Paulo Aguiar

Kinetochore motors drive congression of peripheral polar chromosomes by overcoming random arm-ejection forces

Accurate chromosome segregation during cell division in metazoans relies on proper chromosome congression at the equator. Chromosome congression is achieved after bi-orientation to both spindle poles shortly after nuclear envelope breakdown, or by the coordinated action of motor proteins that slide misaligned chromosomes along pre-existing spindle microtubules. These proteins include the minus-end-directed kinetochore motor dynein, and the plus-end-directed motors CENP-E at kinetochores and chromokinesins on chromosome arms. However, how these opposite and spatially distinct activities are coordinated to drive chromosome congression remains unknown. Here we used RNAi, chemical inhibition, kinetochore tracking and laser microsurgery to uncover the functional hierarchy between kinetochore and arm-associated motors, exclusively required for congression of peripheral polar chromosomes in human cells. We show that dynein poleward force counteracts chromokinesins to prevent stabilization of immature/incorrect end-on kinetochore–microtubule attachments and random ejection of polar chromosomes. At the poles, CENP-E becomes dominant over dynein and chromokinesins to bias chromosome ejection towards the equator. Thus, dynein and CENP-E at kinetochores drive congression of peripheral polar chromosomes by preventing arm-ejection forces mediated by chromokinesins from working in the wrong direction.

Classification of breast cancer histology images using Convolutional Neural Networks

Breast cancer is one of the main causes of cancer death worldwide. The diagnosis of biopsy tissue with hematoxylin and eosin stained images is non-trivial and specialists often disagree on the final diagnosis. Computer-aided Diagnosis systems contribute to reduce the cost and increase the efficiency of this process. Conventional classification approaches rely on feature extraction methods designed for a specific problem based on field-knowledge. To overcome the many difficulties of the feature-based approaches, deep learning methods are becoming important alternatives. A method for the classification of hematoxylin and eosin stained breast biopsy images using Convolutional Neural Networks (CNNs) is proposed. Images are classified in four classes, normal tissue, benign lesion, in situ carcinoma and invasive carcinoma, and in two classes, carcinoma and non-carcinoma. The architecture of the network is designed to retrieve information at different scales, including both nuclei and overall tissue organization. This design allows the extension of the proposed system to whole-slide histology images. The features extracted by the CNN are also used for training a Support Vector Machine classifier. Accuracies of 77.8% for four class and 83.3% for carcinoma/non-carcinoma are achieved. The sensitivity of our method for cancer cases is 95.6%.

Shared Knowledge or Shared Affordances? Insights from an Ecological Dynamics Approach to Team Coordination in Sports

Previous research has proposed that team coordination is based on shared knowledge of the performance context, responsible for linking teammates’ mental representations for collective, internalized action solutions. However, this representational approach raises many questions including: how do individual schemata of team members become reformulated together? How much time does it take for this collective cognitive process to occur? How do different cues perceived by different individuals sustain a general shared mental representation? This representational approach is challenged by an ecological dynamics perspective of shared knowledge in team coordination. We argue that the traditional shared knowledge assumption is predicated on ‘knowledge about’ the environment, which can be used to share knowledge and influence intentions of others prior to competition. Rather, during competitive performance, the control of action by perceiving surrounding informational constraints is expressed in ‘knowledge of’ the environment. This crucial distinction emphasizes perception of shared affordances (for others and of others) as the main communication channel between team members during team coordination tasks. From this perspective, the emergence of coordinated behaviours in sports teams is based on the formation of interpersonal synergies between players resulting from collective actions predicated on shared affordances.

OpenControl : A free opensource software for video tracking and automated control of behavioral mazes

Operant animal behavioral tests require the interaction of the subject with sensors and actuators distributed in the experimental environment of the arena. In order to provide user independent reliable results and versatile control of these devices it is vital to use an automated control system. Commercial systems for control of animal mazes are usually based in software implementations that restrict their application to the proprietary hardware of the vendor. In this paper we present OpenControl: an opensource Visual Basic software that permits a Windows-based computer to function as a system to run fully automated behavioral experiments. OpenControl integrates video-tracking of the animal, definition of zones from the video signal for real-time assignment of animal position in the maze, control of the maze actuators from either hardware sensors or from the online video tracking, and recording of experimental data. Bidirectional communication with the maze hardware is achieved through the parallel-port interface, without the need for expensive AD-DA cards, while video tracking is attained using an inexpensive Firewire digital camera. OpenControl Visual Basic code is structurally general and versatile allowing it to be easily modified or extended to fulfill specific experimental protocols and custom hardware configurations. The Visual Basic environment was chosen in order to allow experimenters to easily adapt the code and expand it at their own needs.

