an in-depth study, 25I-NBOH inhibited potassium channels in the hERG assay, but in ex vivo test, the substance did not affect the left ventricular developed pressure (LVDP) and heart rate of the isolated rat hearts. Taken together, these results suggest that both 25I-NBOH and 25C-NBOH may have adverse cardiovascular effect. Further investigation would be needed to determine which factors mainly influence the relationship between PAK1 expression and cardiotoxicity.

Pentoxifylline, a nonselective phosphodiesterase inhibitor, in adjunctive therapy in patients with irritable bowel syndrome treated with mebeverine.

BACKGROUND: Irritable bowel syndrome (IBS) is a functional gastrointestinal condition marked by chronic bowel pain or discomfort, as well as changes in abdominal motility. Despite its worldwide prevalence and clinical impact, the cause of IBS is unknown. Inflammation could play a fundamental role in the development of IBS. The aim of this study was to examine whether pentoxifylline, a competitive nonselective phosphodiesterase inhibitor, is useful in alleviating abdominal pain in IBS patients treated with mebeverine. METHODS: A randomized, controlled, and prospective clinical study that included 50 outpatients who met the inclusion criteria for IBS. Patients are allocated randomly into two groups (n = 25). Group 1 (mebeverine group) received mebeverine 135 mg three times daily (t.i.d) for three months. Group 2 (pentoxifylline group) received mebeverine 135 mg t.i.d and pentoxifylline 400 mg two times daily for three months. Patients were assessed by a gastroenterologist at baseline and three months after the medication had been started. The serum levels of interleukin-6, interleukin-8 and tumor necrosis factor-alpha, fecal Neutrophil Gelatinase Associated Lipocalin (NGAL), and fecal myeloperoxidase were measured at the start and after three months of therapy. The Numeric Pain Rating scale (NRS) was assessed at baseline and after therapy. RESULTS: the pentoxifylline group showed a significant decrease in the level of measured biomarkers and a significant decrease in NRS. CONCLUSION: Pentoxifylline could be a promising adjuvant anti-inflammatory drug in the treatment of abdominal pain in IBS patients treated with mebeverine.

Formulation development of directly compressible mebevarine tablets using superdisintegrant: A way to investigate quality atributes, in vitro release kinetics and stability profile.

In order for preparing a solid oral dosage form, tablet quality is of significant concern. Compressibility behavior of different powders and mixtures of formulations and release pattern of any tablets are characteristic measures to define prerequisite quality attributes of any compressed formulations. There are basically two major methods that can be adopted for the preparation of tablets including granulation and direct compression. Later process offer fewer processing steps and agreeable release profile with acceptable quality parameters and hence preferred over granulation method. In this investigation Mebeverine hydrochloride an anti-muscarinic drug is studied for compression and release behavior using various concentrations of filler binders and disintegrants via rotatable central composite design (CCRD) option of design expert (software). Nine formulations were developed from F1 to F9 with Crospovidone (superdisintegrant) as (X1) (-α=1.17% to ± α=6.83%) and microcrystalline cellulose (Avicel 102, Filler/binder) as X2 (-α = 29.82% to ± α = 65.18%). Disintegration Time (DT) as (R1) and Hardness in (kg) as (R2) were determined as two dependent response variables. The performance of powder blends and formulations was analyzed by micromeritic and physico-chemical and assessments. Dissolution comparisons were statistically analyzed by ANOVA and model dependent and in-dependent methods. Best fit model was found to be Hixon-crowell's model (r2 = 0.995) followed by Weibull's model (r2 = 0.985). The Trial formulations F2, F4, F6 and F8 were also studied on accelerated conditions (40±5ºC 75%±5% RH) for stability tests and validity of the formulations in months were also determined between 35-39 months.

A deep learning approach to identify gene targets of a therapeutic for human splicing disorders.

