On the challenges and opportunities facing fish biology: a discussion of five key knowledge gaps.

Many fish species face increasing challenges associated with climate change and overfishing. At the same time, aquaculture is becoming vital for food security. Gaining a deeper understanding of the basic biology of fish is therefore more important than ever. Here we synthesize and summarize key questions, opportunities and challenges in fish biology highlighted during a round-table discussion at the 50th Anniversary Symposium of The Fisheries Society of the British Isles, held at the University of Exeter, U.K., in July 2017. We identified several knowledge gaps but also key opportunities for fish biology to inform food security, for collective behaviour, evolutionary history and trait correlations to predict responses to environmental change and for novel analytical approaches to mine existing data sets. Overall, more integrative approaches through stronger collaborations across different fields are needed to advance our understanding of the basic biology of fish.

Quantitative dynamic imaging of immune cell signalling using lentiviral gene transfer.

Live-cell imaging of fluorescent fusion proteins has transformed our understanding of mammalian cell signalling and function. However, some cellular systems such as immune cells are unsuitable or refractory to many existing transgene delivery methods thus limiting systematic analyses. Here, a flexible lentiviral gene transfer platform for dynamic time-lapse imaging has been developed and validated with single-molecule spectroscopy, mathematical modelling and transcriptomics and used for analysis of a set of inflammation-related signalling networks. Time-lapse imaging of nuclear factor kappa B (NF-κB), signal transducer and activator of transcription (STATs) and nuclear factor of activated T-cells (NFAT) in mammalian immune cell lines provided evidence for heterogeneous temporal encoding of inflammatory signals. In particular, the absolute quantification of single-cell responses over time via fluorescent correlation spectroscopy (FCS) showed that NF-κB p65 activation in response to tumour necrosis factor α (TNFα) was differentially encoded in variable amplitude of nuclear translocation between immune and non-immune cells. The absolute number of activated molecules was dictated in part by the cell size, suggesting a morphology-dependent regulatory mechanism. The developed platform will enable further absolute quantitative analyses of the dynamic interactions between signalling networks, in and between individual cells, allowing better integration with mathematical models of signalling networks.

Cannabidivarin is anticonvulsant in mouse and rat.

BACKGROUND AND PURPOSE: Phytocannabinoids in Cannabis sativa have diverse pharmacological targets extending beyond cannabinoid receptors and several exert notable anticonvulsant effects. For the first time, we investigated the anticonvulsant profile of the phytocannabinoid cannabidivarin (CBDV) in vitro and in in vivo seizure models. EXPERIMENTAL APPROACH: The effect of CBDV (1-100 µM) on epileptiform local field potentials (LFPs) induced in rat hippocampal brain slices by 4-aminopyridine (4-AP) application or Mg(2+) -free conditions was assessed by in vitro multi-electrode array recordings. Additionally, the anticonvulsant profile of CBDV (50-200 mg·kg(-1) ) in vivo was investigated in four rodent seizure models: maximal electroshock (mES) and audiogenic seizures in mice, and pentylenetetrazole (PTZ) and pilocarpine-induced seizures in rats. The effects of CBDV in combination with commonly used antiepileptic drugs on rat seizures were investigated. Finally, the motor side effect profile of CBDV was investigated using static beam and grip strength assays. KEY RESULTS: CBDV significantly attenuated status epilepticus-like epileptiform LFPs induced by 4-AP and Mg(2+) -free conditions. CBDV had significant anticonvulsant effects on the mES (≥100 mg·kg(-1) ), audiogenic (≥50 mg·kg(-1) ) and PTZ-induced seizures (≥100 mg·kg(-1) ). CBDV (200 mg·kg(-1) ) alone had no effect against pilocarpine-induced seizures, but significantly attenuated these seizures when administered with valproate or phenobarbital at this dose. CBDV had no effect on motor function. CONCLUSIONS AND IMPLICATIONS: These results indicate that CBDV is an effective anticonvulsant in a broad range of seizure models. Also it did not significantly affect normal motor function and, therefore, merits further investigation as a novel anti-epileptic in chronic epilepsy models. LINKED ARTICLES: This article is part of a themed section on Cannabinoids. To view the other articles in this section visit http://dx.doi.org/10.1111/bph.2012.167.issue-8.