Feedback control of chromosome separation by a midzone Aurora B gradient

Taking a check on chromosome spacing Animal cells divide by mitosis. Chromosomes become condensed and congregate on the mitotic spindle in the center of the cell—the midzone. The spindle then separates sister chromosomes, pulling them to opposite ends of the cell, ready to form new daughter nuclei. Afonso et al. now show that chromosome separation is monitored by the level of midzone-associated Aurora B kinase activity (see the Perspective by Hadders and Lens). This process ensures that daughter nuclei only reassemble after sister chromosomes have successfully separated. Science, this issue p. 332; see also p. 265 A mitotic spindle midzone-associated Aurora B gradient monitors chromosome separation during cell division. [Also see Perspective by Hadders and Lens] Accurate chromosome segregation during mitosis requires the physical separation of sister chromatids before nuclear envelope reassembly (NER). However, how these two processes are coordinated remains unknown. Here, we identified a conserved feedback control mechanism that delays chromosome decondensation and NER in response to incomplete chromosome separation during anaphase. A midzone-associated Aurora B gradient was found to monitor chromosome position along the division axis and to prevent premature chromosome decondensation by retaining Condensin I. PP1/PP2A phosphatases counteracted this gradient and promoted chromosome decondensation and NER. Thus, an Aurora B gradient appears to mediate a surveillance mechanism that prevents chromosome decondensation and NER until effective separation of sister chromatids is achieved. This allows the correction and reintegration of lagging chromosomes in the main nuclei before completion of NER.

Field dimension and skill level constrain team tactical behaviours in small-sided and conditioned games in football.

Abstract This study analysed the influence of field dimension and players’ skill level on collective tactical behaviours during small-sided and conditioned games (SSCGs). Positioning and displacement data were collected using global positioning systems (15 Hz) during SSCGs (Gk+4 v. 4+Gk) played by two groups of participants (NLP- national-level and RLP-regional-level players) on different field dimensions (small: 36.8 × 23.8 m; intermediate: 47.3 × 30.6 and large: 57.8 × 37.4 m). Team tactical performance was assessed through established dynamic team variables (effective playing space, playing length per width ratio and team separateness) and nonlinear signal processing techniques (sample entropy of distances to nearest opponents and the teams’ centroids’ mutual information). Results showed that the effective playing space and team separateness increased significantly with pitch size regardless of participant skill level (P < 0.001, η2 = 0.78 and P < 0.001, η2 = 0.65, respectively). Playing length per width ratio increased with pitch size for the NLP but was maintained at a relatively constant level by RLP across treatments indicating different playing shapes. There was significantly more irregularity in distances to nearest opponents for the NLP in small (P = 0.003) and intermediate fields (P = 0.01). Findings suggest that tactical behaviours in SSCGs are constrained by field size and skill level, which need to be considered by coaches when designing training practices.

Genes involved in centrosome-independent mitotic spindle assembly in Drosophila S2 cells

Significance The mitotic spindle, a structure composed primarily of microtubules, guides the segregation of DNA during cell division. In somatic animal cells, centrosomes (microtubule nucleating structures) reside near the mitotic spindle poles. However, germ cells lack centrosomes, and even somatic cells can execute cell division if centrosome function is compromised. This study on Drosophila cells reports a whole-genome RNAi screen for genes involved in spindle assembly in the absence of functional centrosomes. The results show that spindle assembly pathways with and without centrosomes involve an essentially identical set of genes, demonstrating the constitutive nature of centrosome-independent spindle assembly. However, certain gene knockdowns show distinct phenotypes when centrosomes are absent, thus revealing how spindles adapt to the presence or absence of centrosomes. Animal mitotic spindle assembly relies on centrosome-dependent and centrosome-independent mechanisms, but their relative contributions remain unknown. Here, we investigated the molecular basis of the centrosome-independent spindle assembly pathway by performing a whole-genome RNAi screen in Drosophila S2 cells lacking functional centrosomes. This screen identified 197 genes involved in acentrosomal spindle assembly, eight of which had no previously described mitotic phenotypes and produced defective and/or short spindles. All 197 genes also produced RNAi phenotypes when centrosomes were present, indicating that none were entirely selective for the acentrosomal pathway. However, a subset of genes produced a selective defect in pole focusing when centrosomes were absent, suggesting that centrosomes compensate for this shape defect. Another subset of genes was specifically associated with the formation of multipolar spindles only when centrosomes were present. We further show that the chromosomal passenger complex orchestrates multiple centrosome-independent processes required for mitotic spindle assembly/maintenance. On the other hand, despite the formation of a chromosome-enriched RanGTP gradient, S2 cells depleted of RCC1, the guanine-nucleotide exchange factor for Ran on chromosomes, established functional bipolar spindles. Finally, we show that cells without functional centrosomes have a delay in chromosome congression and anaphase onset, which can be explained by the lack of polar ejection forces. Overall, these findings establish the constitutive nature of a centrosome-independent spindle assembly program and how this program is adapted to the presence/absence of centrosomes in animal somatic cells.