Pre-mRNA splicing is a key controller of human gene expression. Disturbances in splicing due to mutation lead to dysregulated protein expression and contribute to a substantial fraction of human disease. Several classes of splicing modulator compounds (SMCs) have been recently identified and establish that pre-mRNA splicing represents a target for therapy. We describe herein the identification of BPN-15477, a SMC that restores correct splicing of ELP1 exon 20. Using transcriptome sequencing from treated fibroblast cells and a machine learning approach, we identify BPN-15477 responsive sequence signatures. We then leverage this model to discover 155 human disease genes harboring ClinVar mutations predicted to alter pre-mRNA splicing as targets for BPN-15477. Splicing assays confirm successful correction of splicing defects caused by mutations in CFTR, LIPA, MLH1 and MAPT. Subsequent validations in two disease-relevant cellular models demonstrate that BPN-15477 increases functional protein, confirming the clinical potential of our predictions.

Preclinical validation of a potent γ-secretase modulator for Alzheimer's disease prevention.

A potent γ-secretase modulator (GSM) has been developed to circumvent problems associated with γ-secretase inhibitors (GSIs) and to potentially enable use in primary prevention of early-onset familial Alzheimer's disease (EOFAD). Unlike GSIs, GSMs do not inhibit γ-secretase activity but rather allosterically modulate γ-secretase, reducing the net production of Aß42 and to a lesser extent Aß40, while concomitantly augmenting production of Aß38 and Aß37. This GSM demonstrated robust time- and dose-dependent efficacy in acute, subchronic, and chronic studies across multiple species, including primary and secondary prevention studies in a transgenic mouse model. The GSM displayed a >40-fold safety margin in rats based on a comparison of the systemic exposure (AUC) at the no observed adverse effect level (NOAEL) to the 50% effective AUC or AUCeffective, the systemic exposure required for reducing levels of Aß42 in rat brain by 50%.

The involvement of PI3K/Akt/mTOR/GSK3ß signaling pathways in the antidepressant-like effect of AZD6765.

AZD6765 (lanicemine) is a non-competitive NMDA receptor antagonist that induces a fast-acting antidepressant effect without presenting psychotomimetic effects. However, the mechanisms underlying its effects remain to be established. In this context, we demonstrated that a single administration of AZD6765 (1 mg/kg, i.p.) was able to induce an antidepressant-like effect in mice submitted to tail suspension test (TST), an effect reversed by LY294002 (a reversible PI3K inhibitor, 10 nmol/site, i.c.v.), wortmannin (an irreversible PI3K inhibitor, 0.1 µg/site, i.c.v.) and rapamycin (a selective mTOR inhibitor, 0.2 nmol/site, i.c.v.). In addition, the administration of sub-effective doses of AZD6765 (0.1 mg/kg, i.p.) in combination with lithium chloride (non-selective GSK-3ß inhibitor, 10 mg/kg, p.o.) or AR-A014418 (selective GSK-3ß inhibitor, (0.01 µg/site, i.c.v.) caused a synergistic antidepressant-like effect. These results suggest the involvement of PI3K/Akt/mTOR/GSK3ß signaling in the AZD6765 antidepressant-like effect. In addition, western blotting analysis showed an increased immunocontent of synapsin in the prefrontal cortex and a tendency to an increased immunocontent of this protein in the hippocampus 30 min after AZD6765 administration, but no significant effect of AZD6765 was observed in P70S6K (Thr389) phosphorylation and GluA1 immunocontent. A single dose of AZD6765 (3 mg/kg, i.p.), similarly to ketamine (1 mg/kg, i.p.), decreased the latency to feed in the novelty suppressed feeding (NSF) test, a behavioral paradigm that evaluates depression/anxiety-related behavior. This effect was reversed by rapamycin administration, suggesting the activation of mTOR signaling in the effect of AZD in the NSF test. In addition, a single administration of AZD6765 (1 mg/kg, i.p.) or ketamine (1 mg/kg, i.p.) reversed the depressive-like behavior induced by chronic unpredictable stress (CUS). Altogether, the results provide evidence for the fast-acting antidepressant profile of AZD6765, by a mechanism likely dependent on PI3K/Akt/mTOR/GSK3ß.

Effects of zinc propionate supplementation on growth performance, skeletal muscle fiber, and receptor characteristics in beef steers.

