The circadian response of intrinsically photosensitive retinal ganglion cells

Intrinsically photosensitive retinal ganglion cells (ipRGC) signal environmental light level to the central circadian clock and contribute to the pupil light reflex. It is unknown if ipRGC activity is subject to extrinsic (central) or intrinsic (retinal) network-mediated circadian modulation during light entrainment and phase shifting. Eleven younger persons (18–30 years) with no ophthalmological, medical or sleep disorders participated. The activity of the inner (ipRGC) and outer retina (cone photoreceptors) was assessed hourly using the pupil light reflex during a 24 h period of constant environmental illumination (10 lux). Exogenous circadian cues of activity, sleep, posture, caffeine, ambient temperature, caloric intake and ambient illumination were controlled. Dim-light melatonin onset (DLMO) was determined from salivary melatonin assay at hourly intervals, and participant melatonin onset values were set to 14 h to adjust clock time to circadian time. Here we demonstrate in humans that the ipRGC controlled post-illumination pupil response has a circadian rhythm independent of external light cues. This circadian variation precedes melatonin onset and the minimum ipRGC driven pupil response occurs post melatonin onset. Outer retinal photoreceptor contributions to the inner retinal ipRGC driven post-illumination pupil response also show circadian variation whereas direct outer retinal cone inputs to the pupil light reflex do not, indicating that intrinsically photosensitive (melanopsin) retinal ganglion cells mediate this circadian variation.

Influence of culture conditions on the molecular signature of mesenchymal stem cells

