Agonists for 13 trace amine-associated receptors provide insight into the molecular basis of odor selectivity

Trace amine-associated receptors (TAARs) are vertebrate olfactory receptors. However, ligand recognition properties of TAARs remain poorly understood, as most are "orphan receptors" without known agonists. Here, we identify the first ligands for many rodent TAARs and classify these receptors into two subfamilies based on the phylogeny and binding preference for primary or tertiary amines. Some mouse and rat orthologs have similar response profiles, although independent Taar7 gene expansions led to highly related receptors with altered ligand specificities. Using chimeric TAAR7 receptors, we identified an odor contact site in transmembrane helix III that functions as a selectivity filter. Homology models based on the β(2) adrenergic receptor structure indicate spatial proximity of this site to the ligand. Gain-of-function mutations at this site created olfactory receptors with radically altered odor recognition properties. These studies provide new TAAR ligands, valuable tools for studying receptor function, and general insights into the molecular pharmacology of G protein-coupled receptors.     

P2X7 receptor-Pannexin1 complex: pharmacology and signaling

Pannexin 1 (Panx1), an ortholog to invertebrate innexin gap junctions, has recently been proposed to be the pore induced by P2X(7) receptor (P2X(7)R) activation. We explored the pharmacological action of compounds known to block gap junctions on Panx1 channels activated by the P2X(7)R and the mechanisms involved in the interaction between these two proteins. Whole cell recordings revealed distinct P2X(7)R and Panx1 currents in response to agonists. Activation of Panx1 currents following P2X(7)R stimulation or by membrane depolarization was blocked by Panx1 small-interfering RNA (siRNA) and with mefloquine > carbenoxolone > flufenamic acid. Incubation of cells with KN-62, a P2X(7)R antagonist, prevented current activation by 2'(3')-O-(4-benzoylbenzoyl)adenosine 5'-triphosphate (BzATP). Membrane permeabilization to dye induced by BzATP was also prevented by Panx1 siRNA and by carbenoxolone and mefloquine. Membrane permeant (TAT-P2X(7)) peptides, provided evidence that the Src homology 3 death domain of the COOH-terminus of the P2X(7)R is involved in the initial steps of the signal transduction events leading to Panx1 activation and that a Src tyrosine kinase is likely involved in this process. Competition assays indicated that 20 muM TAT-P2X(7) peptide caused 50% reduction in Src binding to the P2X(7)R complex. Src tyrosine phosphorylation following BzATP stimulation was reduced by KN-62, TAT-P2X(7) peptide, and by the Src tyrosine inhibitor PP2 and these compounds prevented both large-conductance Panx1 currents and membrane permeabilization. These results together with the lack Panx1 tyrosine phosphorylation in response to P2X(7)R stimulation indicate the involvement of an additional molecule in the tyrosine kinase signal transduction pathway mediating Panx1 activation through the P2X(7)R.     

Polychaete assemblages inhabiting intertidal soft bottoms associated with mangrove systems in Coiba National Park (East Pacific,Panama)

The Coiba National Park is located off the Pacific coast of Panama andincludes several small islands and a larger one which gives its name to the park. Anumber of rivers in Coiba Island have well developed mangrove systems attheir mouths and two of these were selected for the present study: SantaCruz and El Gambute. The first one comprises an assemblage of Rhizopora mangle, Laguncularia racemosa and Pellicierarhizoporae, and shows no evidence of recent human activity. The secondone is a smaller forest, solely composed of short Rhizoporamangle trees, and is recovering from the impact of human activities.During 1997, the tidal flats associated with each system were studied. Toasses the polychaete distribution within each tidal flat, three transects werearranged from the low to the upper intertidal level. Every transect hadthree sampling sites: the first one was adjacent to the low water level, thesecond one at an intermediate point, and the third one close to the highwater level and the mangrove line. At each site, three randomly chosensamples of sediment were taken for faunal study and a further one wascollected for sedimentology study.The polychaetes were identifiedto species or genus level. For each flat, a matrix summarizing the actualnumber of individuals of each species in every sample was created, andpopulation density, species richness and diversity were computed for eachsite. After matrix data had been transformed, a similarity analysis of thesamples was carried out.     

Enzymatic hydrolysis of wheat straw: Kinetic analysis

Pretreated wheat straw was enzymatically hydrolyzed in 250-ml flasks using the culture filtrate of Trichoderma reesei QM-9414. The influence of the initial enzymatic activity in the liquid phase was studied. The unreacted core model was used to analyze the experimental data obtained at 40, 46 and 50°C. The model adequately describes the data for hydrolysis times lower than 10 h.     

Osteopromotion of Biphasic Calcium Phosphate granules in critical size defects after osteonecrosis induced by focal heating insults

Heat-induced osteonecrosis represents a simple, rapid, and inexpensive method for reproducing the effects of bone disease. In the present study, we employed this technique to induce osteonecrosis in femoral defects in rabbits and assessed the efficacy of treatment using Biphasic Calcium Phosphate (BCP) granules (MBCP+?, Biomatlante SA). After 3 weeks, the osteopromotion effects of BCP granules could be statistically proven (P < 0.05) through image analysis of newly formed bone in osteonecrosed sites containing BCP granules when compared to empty control sites. Increasing mature and woven bone presence was observed after 6 and 12 weeks, forming new trabeculae in necrosed site. Significant statistical differences were evidenced at each time between empty necrosed and filled necrosed defects in terms of new bone volume.     