A randomized complete block design experiment with 32 yearling crossbred steers (average body weight [BW] = 442 ± 17.0 kg) fed a steam-flaked corn-based diet was used to evaluate the effects of dietary Zn (KemTRACE Zn propionate 27; Kemin Industries, Inc., Des Moines, IA) supplementation on live growth performance, skeletal muscle fiber, and beta-adrenergic receptor (ß-AR) characteristics during the finishing phase. Steers were blocked by BW (n = 4 blocks; 8 steers/block), assigned to pens (n = 4 steers/pen), and randomly assigned to the following treatments: control (CON; 0.0 g/[head (hd) · d] of additional Zn) or additional dietary Zn (ZnP; 1.0 g/[hd · d] additional Zn). The basal diet contained Zn (60 ppm dry matter basis) from ZnSO4; additional Zn was top-dressed at feeding. Ractopamine hydrochloride (RH; Optaflexx: Elanco Animal Health, Greenfield, IN) was included at 300 mg/(hd · d) for the final 28 d of the 111-d feeding period. Longissimus muscle biopsy samples, BW, and blood were obtained on days 0, 42, 79, and 107. Final BW was collected prior to shipping on day 111. Biopsy samples were used for immunohistochemical (IHC), mRNA, and protein analysis. Serum urea nitrogen (SUN) and nonesterified fatty acid (NEFA) concentrations were measured. Steers fed ZnP had a greater average daily gain (P = 0.02) and gain to feed ratio (G:F; P = 0.03) during the RH feeding period compared with CON. There were no differences (P > 0.05) in other growth performance variables, carcass traits, mRNA abundance, or relative protein concentration for fiber type and ß-AR. Fiber types I and IIA had no differences in the cross-sectional area; however, the IIX area was greater for CON (P < 0.04) compared with ZnP and increased (P < 0.02) over time. There were no differences between treatments for the ß1-AR density (P > 0.05) in skeletal muscle tissue throughout the study. A treatment × day interaction was observed in ß2-AR density (P = 0.02) and ß3-AR density (P = 0.02) during the RH feeding period, where the abundance of the receptors increased with ZnP but did not change in CON. Compared with CON, ZnP had greater (P < 0.01) mean NEFA concentrations. Mean SUN concentrations did increase by day (P < 0.01). Additional dietary Zn, supplied as Zn propionate, upregulates ß2-AR and ß3-AR and improves growth performance in feedlot steers during the RH feeding period, likely through a shift of resource utilization from lipogenesis to muscle maintenance and hypertrophy.

Classification of drug-induced hERG potassium-channel block from electrocardiographic T-wave features using artificial neural networks.

BACKGROUND: Human ether-à-go-go-related gene (hERG) potassium-channel block represents a harmful side effect of drug therapy that may cause torsade de pointes (TdP). Analysis of ventricular repolarization through electrocardiographic T-wave features represents a noninvasive way to accurately evaluate the TdP risk in drug-safety studies. This study proposes an artificial neural network (ANN) for noninvasive electrocardiography-based classification of the hERG potassium-channel block. METHODS: The data were taken from the "ECG Effects of Ranolazine, Dofetilide, Verapamil, and Quinidine in Healthy Subjects" Physionet database; they consisted of median vector magnitude (VM) beats of 22 healthy subjects receiving a single 500 µg dose of dofetilide. Fourteen VM beats were considered for each subject, relative to time-points ranging from 0.5 hr before to 14.0 hr after dofetilide administration. For each VM, changes in two indexes accounting for the early and the late phases of repolarization, ΔERD30% and ΔTS/A , respectively, were computed as difference between values at each postdose time-point and the predose time-point. Thus, the dataset contained 286 ΔERD30% -ΔTS/A pairs, partitioned into training, validation, and test sets (114, 29, and 143 pairs, respectively) and used as inputs of a two-layer feedforward ANN with two target classes: high block (HB) and low block (LB). Optimal ANN (OANN) was identified using the training and validation sets and tested on the test set. RESULTS: Test set area under the receiver operating characteristic was 0.91; sensitivity, specificity, accuracy, and precision were 0.93, 0.83, 0.92, and 0.96, respectively. CONCLUSION: OANN represents a reliable tool for noninvasive assessment of the hERG potassium-channel block.

Reduction of repetitive behavior by co-administration of adenosine receptor agonists in C58 mice.