Cell based therapies require cells capable of self renewal and differentiation, and a prerequisite is the ability to prepare an effective dose of ex vivo expanded cells for autologous transplants. The in vivo identification of a source of physiologically relevant cell types suitable for cell therapies is therefore an integral part of tissue engineering. Bone marrow is the most easily accessible source of mesenchymal stem cells (MSCs), and harbours two distinct populations of adult stem cells; namely hematopoietic stem cells (HSCs) and bone mesenchymal stem cells (BMSCs). Unlike HSCs, there are yet no rigorous criteria for characterizing BMSCs. Changing understanding about the pluripotency of BMSCs in recent studies has expanded their potential application; however, the underlying molecular pathways which impart the features distinctive to BMSCs remain elusive. Furthermore, the sparse in vivo distribution of these cells imposes a clear limitation to their in vitro study. Also, when BMSCs are cultured in vitro there is a loss of the in vivo microenvironment which results in a progressive decline in proliferation potential and multipotentiality. This is further exacerbated with increased passage number, characterized by the onset of senescence related changes. Accordingly, establishing protocols for generating large numbers of BMSCs without affecting their differentiation potential is necessary. The principal aims of this thesis were to identify potential molecular factors for characterizing BMSCs from osteoarthritic patients, and also to attempt to establish culture protocols favourable for generating large number of BMSCs, while at the same time retaining their proliferation and differentiation potential. Previously published studies concerning clonal cells have demonstrated that BMSCs are heterogeneous populations of cells at various stages of growth. Some cells are higher in the hierarchy and represent the progenitors, while other cells occupy a lower position in the hierarchy and are therefore more committed to a particular lineage. This feature of BMSCs was made evident by the work of Mareddy et al., which involved generating clonal populations of BMSCs from bone marrow of osteoarthritic patients, by a single cell clonal culture method. Proliferation potential and differentiation capabilities were used to group cells into fast growing and slow growing clones. The study presented here is a continuation of the work of Mareddy et al. and employed immunological and array based techniques to identify the primary molecular factors involved in regulating phenotypic characteristics exhibited by contrasting clonal populations. The subtractive immunization (SI) was used to generate novel antibodies against favourably expressed proteins in the fast growing clonal cell population. The difference between the clonal populations at the transcriptional level was determined using a Stem Cell RT2 Profiler TM PCR Array which focuses on stem cell pathway gene expression. Monoclonal antibodies (mAb) generated by SI were able to effectively highlight differentially expressed antigenic determinants, as was evident by Western blot analysis and confocal microscopy. Co-immunoprecipitation, followed by mass spectroscopy analysis, identified a favourably expressed protein as the cytoskeletal protein vimentin. The stem cell gene array highlighted genes that were highly upregulated in the fast growing clonal cell population. Based on their functions these genes were grouped into growth factors, cell fate determination and maintenance of embryonic and neural stem cell renewal. Furthermore, on a closer analysis it was established that the cytoskeletal protein vimentin and nine out of ten genes identified by gene array were associated with chondrogenesis or cartilage repair, consistent with the potential role played by BMSCs in defect repair and maintaining tissue homeostasis, by modulating the gene expression pattern to compensate for degenerated cartilage in osteoarthritic tissues. The gene array also presented transcripts for embryonic lineage markers such as FOXA2 and Sox2, both of which were significantly over expressed in fast growing clonal populations. A recent groundbreaking study by Yamanaka et al imparted embryonic stem cell (ESCs) -like characteristic to somatic cells in a process termed nuclear reprogramming, by the ectopic expression of the genes Sox2, cMyc and Oct4. The expression of embryonic lineage markers in adult stem cells may be a mechanism by which the favourable behaviour of fast growing clonal cells is determined and suggests a possible active phenomenon of spontaneous reprogramming in fast growing clonal cells. The expression pattern of these critical molecular markers could be indicative of the competence of BMSCs. For this reason, the expression pattern of Sox2, Oct4 and cMyc, at various passages in heterogeneous BMSCs population and tissue derived cells (osteoblasts and chondrocytes), was investigated by a real-time PCR and immunoflourescence staining. A strong nuclear staining was observed for Sox2, Oct4 and cMyc, which gradually weakened accompanied with cytoplasmic translocation after several passage. The mRNA and protein expression of Sox2, Oct4 and cMyc peaked at the third passage for osteoblasts, chondrocytes and third passage for BMSCs, and declined with each subsequent passage, indicating towards a possible mechanism of spontaneous reprogramming. This study proposes that the progressive decline in proliferation potential and multipotentiality associated with increased passaging of BMSCs in vitro might be a consequence of loss of these propluripotency factors. We therefore hypothesise that the expression of these master genes is not an intrinsic cell function, but rather an outcome of interaction of the cells with their microenvironment; this was evident by the fact that when removed from their in vivo microenvironment, BMSCs undergo a rapid loss of stemness after only a few passages. One of the most interesting aspects of this study was the integration of factors in the culture conditions, which to some extent, mimicked the in vivo microenvironmental niche of the BMSCs. A number of studies have successfully established that the cellular niche is not an inert tissue component but is of prime importance. The total sum of stimuli from the microenvironment underpins the complex interplay of regulatory mechanisms which control multiple functions in stem cells most importantly stem cell renewal. Therefore, well characterised factors which affect BMSCs characteristics, such as fibronectin (FN) coating, and morphogens such as FGF2 and BMP4, were incorporated into the cell culture conditions. The experimental set up was designed to provide insight into the expression pattern of the stem cell related transcription factors Sox2, cMyc and Oct4, in BMSCs with respect to passaging and changes in culture conditions. Induction of these pluripotency markers in somatic cells by retroviral transfection has been shown to confer pluripotency and an ESCs like state. Our study demonstrated that all treatments could transiently induce the expression of Sox2, cMyc and Oct4, and favourably affect the proliferation potential of BMSCs. The combined effect of these treatments was able to induce and retain the endogenous nuclear expression of stem cell transcription factors in BMSCs over an extended number of in vitro passages. Our results therefore suggest that the transient induction and manipulation of endogenous expression of transcription factors critical for stemness can be achieved by modulating the culture conditions; the benefit of which is to circumvent the need for genetic manipulations. In summary, this study has explored the role of BMSCs in the diseased state of osteoarthritis, by employing transcriptional profiling along with SI. In particular this study pioneered the use of primary cells for generating novel antibodies by SI. We established that somatic cells and BMSCs have a basal level of expression of pluripotency markers. Furthermore, our study indicates that intrinsic signalling mechanisms of BMSCs are intimately linked with extrinsic cues from the microenvironment and that these signals appear to be critical for retaining the expression of genes to maintain cell stemness in long term in vitro culture. This project provides a basis for developing an “artificial niche” required for reversion of commitment and maintenance of BMSC in their uncommitted homeostatic state.