Metachronous testicular teratoma, testicular seminoma and papillary thyroid carcinoma occurring in a single individual: a report of two unrelated cases.

We describe two unrelated men who both developed teratomas in one testis followed by seminomas in the contralateral testis followed by papillary thyroid carcinomas. Neither man had a family history of cancers. Although random occurrence is possible, genetic predisposition and/or environmental influence would seem a likely explanation for this previously unreported combination of tumours.

Rapid onset intratubular calcification following renal transplantation requiring urgent parathyroidectomy.

BACKGROUND: Secondary hyperparathyroidism is a common complication of end-stage renal disease often requiring parathyroidectomy. Renal transplant with the restoration of normal renal function often allows resolution of hyperparathyroidism, avoiding the need for parathyroid surgery. However, a proportion of patients with hyperparathyroidism become overtly hypercalcemic after renal transplantation which poses management dilemmas between medical and surgical treatment. CASE: We present the case of a 48-yearold man with end-stage renal failure known to have secondary hyperparathyroidism who received a living related renal transplant. Postoperatively he developed prompt hypercalcemia, polyuria, polydipsia and rapid onset intratubular calcification, leading to acute tubular necrosis diagnosed on renal biopsy on Day 7 post transplantation. He underwent surgical parathyroidectomy with resolution of his hypercalcemia and improved renal transplant function. DISCUSSION: This case emphasizes the need for good management of secondary hyperparathyroidism together with close surveillance of PTH in patients awaiting renal transplantation. With good renal transplant function hyperparathyroidism usually resolves. Posttransplant surgical parathyroidectomy should be reserved for severe progressive end organ damage.

Phosphodiesterase (PDE) 7 in inflammatory cells from patients with asthma and COPD.

In inflammatory cells, the low K(m) cyclic adenosine monophosphate (cAMP)-specific phosphodiesterase (PDE) 4 subtype is predominant in terms of expression and function, although more recently it has been suggested that PDE 7 may also play a role in regulating inflammatory cell activity. In the present study, PDE 4 and PDE 7 subtype messenger ribonucleic acid (mRNA) transcripts in CD4 and CD8 lymphocytes from healthy (n=10) and asthmatic (n=10) subjects and polymorphonuclear neutrophils (PMNs) and CD8 lymphocytes obtained from healthy (n=10) and chronic obstructive pulmonary disease (COPD) (n=7) subjects were identified and quantified. PDE 4A, PDE 4B, PDE 4D and PDE 7A mRNA were present in similar quantities in both CD4 and CD8 lymphocytes obtained from healthy and asthmatic subjects and in CD8 lymphocytes obtained from healthy and COPD subjects. Expression of PDE 4C and PDE 7B mRNA was also observed, although transcript levels were low and variable between individuals. In addition, the effects of selective PDE 7 inhibition on both phytohaemagluttinin (PHA)-induced human peripheral blood mixed mononuclear cell (HPBMNC) proliferation and fMLP-induced neutrophil elastase (NE) release were studied. HPBMNC and human neutrophils, isolated from the venous blood of healthy volunteers (n=6) were treated with either a novel selective PDE 7 inhibitor PF 0332040 alone or in combination with rolipram. Proliferation of HPBMNC was stimulated by PHA (2microgml(-1)) and assessed by [(3)H]-thymidine incorporation, while fMLP-induced (100nM) NE release was determined using a chromogenic substrate. Both rolipram (0.003-10microM) and PF 0332040 (0.003-10microM) significantly inhibited PHA-stimulated proliferation of HPBMNC ((**)P<0.01). Co-administration of rolipram (0.3-10microM) and PF 0332040 (0.003-10microM) significantly increased the degree of inhibition observed, compared to when either drug was administered alone ((**)P<0.01). PF 0332040 (0.003-10microM) had no inhibitory effect on NE release from human peripheral blood neutrophils stimulated with fMLP (100nM), while rolipram (0.003-10microM) significantly inhibited neutrophil degranulation ((**)P<0.01). These findings suggest no evidence of altered PDE 4 or PDE 7 mRNA transcript levels in inflammatory cells isolated from the peripheral venous blood of mild asymptomatic asthmatic subjects or stable COPD subjects, however, inhibition of PDE 7 may influence mononuclear cell function.