Repetitive behaviors are diagnostic for autism spectrum disorder (ASD) and commonly observed in other neurodevelopmental disorders. Currently, there are no effective pharmacological treatments for repetitive behavior in these clinical conditions. This is due to the lack of information about the specific neural circuitry that mediates the development and expression of repetitive behavior. Our previous work in mouse models has linked repetitive behavior to decreased activation of the subthalamic nucleus, a brain region in the indirect and hyperdirect pathways in the basal ganglia circuitry. The present experiments were designed to further test our hypothesis that pharmacological activation of the indirect pathway would reduce repetitive behavior. We used a combination of adenosine A1 and A2A receptor agonists that have been shown to alter the firing frequency of dorsal striatal neurons within the indirect pathway of the basal ganglia. This drug combination markedly and selectively reduced repetitive behavior in both male and female C58 mice over a six-hour period, an effect that required both A1 and A2A agonists as neither alone reduced repetitive behavior. The adenosine A1 and A2A receptor agonist combination also significantly increased the number of Fos transcripts and Fos positive cells in dorsal striatum. Fos induction was found in both direct and indirect pathway neurons suggesting that the drug combination restored the balance of activation across these complementary basal ganglia pathways. The adenosine A1 and A2A receptor agonist combination also maintained its effectiveness in reducing repetitive behavior over a 7-day period. These findings point to novel potential therapeutic targets for development of drug therapies for repetitive behavior in clinical disorders.

Calcimimetics maintain bone turnover in uremic rats despite the concomitant decrease in parathyroid hormone concentration.

Calcimimetics decrease parathyroid hormone (PTH) secretion in patients with secondary hyperparathyroidism. The decrease in PTH should cause a reduction in bone turnover; however, the direct effect of calcimimetics on bone cells, which express the calcium-sensing receptor (CaSR), has not been defined. In this study, we evaluated the direct bone effects of CaSR activation by a calcimimetic (AMG 641) in vitro and in vivo. To create a PTH "clamp," total parathyroidectomy was performed in rats with and without uremia induced by 5/6 nephrectomy, followed by a continuous subcutaneous infusion of PTH. Animals were then treated with either the calcimimetic or vehicle. Calcimimetic administration increased osteoblast number and osteoid volume in normal rats under a PTH clamp. In uremic rats, the elevated PTH concentration led to reduced bone volume and increased bone turnover, and calcimimetic administration decreased plasma PTH. In uremic rats exposed to PTH at 6-fold the usual replacement dose, calcimimetic administration increased osteoblast number, osteoid surface, and bone formation. A 9-fold higher dose of PTH caused an increase in bone turnover that was not altered by the administration of calcimimetic. In an osteosarcoma cell line, the calcimimetic induced Erk1/2 phosphorylation and the expression of osteoblast genes. The addition of a calcilytic resulted in the opposite effect. Moreover, the calcimimetic promoted the osteogenic differentiation and mineralization of human bone marrow mesenchymal stem cells in vitro. Thus, calcimimetic administration has a direct anabolic effect on bone that counteracts the decrease in PTH levels.

Neuroprotective effect of pharmacological postconditioning on cerebral ischaemia-reperfusion-induced injury in mice.

OBJECTIVES: To investigate the mechanism of neuroprotection rendered via pharmacological postconditioning in cerebral ischaemia-reperfusion-induced injury in mice. METHODS: Pharmacological postconditioning is strategy which either involves hindering deleterious pathway or inducing modest stress level which triggers intracellular defence pathway to sustain more vigorous insult leading to conditioning. Hence, in current research we explored the potentiality of CGS21680 (0.5 mg/kg; i.p), an adenosine A2 A receptor agonist and PTEN inhibitor, SF1670 (3 mg/kg; i.p.) to trigger postconditioning after inducing cerebral global ischaemia (17 min) and reperfusion (24 h)-induced injury via occlusion of both carotid arteries. Mice were also given treatment with LY294002 (1.5 mg/kg; i.p.), a PI3K inhibitor and adenosine A2 A receptor antagonist, Istradefylline (2 mg/kg; i.p.), to establish the precise mechanism of postconditioning. Various biochemical and behavioural parameters were assessed to examine the effect of pharmacological postconditioning. KEY FINDINGS: Pharmacological postconditioning induced with CGS21680 and SF1670 attenuated the infarction along with improved behavioural and biochemical parameters in comparison with ischaemia-reperfusion control group. The outcome of postconditioning with CGS21680 and SF1670 was significantly reversed by LY294002 and Istradefylline, respectively. CONCLUSIONS: The neuroprotective effects of CGS21680 and SF1670 postconditioning on cerebral ischaemia-reperfusion injury may be due to PI3K/Akt pathway activation.

