Notch controls the cell cycle to define leader versus follower identities during collective cell migration.

Coordination of cell proliferation and migration is fundamental for life, and its dysregulation has catastrophic consequences, such as cancer. How cell cycle progression affects migration, and vice versa, remains largely unknown. We address these questions by combining in silico modelling and in vivo experimentation in the zebrafish trunk neural crest (TNC). TNC migrate collectively, forming chains with a leader cell directing the movement of trailing followers. We show that the acquisition of migratory identity is autonomously controlled by Notch signalling in TNC. High Notch activity defines leaders, while low Notch determines followers. Moreover, cell cycle progression is required for TNC migration and is regulated by Notch. Cells with low Notch activity stay longer in G1 and become followers, while leaders with high Notch activity quickly undergo G1/S transition and remain in S-phase longer. In conclusion, TNC migratory identities are defined through the interaction of Notch signalling and cell cycle progression.

Variables associated with administration of analgesia, nurse-initiated analgesia and early analgesia in the emergency department.

OBJECTIVE: To determine the patient and clinical variables associated with administration of any analgesia, nurse-initiated analgesia (NIA, prescribed and administered by a nurse) and early analgesia (within 30 min of presentation). METHODS: We undertook a retrospective cohort study of patients who presented to a metropolitan ED in Melbourne, Australia, during July and August, 2013. The ED has an established NIA programme. Patients were included if they were aged 18 years or more and presented with a painful complaint. The study sample was randomly selected from a list of all eligible patients. Data were extracted electronically from the ED records and by explicit extraction from the medical record. Logistic regression models were constructed to assess associations with the three binary study end points. RESULTS: 1289 patients were enrolled. Patients were less likely to receive any analgesia if they presented 08:00-15:59 hours (OR 0.67, 95% CI 0.46 to 0.98) or 16:00-24:00 hours (OR 0.55, 95% CI 0.37 to 0.80) were triage category 5 (OR 0.20, 95% CI 0.08 to 0.49) or required an interpreter (OR 0.34, 95% CI 0.14 to 0.86). Patients were less likely to receive NIA or early analgesia if they were aged 56 years or more (OR 0.70 and 0.63; OR 0.57 and 0.21, respectively) or if they had received ambulance analgesia (OR 0.59, 95% CI 0.36 to 0.95; OR 0.38, 95% CI 0.20 to 0.74, respectively). CONCLUSIONS: Patients who present during the daytime, have a triage category of 5 or require an interpreter are less likely to receive analgesia. Older patients and those who received ambulance analgesia are less likely to receive NIA or early analgesia.

The TARGET pain study: Lessons from a painful marathon.

This perspective article summarises the experience of conducting a multicentre research project. We describe expected and unexpected hurdles we experienced as well as suggesting possible solutions for researchers embarking on multicentre studies.

Best-practice pain management in the emergency department: A cluster-randomised, controlled, intervention trial.

OBJECTIVES: We aimed to provide 'adequate analgesia' (which decreases the pain score by ≥2 and to <4 [0-10 scale]) and determine the effect on patient satisfaction. METHODS: We undertook a multicentre, cluster-randomised, controlled, intervention trial in nine EDs. Patients with moderate pain (pain score of ≥4) were eligible for inclusion. The intervention was a range of educational activities to encourage staff to provide 'adequate analgesia'. It was introduced into five early intervention EDs between the 0 and 6 months time points and at four late intervention EDs between 3 and 6 months. At 0, 3 and 6 months, data were collected on demographics, pain scores, analgesia provided and pain management satisfaction 48 h post-discharge (6 point scale). RESULTS: Overall, 1317 patients were enrolled. Logistic regression (controlling for site and other confounders) indicated that, between 0 and 3 months, satisfaction increased significantly at the early intervention EDs (OR 2.2, 95% CI 1.5 to 3.4 [P < 0.01]) but was stable at the control EDs (OR 0.8, 95% CI 0.5 to 1.3 [P = 0.35]). Pooling of data from all sites indicated that the proportion of patients very satisfied with their pain management increased from 42.9% immediately pre-intervention to 53.9% after 3 months of intervention (difference in proportions 11.0%, 95% CI 4.2 to 17.8 [P = 0.001]). Logistic regression of all data indicated that 'adequate analgesia' was significantly associated with patient satisfaction (OR 1.4, 95% CI 1.1 to 1.8 [P < 0.01]). CONCLUSIONS: The 'adequate analgesia' intervention significantly improved patient satisfaction. It provides a simple and efficient target in the pursuit of best-practice ED pain management.