Iatrogenic femoropopliteal graft entrapment leading to thrombus formation.

Femoropopliteal bypass graft entrapment by the gastrocnemius muscle and tendons is an unusual cause of graft stenosis or thrombosis. Before graft occlusion occurs, reduced flow may be seen either with the knee in extension or hyperextension or by passive dorsiflexion of the ankle. We report a case of a femoropopliteal bypass graft entrapment causing a thrombus in the distal graft. Duplex imaging, angiography, MRI and graft surveillance programs are useful diagnostic tools. Treatment options include dividing the occluding muscles and tendons and rerouting the graft.

The effect of selective phosphodiesterase isoenzyme inhibition on neutrophil function in vitro.

Neutrophil-derived proteases such as neutrophil elastase (NE) and matrix metalloproteinase (MMP) are implicated in the pathogenesis of Chronic Obstructive Pulmonary Disease (COPD). In this study, the effects of selective phosphodiesterase (PDE) inhibition on NE and MMP-9 release, as well as Myeloperoxidase (MPO) activity and integrin-mediated neutrophil adhesion to human umbilical vein endothelial cells (HUVECs), were investigated. Human neutrophils were treated with PDE inhibitors (10(-11)-10(-4)M) in the absence and presence of TNF-alpha (tumour necrosis factor) (100 U ml(-1)) for 30 min, prior to fMLP activation. After 45 min, the cells were removed and NE, MPO and MMP-9 release assessed. In the adhesion studies, the neutrophils were radio-labelled with 51Cr, stimulated and immediately transferred to cultured HUVEC monolayers for 30 min, prior to assessment of adhesion. TNF-alpha (100 U ml(-1)) acted synergistically with fMLP in stimulating azurophil degranulation with respect to both MPO activity (P<0.01) and NE release (P<0.01). In contrast, an additive effect was observed with TNF-alpha and fMLP with regard to MMP-9 release and neutrophil adhesion to HUVECs. The PDE4 inhibitors, roflumilast, roflumilast N-oxide, cilomilast and rolipram significantly suppressed MPO, NE and MMP-9 release in both the presence and absence of TNF-alpha (P<0.05; n=6-10) and also reduced neutrophil adhesion to HUVECs. In contrast, milrinone, a PDE3 inhibitor and the non-selective PDE inhibitor, theophylline did not inhibit azurophil degranulation under any of the experimental conditions. These data provide further evidence that selective PDE4 isoenzyme inhibitors can inhibit neutrophil degranulation, effects not shared by PDE3 inhibitors or theophylline.

Outcome following surgery for thoracic outlet syndrome.

INTRODUCTION: the diagnosis of thoracic outlet syndrome (TOS) relies heavily on subjective rather than objective assessment criteria. Subsequently, published results after surgical decompression vary considerably. This study aimed to use a symptom-based patient-directed questionnaire to assess the outcome after decompression for TOS. METHODS: sixty patients who underwent decompression procedures were identified from a prospectively maintained vascular database. Patient records were analysed for details regarding initial presentation, investigation, type of procedure used for decompression and management. Outcome questionnaires were sent to all identified patients to give a patient-based outcome measure. RESULTS: eighty-four per cent of patients responded. In 90% of these patients there was an improvement in symptoms post-surgery with a median follow up of 43 months. The results were not influenced by the procedure or approach used. CONCLUSION: surgery remains an effective tool in the management of TOS. A simple patient-directed questionnaire as used in this study could assist in the standardisation of outcome assessment.