Adenosine A2A receptors are up-regulated and control the activation of human alveolar macrophages.

Chronic inflammatory lung diseases remain a health concern and new anti-inflammatory treatments are needed. Targeting adenosine A2A receptors (A2AR) affords robust anti-inflammatory effects in animal models, but the translation of this promising strategy to humans has been challenging, possibly due to interspecies differences in receptor distribution and effects. Thus, we now assessed the efficiency of a selective A2AR agonist to control the activation of fresh human alveolar inflammatory cells. We collected bronchoalveolar lavage fluid from patients with interstitial lung disease and loaded alveolar cells with the intracellular free calcium probe FURA-2/AM. Calcium transients were then recorded in response to superfusion with a proinflammatory peptide (N-formylmethionyl-leucyl-phenylalanine - FMLP), in the absence or presence of the selective A2AR agonist CGS21680. In a second experiment, cells were continuously exposed to FMLP and A2AR density was assessed by immunocytochemistry. Sixteen patients were included, nine for analysis of calcium transients, and seven for immunocytochemistry. When alveolar macrophages were exposed to 100 nM FMLP for 120 s, a peak elevation of intracellular free calcium levels (97.0% over baseline) was recorded; CGS21680 (100 and 300 mM) significantly reduced this peak to 89.5% and 81.5%, respectively. The immunofluorescence analysis revealed a time-dependent increase of A2AR density in alveolar macrophage upon exposure to 1 µM FMLP, up to 148% of control at 6 h. These results show that pro-inflammatory stimuli up-regulate A2AR and their activation dampens the impact of pro-inflammatory stimuli. This supports that targeting A2AR is a promising therapy for human lung inflammatory diseases, especially for diseases with a strong inflammatory component.

Effects of immunocastration and ß-adrenergic agonists on the performance and carcass traits of feedlot finished Nellore cattle.

ß-Adrenergic agonists (ß-AA) are non-hormonal growth promoters which promote muscle hypertrophy in supplemented animals. The effects of two ß-AA in combination with the immunocastration technique on the performance and carcass traits were evaluated using 96 feedlot Nellore males in a randomized complete block design with two sex conditions (immunocastrated (IC) v. non-castrated (NC)) and three treatments: CON (no ß-agonists added), RH (300 mg of ractopamine hydrochloride/day, for 33 days) or ZH (80 mg of zilpaterol·hydrochloride animal/day for 30 days, removed 3 days for required withdrawal period). The trial was carried for 100 days where in the first 70 days animals did not receive ß-AA (phase 1) and during the last 30 days they were treated with ß-AA (phase 2). The performance and ultrasound measurements of longissimus muscle area (LMA), backfat thickness (BFT) and rump fat thickness (RFT) were evaluated in both phases. No sex condition v. treatment interactions were observed for any trait. The NC animals had higher average daily gain (ADG) and final BW than the IC animals, but they did not differ in dry matter intake (DMI) and feed efficiency (gain to feed). The NC animals showed greater LMA (P=0.0001) and hot carcass weight (P=0.0006), and smaller BFT (P=0.0007), RFT (P=0.0039) and percentage of kidney, pelvic and heart fat (P<0.0001) when compared with IC animals. The animals fed ZH showed greater ADG (P=0.0002), G : F (P<0.0001) and dressing per cent (P=0.0136) than those fed RH and CON diets. No differences in BW and DMI were observed. A interaction between treatment and time on feed was observed for LMA and BFT, in which the animals fed ZH diet showed greater LMA (P<0.01) and lower BFT (P<0.01) at 100 days than the animals fed RH and CON diets, whereas RH and CON diets did not differ. Immunocastration decreases muscle development and increases carcass finishing. In contrast, ß-AA increases muscle and decreases fat deposition. The ZH has a higher action on the muscle metabolism than animals fed RH diet. However, RH diet achieves a better balance because it has an intermediary performance between non-supplemented and ZH animals and does not decrease the carcass fat.