Par3 controls neural crest migration by promoting microtubule catastrophe during contact inhibition of locomotion.

There is growing evidence that contact inhibition of locomotion (CIL) is essential for morphogenesis and its failure is thought to be responsible for cancer invasion; however, the molecular bases of this phenomenon are poorly understood. Here we investigate the role of the polarity protein Par3 in CIL during migration of the neural crest, a highly migratory mesenchymal cell type. In epithelial cells, Par3 is localised to the cell-cell adhesion complex and is important in the definition of apicobasal polarity, but the localisation and function of Par3 in mesenchymal cells are not well characterised. We show in Xenopus and zebrafish that Par3 is localised to the cell-cell contact in neural crest cells and is essential for CIL. We demonstrate that the dynamics of microtubules are different in different parts of the cell, with an increase in microtubule catastrophe at the collision site during CIL. Par3 loss-of-function affects neural crest migration by reducing microtubule catastrophe at the site of cell-cell contact and abrogating CIL. Furthermore, Par3 promotes microtubule catastrophe by inhibiting the Rac-GEF Trio, as double inhibition of Par3 and Trio restores microtubule catastrophe at the cell contact and rescues CIL and neural crest migration. Our results demonstrate a novel role of Par3 during neural crest migration, which is likely to be conserved in other processes that involve CIL such as cancer invasion or cell dispersion.

Factors associated with high levels of patient satisfaction with pain management.

OBJECTIVES: The objective was to determine, among emergency department (ED) patients, the factors associated with a high level of satisfaction with pain management. METHODS: This was a prospective cohort study in a single ED. Consecutive adult patients, with triage pain scores of ≥4 (numerical rating scale=0 to 10), were enrolled. Variables examined included demographics, presenting complaint, pain scores, nurse-initiated analgesia, analgesia administered, time to first analgesia, specific pain communication, and whether "adequate analgesia" was provided (defined as a decrease in pain score to <4 and a decrease from the triage pain score of ≥2). The level of patient satisfaction with their pain management (six-point scale: very unsatisfied to very satisfied) was determined by a blinded investigator 48 hours post discharge. Logistic regression analyses were undertaken. RESULTS: Data were complete for 476 patients: mean (±standard deviation [SD]) age was 43.6 (±17.2) years, and 237 were males (49.8%, 95% confidence interval [CI]=45.2% to 54.4%). A total of 190 (39.9%, 95% CI=35.5% to 44.5%) patients were "very satisfied" with their pain management, and 207 (43.5%, 95% CI=39.0% to 48.1%) patients received adequate analgesia. Three variables were associated with the patient being very satisfied: the provision of adequate analgesia (odds ratio [OR]=7.8, 95% CI=4.9 to 12.4), specific pain communication (OR=2.3, 95% CI=1.3 to 4.1), and oral opioid administration (OR=2.0, 95% CI=1.1 to 3.4). Notably, the provision of nurse-initiated analgesia to 211 patients (44.3%, 95% CI=39.8% to 48.9%) and the short time to analgesia (median=11.5 minutes; interquartile range [IQR]=2.0 to 85.8 minutes) were not associated with being very satisfied. CONCLUSIONS: The receipt of adequate analgesia (as defined) is highly associated with patient satisfaction. This variable may serve as a clinically relevant and achievable target in the pursuit of best-practice pain management.

Evolution of the Alx homeobox gene family: parallel retention and independent loss of the vertebrate Alx3 gene.

The Alx gene family is implicated in craniofacial development and comprises two to four homeobox genes in each vertebrate genome analyzed. Using phylogenetics and comparative genomics, we show that the common ancestor of jawed vertebrates had three Alx genes descendent from the two-round genome duplications (Alx1, Alx3, Alx4), compared with a single amphioxus gene. Later in evolution one of the paralogues, Alx3, was lost independently from at least three different vertebrate lineages, whereas Alx1 and Alx4 were consistently retained. Comparison of spatial gene expression patterns reveals that the three mouse genes have equivalent craniofacial expression to the two chick and frog genes, suggesting that redundancy compensated for gene loss. We suggest that multiple independent loss of one Alx gene was predisposed by extensive and persistent overlap in gene expression between Alx paralogues. Even so, it is unclear whether it was coincidence or evolutionary bias that resulted in the same Alx gene being lost on each occasion, rather than different members of the gene family.