Dynamics of spontaneous activity in random networks with multiple neuron subtypes and synaptic noise

During slow-wave sleep, brain electrical activity is dominated by the slow (< 1 Hz) electroencephalogram (EEG) oscillations characterized by the periodic transitions between active (or Up) and silent (or Down) states in the membrane voltage of the cortical and thalamic neurons. Sleep slow oscillation is believed to play critical role in consolidation of recent memories. Past computational studies, based on the Hodgkin-Huxley type neuronal models, revealed possible intracellular and network mechanisms of the neuronal activity during sleep, however, they failed to explore the large-scale cortical network dynamics depending on collective behavior in the large populations of neurons. In this new study, we developed a novel class of reduced discrete time spiking neuron models for large-scale network simulations of wake and sleep dynamics. In addition to the spiking mechanism, the new model implemented nonlinearities capturing effects of the leak current, the Ca²⁺ dependent K⁺ current and the persistent Na⁺ current that were found to be critical for transitions between Up and Down states of the slow oscillation. We applied the new model to study large-scale two-dimensional cortical network activity during slow-wave sleep. Our study explained traveling wave dynamics and characteristic synchronization properties of transitions between Up and Down states of the slow oscillation as observed in vivo in recordings from cats. We further predict a critical role of synaptic noise and slow adaptive currents for spike sequence replay as found during sleep related memory consolidation.     

The Effect of Persantin on Intercoronary Collateral Circulation and Survival During Gradual Experimental Coronary Occlusion: A Preliminary Report

Twenty-five dogs were exposed to gradual coronary occlusion by placing Ameroid constrictors around the origins of the left circumflex and anterior descending coronary arteries. Previous experiments have demonstrated that these constrictors absorb water and, over a period of three weeks, narrow the cross-sectional area of the two arteries to 50% or less, and consequently cause the death of 80% of the experimental animals. Twelve of the 25 animals were fed 50 mg. of Persantin three times a day by mouth commencing one day before the operative procedure. Determinations of the concentration of the drug in the blood revealed a level consistent with that obtained in humans after the administration of therapeutic doses. Eleven of the 13 control animals died in the three-month experimental period while only six of the 12 treated animals expired. Injections of Schlesinger mass in all animals dying or killed following the experimental period demonstrated that Persantin significantly accelerated the development of intercoronary anastomoses in the treated group, and in the surviving animals produced a rich anastomotic network much in excess of that seen in the surviving animals in the control series that were exposed to hypoxia alone. On the basis of these experimental findings, it is suggested that Persantin may favourably alter the prognosis of many patients with coronary artery disease.     

Quantification of protopectinase SE, an endopolygalacturonase with pectin-releasing activity from Geotrichum klebahnii

Pectin releasing activity of protopectinase SE (PPase-SE) from Geotrichum klebahnii (= G. penicillatum = Trichosporon penicillatum) ATCC 42397 was determined using different batches of lemon protopectin as substrate. Results obtained showed a high degree of variability depending on the batch of protopectin used. As PPase-SE also shows polygalacturonase (PGase) activity, a method for the assay of this activity was optimised. The best assay conditions were: substrate (polygalacturonic acid) concentration of 2.0 g 1^–1, reaction time of 10 min and up to 0.17 PGase units per test tube.     

Association of genetic variants rs641153 (CFB), rs2230199 (C3), and rs1410996 (CFH) with age-related macular degeneration in a Brazilian population

This study aimed to investigate the association among genetic variants of the complement pathway CFB R32Q (rs641153), C3 R102G (rs2230199), and CFH (rs1410996) with age-related macular degeneration (AMD) in a sample of the Brazilian population. In a case-control study, 484 AMD patients were classified according to the clinical age-related maculopathy grading system (CARMS) and compared to 479 unrelated controls. The genetic variants rs1410996 of complement H (CFH), rs641153 of complement factor B (CFB), and rs2230199 of complement 3 (C3) were evaluated through polymerase chain reaction (PCR) and direct sequencing. The associations between single nucleotide polymorphisms (SNPs) and AMD, adjusted by age, were assessed by using logistic regression models. A statistically significant association was observed between AMD risk and rs2230199 variant with an OR of 2.01 (P  = 0.0002) for CG individuals compared to CC individuals. Regarding the comparison of advanced AMD versus the control group, the OR was 2.12 (P = 0.0036) for GG versus AA genotypes for rs1410996 variant. Similarly, the OR for rs2230199 polymorphism was 2.3034 (P  = 5.47^e-05) when comparing CG individuals to CC carriers. In contrast, the rs641153 variant showed a significant protective effect against advanced AMD for GA versus GG genotype (OR = 0.4406; P  = 0.0019). When comparing wet AMD versus controls, a significant association was detected for rs1410996 variant (OR = 2.16; P  = 0.0039) comparing carriers of the homozygous GG versus AA genotype, as well as in the comparisons of GG (OR = 3.0713; P  = 0.0046) and CG genotypes (OR = 2.2249; P  = 0.0002) versus CC genotype for rs2230199 variant, respectively. The rs641153 variant granted a significant protective effect against wet AMD for GA versus GG genotypes (OR = 0.4601; P  = 0.0044). Our study confirmed the risk association between rs2230199 and rs1410996 variants and AMD, and the protective role against AMD for rs641153 variant.