Short- and long-term clinical predictors of pharmacological cardioversion of persistent atrial fibrillation by dofetilide: A retrospective cohort study of 160 patients.

INTRODUCTION: Dofetilide is a class III antiarrhythmic prescribed to cardiovert persistent atrial fibrillation (AF) to sinus rhythm (SR). HYPOTHESIS: To determine the clinical predictors of cardioversion and readmission in persistent AF patients on dofetilide. METHODS: We analyzed 160 patients with persistent AF who were started on dofetilide and followed for 1 year. We examined age, sex, race, hypertension, diabetes, smoking, dyslipidemia, CAD, left ventricular ejection fraction (LVEF), creatinine, BMI and concomitant use of calcium channel blockers (CCB), ß-blockers in a multivariable logistic regression model. We also examined the same predictors in Cox regression model for AF-related readmission within 1 year of follow-up. RESULTS: 13.5% individuals did not convert to SR on dofetilide. 55.6% converted on the first dose and 83.1% converted by the fourth dose. In multivariable logistic models, dyslipidemia (OR: 2.4, CI: 1.12-5.16) and LVEF (OR: 3.83,CI: 1.37-10.8) were associated with failure to convert with the first dose. Female sex and LVEF also were associated with increased risk of failure to convert at all. Concomitant use of CCB associated with decreased risk of failure to convert to SR. In Cox proportional model, female sex, age <63 years and CAD were associated with increased AF readmission within 1 year. CONCLUSIONS: Dyslipidemia and LVEF <40% were associated with failure to cardiovert after first dose, and female sex and LVEF 40% were related to failure to convert at all on dofetilide in persistent AF patients. After 1-year follow-up, female sex, known CAD, and age <63 years were associated with increased AF readmissions.

The effects of increasing supplementation of zinc-amino acid complex on growth performance, carcass characteristics, and inflammatory response of beef cattle fed ractopamine hydrochloride.

Forty-two Angus crossbred steers (380 ± 5.3 kg) were enrolled in a finishing study to evaluate the influence of a supplemental Zn amino-acid complex (ZnAA; Availa-Zn) on performance and carcass characteristics of finishing steers in combination with ractopamine hydrochloride (RAC). Steers were stratified by BW into 7 pens of 6 steers each, and individual feed intake was measured. Steers were assigned to 1 of 4 treatments for 86 d (pre-RAC period): a dry-rolled corn-based diet supplemented with 60 mg Zn/kg DM from ZnSO and no supplemental ZnAA (CON; analyzed 88 mg Zn/kg DM; = 6) or CON diet supplemented with 30 (Zn30; = 12), 60 (Zn60; = 12), or 90 (Zn90; = 11) mg Zn/kg DM from ZnAA. Day 86 BW and G:F displayed a quadratic tendency ( = 0.09) with Zn60 steers being greater than the other treatments. Plasma cyclic adenosine monophosphate tended to linearly increase with increasing ZnAA ( = 0.10). On d 88, 6 of 12 steers (one of the 2 pens) receiving supplemental ZnAA was randomly selected to be supplemented with RAC at 300 mg∙steer∙d for the final 28 d of the experiment (RAC period). This created 7 final treatments: CON: no supplemental ZnAA, no RAC ( = 5); Zn30: Zn30, no RAC ( = 5); Zn30R: Zn30 + RAC ( = 6); Zn60: Zn60, no RAC ( = 6); Zn60R: Zn60 + RAC ( = 6); Zn90: Zn90, no RAC ( = 5); and Zn90R: Zn90 + RAC ( = 6). During the RAC period, as supplemental ZnAA increased within RAC-supplemented treatments, there was a linear increase in final BW, ADG, and G:F ( < 0.05). However, there was no effect of supplemental ZnAA on BW, ADG, or G:F during this period in non-RAC fed steers ( ≥ 0.44). Day 111 plasma Cu was increased, plasma Fe decreased, and leukocyte counts and serum interleukin-8 concentrations were greater ( < 0.05) in RAC-fed steers suggesting that RAC may elicit a mild inflammatory response. There was a tendency for increasing Zn supplementation to decrease plasma haptoglobin within RAC-fed steers ( = 0.07), suggesting that Zn may alter the inflammatory response. Overall, Zn60 improved growth performance during the pre-RAC period. Zinc supplemented as ZnAA appears to improve growth in combination with RAC supplementation, suggesting that Zn may enhance or support the biological function of RAC. Additionally, these results indicate that feeding RAC impacts trace mineral status, and potentially causes a non-specific inflammatory response, but further research is required to define this response.