Particulate texture image analysis with applications

Texture analysis and textural cues have been applied for image classification, segmentation and pattern recognition. Dominant texture descriptors include directionality, coarseness, line-likeness etc. In this dissertation a class of textures known as particulate textures are defined, which are predominantly coarse or blob-like. The set of features that characterise particulate textures are different from those that characterise classical textures. These features are micro-texture, macro-texture, size, shape and compaction. Classical texture analysis techniques do not adequately capture particulate texture features. This gap is identified and new methods for analysing particulate textures are proposed. The levels of complexity in particulate textures are also presented ranging from the simplest images where blob-like particles are easily isolated from their back- ground to the more complex images where the particles and the background are not easily separable or the particles are occluded. Simple particulate images can be analysed for particle shapes and sizes. Complex particulate texture images, on the other hand, often permit only the estimation of particle dimensions. Real life applications of particulate textures are reviewed, including applications to sedimentology, granulometry and road surface texture analysis. A new framework for computation of particulate shape is proposed. A granulometric approach for particle size estimation based on edge detection is developed which can be adapted to the gray level of the images by varying its parameters. This study binds visual texture analysis and road surface macrotexture in a theoretical framework, thus making it possible to apply monocular imaging techniques to road surface texture analysis. Results from the application of the developed algorithm to road surface macro-texture, are compared with results based on Fourier spectra, the auto- correlation function and wavelet decomposition, indicating the superior performance of the proposed technique. The influence of image acquisition conditions such as illumination and camera angle on the results was systematically analysed. Experimental data was collected from over 5km of road in Brisbane and the estimated coarseness along the road was compared with laser profilometer measurements. Coefficient of determination R2 exceeding 0.9 was obtained when correlating the proposed imaging technique with the state of the art Sensor Measured Texture Depth (SMTD) obtained using laser profilometers.

Order of instruction effects - do they make a difference when teaching senior chemistry with computer based visualizations?

This study investigated whether conceptual development is greater if students learning senior chemistry hear teacher explanations and other traditional teaching approaches first then see computer based visualizations or vice versa. Five Canadian chemistry classes, taught by three different teachers, studied the topics of Le Chatelier’s Principle and dynamic chemical equilibria using scientific visualizations with the explanation and visualizations in different orders. Conceptual development was measured using a 12 item test based on the Chemistry Concepts Inventory. Data was obtained about the students’ abilities, learning styles (auditory, visual or kinesthetic) and sex, and the relationships between these factors and conceptual development due to the teaching sequences were investigated. It was found that teaching sequence is not important in terms of students’ conceptual learning gains, across the whole cohort or for any of the three subgroups.

Which way from here? The spatiality of maps & navigation in first-person single-player video games

Player experience of spatiality in first-person, single-player games is informed by the maps and navigational aids provided by the game. This project uses textual analysis to examine the way these maps and navigational aids inform the experience of spatiality in Fallout 3, BioShock and BioShock 2. Spatiality is understood as trialectic, incorporating perceived, conceived and lived space, drawing on the work of Henri Lefebvre and Edward Soja. The most prominent elements of the games’ maps and navigational aids are analysed in terms of how they inform players’ experience of the games’ spaces. In particular this project examines the in-game maps these games incorporate, the waypoint navigation and fast-travel systems in Fallout 3, and the guide arrow and environmental cues in the BioShock games.