A novel testosterone gel formulation normalizes androgen levels in hypogonadal men, with improvements in body composition and sexual function.

OBJECTIVE: To compare the safety and efficacy of two doses of a new testosterone gel formulation (Testim Auxilium Pharmaceuticals, Inc., Norristown, PA, USA) to a permeation-enhanced testosterone patch (Andropatch), GlaxoSmithKline, UK) for treating men with confirmed low serum testosterone levels, and associated signs and symptoms of hypogonadism. PATIENTS AND METHODS: In all, 208 men were randomized and treated at 29 centres in Denmark, Germany, Netherlands, Sweden and the UK. The men were treated for 90 days, and the pharmacokinetics and treatment effectiveness of Testim at two doses (50 and 100 mg/day, delivering a daily dose of 5 and 10 mg testosterone, respectively) and Andropatch (2 x 2.5 mg patches, each delivering 2.5 mg testosterone and containing 12.2 mg of testosterone) were compared. Pharmacokinetic profiles were obtained, body composition measured, and mood and sexual function data recorded. RESULTS: Testim produced dose-dependent improvements in all pharmacokinetic variables compared with Andropatch. The mean increases from baseline to 90 days in testosterone were 12.41, 6.54 and 3.82 nmol/L for Testim 100 and 50 mg/day and the Andropatch, respectively. Both doses of Testim significantly improved positive and negative mood over baseline; Andropatch did not. All three treatments increased lean body mass, and the higher dose of Testim produced a significant decrease in percentage body fat. At all sample times both doses of Testim significantly improved sexual performance, sexual motivation, sexual desire and spontaneous erections. Andropatch provided insignificant improvements from baseline at all sample times for sexual desire, an inconsistent improvement in sexual motivation, but no effect on spontaneous erections. These results are similar to those previously reported for testosterone replacement therapy in hypogonadal men, suggesting that normalization of serum testosterone restores sexual function. However, the present data suggest that higher serum testosterone levels may further improve sexual function. Gel treatment was well tolerated, while patch treatment produced higher rates of application-site reactions and study discontinuation. CONCLUSION: The favourable pharmacokinetic profile and treatment outcome, combined with the enhanced tolerability of Testim, suggest that this new gel formulation is a safe and effective treatment in men with low serum testosterone levels and associated signs and symptoms of hypogonadism.

The value of risks scores for predicting postoperative nausea and vomiting when used to compare patient group in a randomised controlled trial.

Whilst conducting a randomised controlled trial into the effects of combination anti-emetics, we endeavoured to confirm that our patient groups were matched using the predictive scoring systems for postoperative nausea and vomiting (PONV) and postoperative vomiting (POV) reported in the literature. One hundred and seventy-seven female patients attending for day case gynaecological surgery were studied and their individual risks of PONV and POV were calculated using four predictive models for PONV and two predictive models for POV. The scoring systems were then evaluated to see if agreement existed between them using the method described by Bland and Altman. Bias and 95% limits of agreement were calculated for each combination. Agreement between scoring systems was poor. As the scoring systems gave widely divergent predictions, we concluded that the predictive risk for PONV or POV would be dependent upon the scoring system chosen, thus limiting their usefulness in this role.

The use of anaesthetic rooms for induction of anaesthesia: a postal survey of current practice and attitudes in Great Britain and Northern Ireland.