Adenosine A(2A) receptors are necessary and sufficient to trigger memory impairment in adult mice.

BACKGROUND AND PURPOSE: Caffeine (a non-selective adenosine receptor antagonist) prevents memory deficits in aging and Alzheimer's disease, an effect mimicked by adenosine A2 A receptor, but not A1 receptor, antagonists. Hence, we investigated the effects of adenosine receptor agonists and antagonists on memory performance and scopolamine-induced memory impairment in mice. EXPERIMENTAL APPROACH: We determined whether A2 A receptors are necessary for the emergence of memory impairments induced by scopolamine and whether A2 A receptor activation triggers memory deficits in naïve mice, using three tests to assess short-term memory, namely the object recognition task, inhibitory avoidance and modified Y-maze. KEY RESULTS: Scopolamine (1.0 mg·kg(-1) , i.p.) impaired short-term memory performance in all three tests and this scopolamine-induced amnesia was prevented by the A2 A receptor antagonist (SCH 58261, 0.1-1.0 mg·kg(-1) , i.p.) and by the A1 receptor antagonist (DPCPX, 0.2-5.0 mg·kg(-1) , i.p.), except in the modified Y-maze where only SCH58261 was effective. Both antagonists were devoid of effects on memory or locomotion in naïve rats. Notably, the activation of A2 A receptors with CGS 21680 (0.1-0.5 mg·kg(-1) , i.p.) before the training session was sufficient to trigger memory impairment in the three tests in naïve mice, and this effect was prevented by SCH 58261 (1.0 mg·kg(-1) , i.p.). Furthermore, i.c.v. administration of CGS 21680 (50 nmol) also impaired recognition memory in the object recognition task. CONCLUSIONS AND IMPLICATIONS: These results show that A2 A receptors are necessary and sufficient to trigger memory impairment and further suggest that A1 receptors might also be selectively engaged to control the cholinergic-driven memory impairment.

Promotion of Wound Healing by an Agonist of Adenosine A2A Receptor Is Dependent on Tissue Plasminogen Activator.

Impaired wound healing, as it occurs in diabetes mellitus or long-term corticoid treatment, is commonly associated with disability, diminished quality of life, and high economic costs. Selective agonists of the A2A receptor subtype of adenosine, an endogenous regulator of inflammation, promote tissue repair in animal models, both healthy and with impaired healing. Plasmin-mediated proteolysis of fibrin and other matrix proteins is essential for cell migration at sites of injury. Since adenosine A2A receptor activation increases plasminogen activator release from macrophages and mast cells, we studied the effect of a selective agonist, CGS-21680, on full-thickness excisional wound closure in wild-type, urokinase plasminogen activator (uPA)-deficient, and tissue plasminogen activator (tPA)-deficient mice. Wound closure was impaired in tPA- and uPA-deficient mice as compared with wild-type mice, and topical application of CGS-21680 significantly increased the rate at which wounds closed in wild-type mice and uPA-deficient mice, but not in tPA-deficient mice. Immunostaining of tissue sections showed that tPA was present in endothelial cells and histiocytes by day 3 post-wound and also by day 6. In contrast, uPA was more prominent in these cell types only by day 6 post-wound. Our results confirm that plasminogen activation contributes to wound repair and are consistent with the hypothesis that adenosine A2A receptor activation promotes wound closure by a mechanism that depends upon tPA, but not uPA. Moreover, our results suggest that topical adenosine A2A receptor agonists may be useful in promotion of wound closure in patients with impaired wound healing.

Targeting adenosine receptors to prevent inflammatory skin diseases.