Assessment of preterm infants' readiness to commence breastfeeding

Preterm infants commence breastfeeding when health-care professionals deem them to be ready. However, the optimal timing for commencement of breastfeeding is unclear. Currently, there is little guidance for neonatal care providers to decide when to initiate breastfeeding among preterm infants. A mixed-methods study was conducted to develop and test the Preterm Sucking Readiness (PTSR) scale in four phases. The first phase involved a chart audit to explore the use of age as a criterion by investigating when preterm infants meet feeding milestones as well as other factors that may affect an infant’s readiness to engage in nutritive sucking behaviour. The second phase utilised focus groups to explore and define how neonatal care providers decide when to commence breastfeeding. To gain consensus on the criteria mentioned by the focus groups, a Delphi survey was conducted in phase 3, involving neonatal providers across Australia and New Zealand. Phase 4 of the study involved an observational study that was used to test the six-item PTSR. The age at which specific feeding milestones were reached was consistent with what has been previously described in the literature. The chart audit showed that the time taken to the first feeding attempt in the preterm infant population was affected by gestational age at birth, birth weight, and specific interventions. Staff also considered age along with other criteria when deciding when to initiate feeding. Consensus on nine criteria for inclusion into the six-item PTSR was achieved using the Delphi technique. Three items of PTSR showed significant differences between the preterm and fullterm infant groups. Only two items, feeding-readiness behaviour and low pulse oximetry during handling, explained the variance in breastfeeding behaviour. The inter-rater variability ranged between moderate and very good for the PTSR items. The results of this study indicate the importance of assessing behavioural cues as an indication of breastfeeding readiness in the preterm infant population, once an infant is deemed physiologically stable. Age continues to be a factor in some clinicians' decisions to commence breastfeeding. However, age alone cannot be used to decide if an infant is ready to engage in breastfeeding. Further research is needed to confirm these findings.

Using strategic movement to calibrate a neural compass : a spiking network for tracking head direction in rats and robots

The head direction (HD) system in mammals contains neurons that fire to represent the direction the animal is facing in its environment. The ability of these cells to reliably track head direction even after the removal of external sensory cues implies that the HD system is calibrated to function effectively using just internal (proprioceptive and vestibular) inputs. Rat pups and other infant mammals display stereotypical warm-up movements prior to locomotion in novel environments, and similar warm-up movements are seen in adult mammals with certain brain lesion-induced motor impairments. In this study we propose that synaptic learning mechanisms, in conjunction with appropriate movement strategies based on warm-up movements, can calibrate the HD system so that it functions effectively even in darkness. To examine the link between physical embodiment and neural control, and to determine that the system is robust to real-world phenomena, we implemented the synaptic mechanisms in a spiking neural network and tested it on a mobile robot platform. Results show that the combination of the synaptic learning mechanisms and warm-up movements are able to reliably calibrate the HD system so that it accurately tracks real-world head direction, and that calibration breaks down in systematic ways if certain movements are omitted. This work confirms that targeted, embodied behaviour can be used to calibrate neural systems, demonstrates that ‘grounding’ of modeled biological processes in the real world can reveal underlying functional principles (supporting the importance of robotics to biology), and proposes a functional role for stereotypical behaviours seen in infant mammals and those animals with certain motor deficits. We conjecture that these calibration principles may extend to the calibration of other neural systems involved in motion tracking and the representation of space, such as grid cells in entorhinal cortex.

Lingodroids : studies in spatial cognition and language

The Lingodroids are a pair of mobile robots that evolve a language for places and relationships between places (based on distance and direction). Each robot in these studies has its own understanding of the layout of the world, based on its unique experiences and exploration of the environment. Despite having different internal representations of the world, the robots are able to develop a common lexicon for places, and then use simple sentences to explain and understand relationships between places even places that they could not physically experience, such as areas behind closed doors. By learning the language, the robots are able to develop representations for places that are inaccessible to them, and later, when the doors are opened, use those representations to perform goal-directed behavior.

Associations between young children's emotion attributions and prediction of outcome in differing social situations

Associations between young children's attributions of emotion at different points in a story, and with regard to their own prediction about the story's outcome, were investigated using two hypothetical scenarios of social and emotional challenge (social entry and negative event). First grade children (N = 250) showed an understanding that emotions are tied to situational cues by varying the emotions they attributed both between and within scenarios. Furthermore, emotions attributed to the main protagonist at the beginning of the scenarios were differentially associated with children's prediction of a positive or negative outcome and with the valence of the emotion attributed at the end of the scenario. Gender differences in responses to some items were also found. © 2010 The British Psychological Society.