Gli2a protein localization reveals a role for Iguana/DZIP1 in primary ciliogenesis and a dependence of Hedgehog signal transduction on primary cilia in the zebrafish.

BACKGROUND: In mammalian cells, the integrity of the primary cilium is critical for proper regulation of the Hedgehog (Hh) signal transduction pathway. Whether or not this dependence on the primary cilium is a universal feature of vertebrate Hedgehog signalling has remained contentious due, in part, to the apparent divergence of the intracellular transduction pathway between mammals and teleost fish. RESULTS: Here, using a functional Gli2-GFP fusion protein, we show that, as in mammals, the Gli2 transcription factor localizes to the primary cilia of cells in the zebrafish embryo and that this localization is modulated by the activity of the Hh pathway. Moreover, we show that the Igu/DZIP1protein, previously implicated in the modulation of Gli activity in zebrafish, also localizes to the primary cilium and is required for its proper formation. CONCLUSION: Our findings demonstrate a conserved role of the primary cilium in mediating Hedgehog signalling activity across the vertebrate phylum and validate the use of the zebrafish as a representative model for the in vivo analysis of vertebrate Hedgehog signalling.

Effect of topical alkane vapocoolant spray on pain with intravenous cannulation in patients in emergency departments: randomised double blind placebo controlled trial.

OBJECTIVE: To assess the efficacy, acceptability, and safety of a topical alkane vapocoolant in reducing pain during intravenous cannulation in adults. DESIGN: Randomised double blind placebo controlled trial. SETTING: Emergency department of a metropolitan teaching hospital. PARTICIPANTS: 201 adult patients (54% male), mean (SD) age 58.2 (19.5) years, who required intravenous cannulation. INTERVENTIONS: Less than 15 seconds before cannulation, the skin area was sprayed with either water (control, n=98) or vapocoolant (intervention, n=103), from a distance of 12 cm for 2 seconds. The intervention spray was a blend of propane, butane, and pentane. MAIN OUTCOME MEASURES: Pain with cannulation and discomfort with spray, measured with a 100 mm visual analogue scale. RESULTS: Groups did not differ significantly in age, sex, indication for or site of cannulation, cannula size, or who cannulated the patient (P>0.05). Median (interquartile range) pain scores for cannulation in the control and intervention groups were 36 (19-51) and 12 (5-40) mm, respectively (P<0.001), and 59 (60%) and 33 (32%) reported pain scores >or=30 mm (P<0.001). Scores for spray discomfort also differed significantly (P<0.001) because of skewing to the right within the intervention group. The median discomfort scores, however, were 0 mm in both groups. Success rates for first cannulation attempt did not differ between groups (P=0.39). Thirty four (39%) and 62 (62%) patients said they would choose the spray they received for analgesia in the future (P=0.002). At follow-up at five days, two patients in the intervention group reported transient skin redness. CONCLUSIONS: Topical alkane vapocoolant spray is effective, acceptable, and safe in reducing pain with peripheral intravenous cannulation in adults in the emergency department. TRIAL REGISTRATION: Australian Clinical Trials ACTRN12607000470493.

Conservation of upstream regulators of scute on the notum of cyclorraphous Diptera.

Bristles on the notum of many cyclorraphous flies are arranged into species-specific stereotyped patterns. Differences in the spatial expression of the proneural gene scute correlate with the positions of bristles in those species looked at so far. However, the examination of a number of genes encoding trans-regulatory factors, such as pannier, stripe, u-shaped, caupolican and wingless, indicates that they are expressed in conserved domains on the prospective notum. This suggests that the function of a trans-regulatory network of genes is relatively unchanged in derived Diptera, and that many differences are likely to be due to changes in cis-regulatory sequences of scute. In contrast, in Anopheles gambiae, a basal species with no stereotyped bristle pattern, the expression patterns of pannier and wingless are not conserved, and expression of AgASH, the Anopheles proneural gene, does not correlate in a similar manner with the bristle pattern. We discuss the possibility that independently acting cis-regulatory sequences at the scute locus may have arisen in the lineage giving rise to cyclorraphous flies.

A conserved trans-regulatory landscape for scute expression on the notum of cyclorraphous Diptera.