Adenosine mediates its effects through activation of a family of four G-protein-coupled receptors, named A1 , A2A , A2B and A3 . This nucleoside plays an important role in immunity and inflammation, and the A2A adenosine receptor subtype has a key role in the inhibition of inflammatory processes besides promoting wound healing. In this issue of Experimental Dermatology, Arasa et al. show that the topical application of a selective A2A agonist, CGS 21680, to mouse skin reduced epidermal hyperplasia as well as skin inflammation, similarly to topical corticoids, without side effects like skin atrophy. Rigorously following up this work is important for the development of novel treatment strategies for chronic hyperproliferative inflammatory dermatoses, such as targeting the A2A adenosine receptor family.

Topical application of the adenosine A2A receptor agonist CGS-21680 prevents phorbol-induced epidermal hyperplasia and inflammation in mice.

The nucleoside adenosine is a known regulator of immunity and inflammation that mediates, at least in part, the anti-inflammatory effect of methotrexate, an immunosuppressive agent widely used to treat autoimmune inflammatory diseases. Adenosine A2A receptors play a key role in the inhibition of the inflammatory process besides promoting wound healing. Therefore, we aimed to determine the topical effect of a selective agonist, CGS-21680, on a murine model of skin hyperplasia with a marked inflammatory component. Pretreatment with either CGS-21680 (5 µg per site) or the reference agent dexamethasone (200 µg/site) prevented the epidermal hyperplasia and inflammatory response induced by topical application of 12-O-tetradecanoylphorbol-13-acetate (TPA, 2 nmol/site) for three consecutive days. The histological analysis showed that both CGS-21680 and dexamethasone produced a marked reduction of inflammatory cell infiltrate, which correlated with diminished myeloperoxidase (MPO) activity in skin homogenates. Both treatments reduced the levels of the chemotactic mediators LTB4 and CXCL-1, and the inflammatory cytokine TNF-α, through the suppression of NFκB phosphorylation. The immunohistochemical analysis of the hyperproliferative markers cytokeratin 6 (CK6) and Ki67 revealed that while both agents inhibit the number of proliferating cells in the epidermis, CGS-21680 treatment promoted dermal fibroblasts proliferation. Consistently, increased collagen deposition in dermis was observed in tissue sections from agonist-treated mice. Our results showed that CGS 21680 efficiently prevents phorbol-induced epidermal hyperplasia and inflammation in mice without the deleterious atrophic effect of topical corticosteroids.

Nucleus tractus solitarii A(2a) adenosine receptors inhibit cardiopulmonary chemoreflex control of sympathetic outputs.

Previously we have shown that stimulation of inhibitory A1 adenosine receptors located in the nucleus tractus solitarii (NTS) attenuates cardiopulmonary chemoreflex (CCR) evoked inhibition of renal, adrenal and lumbar sympathetic nerve activity and reflex decreases in arterial pressure and heart rate. Activation of facilitatory A2a adenosine receptors, which dominate over A1 receptors in the NTS, contrastingly alters baseline activity of regional sympathetic outputs: it decreases renal, increases adrenal and does not change lumbar nerve activity. Considering that NTS A2a receptors may facilitate release of inhibitory transmitters we hypothesized that A2a receptors will act in concert with A1 receptors differentially inhibiting regional sympathetic CCR responses (adrenal>lumbar>renal). In urethane/chloralose anesthetized rats (n=38) we compared regional sympathetic responses evoked by stimulation of the CCR with right atrial injections of serotonin 5HT3 receptor agonist, phenylbiguanide, (1-8µg/kg) before and after selective stimulation, blockade or combined blockade and stimulation of NTS A2a adenosine receptors (microinjections into the NTS of CGS-21680 0.2-20pmol/50nl, ZM-241385 40pmol/100nl or ZM-241385+CGS-21680, respectively). We found that stimulation of A2a adenosine receptors uniformly inhibited the regional sympathetic and hemodynamic reflex responses and this effect was abolished by the selective blockade of NTS A2a receptors. This indicates that A2a receptor triggered inhibition of CCR responses and the contrasting shifts in baseline sympathetic activity are mediated via different mechanisms. These data implicate that stimulation of NTS A2a receptors triggers unknown inhibitory mechanism(s) which in turn inhibit transmission in the CCR pathway when adenosine is released into the NTS during severe hypotension.