A postal survey was sent to all anaesthetic departments in the UK to identify current practice and gain insight into anaesthetists' attitudes regarding the use of anaesthetic rooms for induction of general anaesthesia. Replies were received from 247 (88%) departments. Of these, 10 (4%) departments routinely anaesthetise all patients in theatre. The main reason for change was patient safety. Of those who routinely use the anaesthetic room for induction of anaesthesia, only 5% have made provision to change to in-theatre induction. An estimated pound 30 million has been spent on equipping anaesthetic rooms since 1994; with the result that 91% of departments where anaesthetic room induction occurs, now have monitoring that complies with the current Association of Anaesthetists of Great Britain and Ireland guidelines. The majority of the respondents who use anaesthetic rooms perceived induction in theatre to result in reduced efficiency, increased patient anxiety, a worse teaching environment and no improvement in patient safety. This was in contrast to the attitudes of respondents from hospitals where in-theatre induction occurs. Only 9.7% of all respondents believed that clinical governance would necessitate a change to anaesthetizing all patients in theatre compared to 25% who believed that the increasing costs of monitoring equipment would lead to a change. Overall 79% of respondents prefer to use the anaesthetic room, 16% prefer in-theatre induction and 5% expressed no preference. However, of those who routinely anaesthetise in theatre, 70% thought it to be preferable.

A comparison between midazolam co-induction and propofol predosing for the induction of anaesthesia in the elderly.

In a prospective, double-blind, randomised, placebo-controlled trial, we have compared the effects of midazolam co-induction with propofol predosing on the induction dose requirements of propofol in elderly patients. We enrolled 60 patients aged > 70 years, attending for urological surgery. The patients were allocated randomly to one of three groups, to receive either midazolam 0.02 mg.kg(-1), propofol 0.25 mg.kg(-1), or normal saline 2 ml (placebo) 2 min prior to induction of anaesthesia using propofol 1% infusion at 300 ml.h(-1). The propofol dose requirements for induction were recorded for two end-points (loss of verbal contact and insertion of an oropharyngeal airway). Cardiovascular parameters were recorded at 1-min intervals for each patient until induction was complete. The midazolam group showed a significant reduction in propofol dose requirements for induction (p = 0.05) compared to the placebo group. The propofol group did not show a significant dose reduction compared to placebo. There were no demonstrable differences in terms of improved cardiovascular stability between groups. We conclude that propofol predosing does not significantly reduce the induction dose of propofol required in the elderly, and there were no cardiovascular benefits to either midazolam co-induction or propofol predosing in the elderly.

Management of life-threatening autonomic hyper-reflexia using magnesium sulphate in a patient with a high spinal cord injury in the intensive care unit.

We report the successful use of i.v. magnesium sulphate to control life-threatening autonomic hyper-reflexia associated with chronic spinal cord injury in the intensive care environment. A 37-yr-old, male was admitted to the intensive care unit with a diagnosis of septic shock and acute renal failure secondary to pyelonephritis. He had been found unresponsive at home following a 2-day history of pyrexia and purulent discharge from his suprapubic catheter. He had sustained a T5 spinal cord transection 20 yr previously. Initial management included assisted ventilation, fluid resuscitation, vasopressor support, and continuous veno-venous haemofiltration. The sepsis was treated with antibiotic therapy and percutaneous nephrostomy drainage of the pyonephrosis. On the fifth day, the patient developed profuse diarrhoea. This was associated with paroxysms of systemic hypertension and diaphoresis, his arterial pressure rising on occasion to 240/140 mm Hg. A diagnosis of autonomic hyper-reflexia was made and a bolus dose of magnesium sulphate 5 g was administered over 15 min followed by an infusion of 1-2 g h(-1). There was an almost immediate decrease in the severity and frequency of the hypertensive episodes. There were no adverse cardiac effects associated with the administration of magnesium, only a slight decrease in minute ventilation as the plasma level approached the upper end of the therapeutic range (2-4 mmol litre(-1)). In view of the beneficial effects observed in this case we advocate further research into the use of magnesium sulphate in the treatment or prevention of autonomic hyper-reflexia secondary to chronic spinal cord injury in the intensive care unit.

Identification and quantification of phosphodiesterase 4 subtypes in CD4 and CD8 lymphocytes from healthy and asthmatic subjects.

