Background and purpose: cAMP is a key intracellular signalling molecule that regulates multiple processes of the vertebrate skeletal muscle. We have shown that cAMP can be actively pumped out from the skeletal muscle cell. Since in other tissues, cAMP efflux had been associated with extracellular generation of adenosine, in the present study we have assessed the fate of interstitial cAMP and the existence of an extracellular cAMP-adenosine signalling pathway in skeletal muscle. Experimental approach: cAMP efflux and/or its extracellular degradation were analysed by incubating rat cultured skeletal muscle with exogenous cAMP, forskolin or isoprenaline. cAMP and its metabolites were quantified by radioassay or HPLC, respectively. Key results: Incubation of cells with exogenous cAMP was followed by interstitial accumulation of 5 0 -AMP and adenosine, a phenomenon inhibited by selective inhibitors of ecto-phosphodiesterase (DPSPX) and ecto-nucleotidase (AMPCP). Activation of adenylyl cyclase (AC) in cultured cells with forskolin or isoprenaline increased cAMP efflux and extracellular generation of 5 0 -AMP and adenosine. Extracellular cAMP-adenosine pathway was also observed after direct and receptor-dependent stimulation of AC in rat extensor muscle ex vivo. These events were attenuated by probenecid, an inhibitor of ATP binding cassette family transporters. Conclusions and implications:Our results show the existence of an extracellular biochemical cascade that converts cAMP into adenosine. The functional relevance of this extracellular signalling system may involve a feedback modulation of cellular response initiated by several G protein-coupled receptor ligands, amplifying cAMP influence to a paracrine mode, through its metabolite, adenosine.

1 The present study analyses the short-(15 min ± 2 h) and long-term (24 ± 48 h) in¯uences of calcitonin gene-related peptide (CGRP) on acetylcholinesterase (AChE) expression in the rat cultured skeletal muscle and the signal transduction events underlying CGRP actions. 2 To assess the eect of CGRP on AChE synthesis, myotubes were pre-exposed to the irreversible AChE inhibitor diisopropyl¯uorophosphate (DFP) and treated with CGRP or forskolin, an adenylyl cyclase (AC) activator. Treatment of myotubes with 1 ± 100 nM CGRP for 2 h increased by up to 42% the synthesis of catalytically active AChE with a parallel increase in the intracellular cyclic AMP. 3 The stimulation of AChE synthesis induced by CGRP was mimicked by direct activation of AC with 3 ± 30 mM forskolin. In contrast, pre-treatment of cultures with 100 nM CGRP for 20 h reduced by 37% the subsequent synthesis of AChE, resulting in a 15% decrease in total AChE activity after 48 h CGRP treatment. 4 Moreover, 24 h treatment of myotubes with 100 nM CGRP reduced by 54% the accumulation of cyclic AMP induced by a subsequent CGRP treatment. 5 These ®ndings indicate that, in skeletal muscle cells, CGRP modulates the AChE expression in a time-dependent manner, initially stimulating the enzyme synthesis through a cyclic AMP-dependent mechanism. The decreased AChE synthesis observed after long-term CGRP treatment suggests that CGRP signalling system is subject to desensitization or down-regulation, that might function as an important adaptative mechanism of the muscle ®bre in response to long-term changes in neuromuscular transmission.

Unique Objects (UNOs) are relevant for real-world applications such as anti-counterfeiting systems. In this work, cork is demonstrated as a UNO, part of the Physical Unclonability and Disorder (PUD) system. An adequate measurement kit (illumination device) and recognition method are also devised and evaluated. Natural hills and valleys of the cork are enhanced using the illumination device and the overall robustness of the recognition application inherent to UNOs is presented. The lighting device is based on grazing light and the recognition task is based on a local feature detector and descriptor called ORB - Oriented FAST (Features from Accelerated Segment Test) and Rotated BRIEF (Binary Robust Independent Elementary Features). The performance evaluation utilizes a private cork database (1500 photos of 500 cork stoppers) and three public iris databases. In the tests carried out on the illumination device, the results clearly show the success of capturing stable/repeatable features needed for the recognition task in the cork database. This achievement is also reflected in the perfect recognition score achieved in the cork database, in the intra-distance measure μ i n t r a , which gives the notion of average noise between measures, and in the inter-distance μ i n t e r which provides hints about the randomness/uniqueness of a cork. Regarding the recognition application, its effectiveness is further tested using the iris databases. Regardless of the fact that the recognition algorithm was not designed for the iris recognition problem, the results show that the proposed approach is capable of competing with the techniques found in the literature specially designed for iris recognition. Furthermore, the evaluation shows that the three requirements that constitute a UNO (Disorder, Operability, and Unclonability) are fulfilled, thus supporting the main assertion of this work: that cork is a UNO.

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Interest in robotics field as a teaching tool to promote the STEM areas has grown in the past years. The search for solutions to promote robotics is a major challenge and the use of real robots always increases costs. An alternative is the use of a simulator. The construction of a simulator related with the Portuguese Autonomous Driving Competition using Gazebo as 3D simulator and ROS as a middleware connection to promote, attract, and enthusiasm university students to the mobile robotics challenges is presented. It is intended to take advantage of a competitive mindset to overcome some obstacles that appear to students when designing a real system. The proposed simulator focus on the autonomous driving competition task, such as semaphore recognition, localization, and motion control. An evaluation of the simulator is also performed, leading to an absolute error of 5.11% and a relative error of 2.76% on best case scenarios relating to the odometry tests, an accuracy of 99.37% regarding to the semaphore recognition tests, and an average error of 1.8 pixels for the FOV tests performed.

Wine counterfeiting is a major problem worldwide. Within this context, an approach to the problem of discerning original wine bottles from forged ones is the use of natural features present in the product, object and/or material (using it "as is"). The proposed application uses the cork stopper as a unique fingerprint, combined with state of the art image processing techniques to achieve individual object recognition and smartphones as the authentication equipment. The anti-counterfeiting scheme is divided into two phases: an enrollment phase, where every bottle is registered in a database using a photo of its cork stopper inside the bottle; and a verification phase, where an end-user/retailer captures a photo of the cork stopper using a regular smartphone, compares the photo with the previously-stored one and retrieves it if the wine bottle was previously registered. To evaluate the performance of the proposed application, two datasets of natural/agglomerate cork stoppers were built, totaling 1000 photos. The worst case results show a 100% precision ratio, an accuracy of 99.94% and a recall of 94.00%, using different smartphones. The perfect score in precision is a promising result, proving that this system can be applied to the prevention of wine counterfeiting and consumer/retailer security when purchasing a wine bottle.

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