Parenting behaviours and maternal infant-feeding practices in first-time Australian mothers

Introduction: There is emerging evidence that parenting style and early feeding practices are associated with child intake, eating behaviours and weight status. The aim of this cross sectional study was to examine the relationships between general maternal parenting behaviour and feeding practices and beliefs. Methods: Participants were 421 first-time mothers of 9-22 week old healthy term infants (49% male, mean±sd age 19±4 weeks) enrolled in the NOURISH trial. At baseline mothers self-reported their parenting behaviours (self-efficacy, warmth, irritability) and infant-feeding beliefs using questions from the Longitudinal Study of Australian Children and the Infant Feeding Questionnaire (Baughcum, 2001), respectively. Multivariable regression analyses were used with feeding practices (four factors) as the dependent variables, Independent variables were maternal BMI, weight concern, age, education level perception of infant weight status, feeding mode (breast vs formula) and infant gender, age and weight gain z-score. Results: Parenting behaviours partly were associated with feeding beliefs (adjusted R2 =0.21-0.30). Higher maternal parenting self-efficacy was inversely associated with concerns that the baby would become underweight (p=0.006); become overweight (p<0.001); and lack of awareness of infant hunger/satiety cues (p<0.001). Higher maternal irritability was positively associated with lack of awareness of cues (p<0.05). Maternal warmth was not associated with any feeding beliefs. Infant weight- gain (from birth) z-score and age, maternal BMI and education level and mothers’ perception of infant weight status and feeding mode were covariates. Conclusions: These findings suggest strategies to improve early feeding practices need to be address broader parenting approaches, particularly self-efficacy and irritability.

Association between feeding style and weight gain in infants aged 2–7 months

Introduction: Feeding on demand supports an infant’s innate capacity to respond to hunger and satiety cues and may promote later self-regulation of intake. Our aim was to examine whether feeding style (on demand vs to schedule) is associated with weight gain in early life. Methods: Participants were first-time mothers of healthy term infants enrolled NOURISH, an RCT evaluating an intervention to promote positive early feeding practices. Baseline assessment occurred when infants were aged 2-7 months. Infants able to be categorised clearly as feeding on demand or to schedule (mothers self report) were included in the logistic regression analysis. The model was adjusted for gender, breastfeeding and maternal age, education, BMI. Weight gain was defined as a positive difference in baseline minus birthweight z-scores (WHO standards) which indicated tracking above weight percentile. Results: Data from 356 infants with a mean age of 4.4 (SD 1.0) months were available. Of these, 197 (55%) were fed on demand, 42 (12%) were fed on schedule. There was no statistical association between feeding style and weight gain [OR=0.72 (95%CI 0.35-1.46), P=0.36]. Formula fed infants were three times more likely to be fed on schedule and formula feeding was independently associated with increased weight gain [OR=2.02 (95%CI 1.11-3.66), P=0.021]. Conclusion: In this preliminary analysis the association between feeding style and weight gain did not reach statistical significance, however , the effect size may be clinically relevant and future analysis will include the full study sample (N=698).

Associations between parenting behaviours and feeding beliefs of first-time Australian mothers in the NOURISH obesity prevention trial

Purpose: Parenting style and early feeding practices have been linked to child intake, eating behaviour and weight status. The purpose of this study was to examine associations between general maternal parenting behaviours and feeding beliefs in Australian mothers of 11-17 month-old children. Methods: This cross-sectional analysis included 223 first-time mothers and their children (49% male, mean age 14 [sd 1] months) enrolled in the control group of the NOURISH trial. Mothers self-reported their feeding beliefs and parenting behaviours (overprotection, irritability, warmth, autonomy-encouraging) using modified questions from the Infant Feeding Questionnaire (Baughcum, 2001) and the Longitudinal Study of Australian Children. Multiple regression analyses were conducted, using feeding beliefs (four factors) as dependent and parenting behaviours as independent variables while adjusting for child gender, age, weight-for-age z-score and maternal age, education level, feeding mode (breast vs. non-breast), and perception of own pre-pregnancy and child weight status. Results/Findings: Two of four parenting behaviours were significantly associated with feeding beliefs. Maternal warmth was inversely associated with concerns that the child would become underweight (β=-0.156, p=0.022) and positively associated with mothers’ awareness of child’s hunger/satiety cues (β=0.303, p<0.001). Mothers’ overprotection was positively associated with concerns that the child would become underweight (β=0.213, p=0.001); become overweight (β=0.174, p=0.005); and mother’s responsive feeding (β=0.135, p=0.057). Weight-for-age z-score, mothers’ perception of their child and own weight status, education, feeding mode, and child’s age were significant covariates. Conclusions: Feeding occurs within the broader parenting context. Improving early feeding beliefs and practices may require addressing mother’s approaches to parenting, especially warmth and overprotection.