Bristles on the notum of many cyclorraphous flies are arranged into species-specific stereotyped patterns. The positions of bristles correlate with differences in the spatial expression of the scute (sc) gene in those species examined so far. However, a major upstream activator of scute, Pannier (Pnr), is expressed in a conserved domain over the entire medial notum. Here we examine the expression patterns in Calliphora vicina of stripe (sr), u-shaped (ush), caupolican (caup) and wingless (wg), genes known to modify the activity of Pnr or to act downstream of Pnr in Drosophila. We find that, with minor differences, their expression patterns are conserved. This suggests that the function of a trans-regulatory network of genes is relatively unchanged in derived Diptera and that many differences are likely to be due to changes in cis-regulatory sequences of scute.

Multidisciplinary assessment at triage: a new way forward.

OBJECTIVE: To evaluate a dual doctor and nurse triage system at a tertiary referral hospital. METHODS: Data were compared between periods of multidisciplinary triage and periods of standard triage. Data comparison was also made between rostered multidisciplinary triage shifts and non-multidisciplinary triage shifts. Staff satisfaction with the process was assessed. RESULTS: The percentage of patients seen within Australasian Triage Scale performance indicator thresholds increased from 75% to 81% in Category 2 patients (P = 0.12) and 56% to 78% in Category 3 patients (P < 0.0001). There was a reduction of 50% in the number of patients who left prior to being seen by a doctor (P = 0.024). Surveys showed high staff satisfaction with the process. CONCLUSIONS: We feel that multidisciplinary triage performs a useful function in our department enabling us to reduce waiting times. The process is widely accepted amongst the staff and it ensures a senior doctor assesses most patients. It reduces the number of patients leaving prior to being seen by a doctor and it provides one way of getting around access block and a physically small department.

Agonist-specific coupling of a cloned human alpha1A-adrenoceptor to different second messenger pathways.

The agonist-specific coupling properties of the three cloned human alpha(1)-adrenoceptor subtypes have been compared, when expressed at similar levels in Chinese hamster ovary (CHO) cell lines, using noradrenaline and the (+/-)- meta- and (+/-)- para- structural isomers of octopamine as agonists. The alpha(1A)- and the alpha(1B)-adrenoceptor subtypes coupled to both the release of arachidonic acid and to the accumulation of inositol phosphates, whereas the alpha(1D)-adrenoceptor subtype only coupled effectively to the accumulation of inositol phosphates. The rank order of potencies of the three agonists tested was the same for all the three receptor subtypes when coupled to either signalling pathway: noradrenaline > meta-octopamine > para-octopamine. Meta-octopamine was a partial agonist of the alpha(1A)-adrenoceptor subtype when coupled to arachidonic acid release, whereas para-octopamine was a full agonist of this pathway. In contrast, meta-octopamine was a full agonist at the alpha(1B)-adrenoceptor subtype when coupled to arachidonic acid release, whereas para-octopamine was a partial agonist of this pathway. Neither meta-octopamine, nor para-octopamine acted as full agonists when coupling any of the three alpha(1)-adrenoceptor subtypes to the accumulation of inositol phosphates. Para-octopamine was only a weak partial agonist of this pathway for all three receptor subtypes. The results show that the modulation of arachidonic acid release and inositol 1,4,5-trisphosphate production occurs in both a subtype- and agonist-specific manner for the alpha(1A)-, alpha(1B)- and alpha(1D)-adrenoceptor subtypes. In addition, the alpha(1A)-adrenoceptor exhibits agonist-specific coupling (agonist trafficking) to the different second messenger pathways.

olig2 is required for zebrafish primary motor neuron and oligodendrocyte development.

Oligodendrocytes are produced from the same region of the ventral spinal cord that earlier generated motor neurons in bird and rodent embryos. Motor neuron and oligodendrocyte precursor cells express Olig genes, which encode basic helix-loop-helix transcription factors that play important roles in the development of both motor neurons and oligodendrocytes. We found that oligodendrocytes develop similarly in zebrafish embryos, in that they arise from ventral spinal cord and migrate to new positions. Developing primary motor neurons and oligodendrocytes express olig2 as do neural plate cells that give rise to both primary motor neurons and oligodendrocytes. Loss of olig2 function prevented primary motor neuron and oligodendrocyte development, whereas olig2 overexpression promoted formation of excess primary motor neurons and oligodendrocytes. We provide genetic evidence that Hedgehog signaling is required for zebrafish olig2 expression and oligodendrocyte development. However, olig2 overexpression did not promote primary motor neuron or oligodendrocyte development in embryos with reduced Hedgehog signaling activity. One possibility consistent with these data is that Hedgehog signaling, partly by inducing olig2 expression, specifies neural precursor cells that have potential for primary motor neuron or oligodendrocyte fate.