Resistive switching and activity-dependent modifications in Ni-doped graphene oxide thin films

The resistive switching (RS) mechanism in Ni-doped graphene oxide (GO) devices is studied. We found that RS depends strongly on the fabrication method of the GO sheet and on the electrode material. Resistive switching in GO-devices can be caused by the diffusion of ions from metallic electrode or by the migration of oxygen groups, depending on the fabrication process. We also show that GO-based structures possess activity-dependent modification capabilities, emphasized by the increase/decrease of device conductance after consecutive voltage sweeps of opposite polarity. Our results allow a better understanding of bipolar RS, towards future non-volatile memories and neuromorphic systems.

Axon Diversity of Lamina I Local-Circuit Neurons in the Lumbar Spinal Cord

Spinal lamina I is a key area for relaying and integrating information from nociceptive primary afferents with various other sources of inputs. Although lamina I projection neurons have been intensively studied, much less attention has been given to local‐circuit neurons (LCNs), which form the majority of the lamina I neuronal population. In this work the infrared light‐emitting diode oblique illumination technique was used to visualize and label LCNs, allowing reconstruction and analysis of their dendritic and extensive axonal trees. We show that the majority of lamina I neurons with locally branching axons fall into the multipolar (with ventrally protruding dendrites) and flattened (dendrites limited to lamina I) somatodendritic categories. Analysis of their axons revealed that the initial myelinated part gives rise to several unmyelinated small‐diameter branches that have a high number of densely packed, large varicosities and an extensive rostrocaudal (two or three segments), mediolateral, and dorsoventral (reaching laminae III–IV) distribution. The extent of the axon and the occasional presence of long, solitary branches suggest that LCNs may also form short and long propriospinal connections. We also found that the distribution of axon varicosities and terminal field locations show substantial heterogeneity and that a substantial portion of LCNs is inhibitory. Our observations indicate that LCNs of lamina I form intersegmental as well as interlaminar connections and may govern large numbers of neurons, providing anatomical substrate for rostrocaudal “processing units” in the dorsal horn. J. Comp. Neurol. 521:2719–2741, 2013.

Inflammatory pain disrupts the orbitofrontal neuronal activity and risk-assessment performance in a rodent decision-making task.

Summary Multielectrode recordings in awake behaving rats show that the prefrontal encoding of risk in a rodent gambling task is compromised by the onset of an inflammatory pain model. Abstract It has been recently described that disruption of the neural mechanisms of emotion‐based decision making occurs in both chronic pain patients and in animal models of pain; moreover, it also has been shown that chronic pain causes morphological and functional changes in the prefrontal cortex that may be crucial for this decision‐making dysfunction. However, it is not known whether pain alone is capable of altering the neuronal encoding of decision exhibited by prefrontal neurons. We have previously shown that naïve animals have risk‐averse performance in the rodent gambling task, whereas chronic pain animals reverse their choice preference and become risk prone. Using this paradigm, we chronically implanted arrays of multielectrodes and recorded from neuronal ensembles in the orbitofrontal cortex of freely moving animals performing 4 sessions of the rodent gambling task: 2 in control conditions and 2 after the onset of inflammatory pain induced by complete Freund’s adjuvant injection. Our results show that the instantaneous neuronal firing rate was correlated with the probability of choosing a specific lever in 62.5% of the neurons; however, although in the control sessions 61% of the neurons encoded the reward magnitude, after the pain onset only 16% of the neurons differentiated small from large rewards. Moreover, we found that the fraction of risk‐sensitive neurons recorded in each session predicted the overall risk bias of the animal. Our data suggest that orbitofrontal cortex encoding of risk preference is compromised in chronic pain animals.