An exploration of Online Shopping Cart Abandonment Syndrome – a matter of risk and reputation

ABSRACT. Despite the surge in online retail sales in recent years there still remains reluctance by consumers to complete the online shopping process. A number of authors have attributed consumers’ reluctance to purchase online to apparent barriers. However, such barriers as yet have not been fully examined within a theoretical context. This research explores the application of the perceived risk theoretical framework. Specifically, performance risk and the influence of perceived performance risk has on the phenomenon of Internet Abandoned Cart Syndrome (ACS) is evaluated. To explore this phenomenon, a number of extrinsic cues are identified as playing a major role in the performance evaluation process of online purchases. The results of this study suggest the extrinsic cues of brand, reputation, design and price have an overall impact on the performance evaluation process just prior to an online purchase. Varying these cues either positively or negatively had a strong impact on performance evaluation. Further, it was found that positive or negative reputation was heavily associated with shopping cart abandonment.

Overlapping medical alarms are almost indiscriminable

Objective: We explore how accurately and quickly nurses can identify melodic medical equipment alarms when no mnemonics are used, when alarms may overlap, and when concurrent tasks are performed. Background: The international standard IEC 60601-1-8 (International Electrotechnical Commission, 2005) has proposed simple melodies to distinguish seven alarm sources. Previous studies with nonmedical participants reveal poor learning of melodic alarms and persistent confusions between some of them. The effects of domain expertise, concurrent tasks, and alarm overlaps are unknown. Method: Fourteen intensive care and general medical unit nurses learned the melodic alarms without mnemonics in two sessions on separate days. In the second half of Day 2 the nurses identified single alarms or pairs of alarms played in sequential, partially overlapping, or nearly completely overlapping configurations. For half the experimental blocks nurses performed a concurrent mental arithmetic task. Results: Nurses' learning was poor and was no better than the learning of nonnurses in a previous study. Nurses showed the previously noted confusions between alarms. Overlapping alarms were exceptionally difficult to identify. The concurrent task affected response time but not accuracy. Conclusion: Because of a failure of auditory stream segregation, the melodic alarms cannot be discriminated when they overlap. Directives to sequence the sounding of alarms in medical electrical equipment must be strictly adhered to, or the alarms must redesigned to support better auditory streaming. Application: Actual or potential uses of this research include the implementation of IEC 60601-1-8 alarms in medical electrical equipment.

Useful field of view predicts driving in the presence of distracters

Purpose. The Useful Field of View (UFOV(R)) test has been shown to be highly effective in predicting crash risk among older adults. An important question which we examined in this study is whether this association is due to the ability of the UFOV to predict difficulties in attention-demanding driving situations that involve either visual or auditory distracters. Methods. Participants included 92 community-living adults (mean age 73.6 +/- 5.4 years; range 65-88 years) who completed all three subtests of the UFOV involving assessment of visual processing speed (subtest 1), divided attention (subtest 2), and selective attention (subtest 3); driving safety risk was also classified using the UFOV scoring system. Driving performance was assessed separately on a closed-road circuit while driving under three conditions: no distracters, visual distracters, and auditory distracters. Driving outcome measures included road sign recognition, hazard detection, gap perception, time to complete the course, and performance on the distracter tasks. Results. Those rated as safe on the UFOV (safety rating categories 1 and 2), as well as those responding faster than the recommended cut-off on the selective attention subtest (350 msec), performed significantly better in terms of overall driving performance and also experienced less interference from distracters. Of the three UFOV subtests, the selective attention subtest best predicted overall driving performance in the presence of distracters. Conclusions. Older adults who were rated as higher risk on the UFOV, particularly on the selective attention subtest, demonstrated poorest driving performance in the presence of distracters. This finding suggests that the selective attention subtest of the UFOV may be differentially more effective in predicting driving difficulties in situations of divided attention which are commonly associated with crashes.