In the present study, for the first time, PDE4 subtypes were identified and semi-quantified in both CD4 and CD8 lymphocytes from healthy and asthmatic individuals. CD4 and CD8 lymphocytes from healthy and mild asymptomatic asthmatic subjects (receiving beta-agonist therapy only) were isolated from peripheral venous blood using appropriate antibody coated paramagnetic beads. PDE4 subtypes and beta-actin were identified by digoxigenin (DIG)-labelling reverse transcriptase-polymerase chain reaction and semi-quantified by DIG-detection enzyme-linked immunosorbance assay. In CD4 and CD8 lymphocytes PDE4A, PDE4B and PDE4D were detected, with no significant differences observed between healthy and asthmatic groups. In CD8 lymphocytes, enzyme subtype expression was lower and showed more intersubject variability. In functional studies investigating the effects of various PDE inhibitors on PHA-induced proliferation of mononuclear cells from healthy and asthmatic subjects, CDP840 (0.03 - 10 microM), rolipram (0.1 - 10 microM) and theophylline (10 microM - 1 mM) inhibited PHA-induced proliferation of mononuclear cells from healthy and asthmatic subjects in a concentration-dependent manner, although no significant difference was observed between the groups investigated. In additional studies, total monocyte cyclic AMP PDE activity was investigated in cells isolated from asthmatic subjects both prior to and 24 h after allergen challenge. Total monocyte cyclic AMP PDE activity remained unaffected following challenge of asthmatic subjects with either house dust mite or cat dander and was inhibited in a concentration-dependent manner by rolipram (0.01 - 100 microM) both before and after allergen challenge.

Maternal depression and increased fetal activity.

Pregnant women with (n = 45) and without (n = 45) symptoms of depression (CES-D score of 16 or more) were provided ultrasound examinations during the second and third trimesters. An analysis of variance on the cross-sectional data yielded a significant diagnosis by gestational month interaction effect (P < 0.05). The fetuses of depressed mothers spent significantly more time being active during the fifth, sixth and seventh gestational months than fetuses of non-depressed mothers. A stepwise regression analysis revealed that 35% of the variance in time being active could be accounted for by the combined depression and trait anxiety scores of the mothers. These findings suggest that maternal depression correlated with increased fetal activity. These data also highlight the need for research on the potential effects of stress hormones on fetal activity.

Polymer chemical structure is a key determinant of physicochemical and colloidal properties of polymer-DNA complexes for gene delivery.

Polyplexes are now emerging as potentially useful vectors for gene therapy. To improve our understanding of how the chemical structure of the polymer affects the properties of these systems, a series of structurally related polymers, the linear poly(amidoamine)s (PAAs), have been examined for their abilities to form complexes with DNA. Structure-dependent differences in DNA binding are shown by gel electrophoretic retardation of DNA and thermal transition analyses. Two PAAs, NG28 and NG30, stand out as having high affinity DNA binding characteristics, similar to the model homopolypeptide, poly-L-lysine. In addition, differences in complex formation, particle size and surface charge are displayed for the different polymer-DNA systems. Electron microscopy studies showed that the polymers condensed DNA into similar unit structures but only complexes with NG30 did not undergo agglomeration. This was attributed to an excess of complexed polymer forming a shell of uncomplexed polymer chain segments around a condensed DNA-polymer core. The transfection activities of these polymer complexes differ greatly, and some of these differences can be explained in a multifactorial way by the physicochemical and colloidal properties. It is concluded that polymer chemical structure dictates the apparent affinity of DNA binding, and also several of the important colloidal characteristics of the resulting complexes.

Right frontal EEG asymmetry and lack of empathy in preschool children of depressed mothers.

EEG activity, empathic reactions to emotion-inducing stimuli, and the ability to complete a teaching task were examined in preschool children of depressed and non-depressed mothers. EEG activity from frontal and parietal regions was recorded. Repeated measures MANOVAs indicated that the children of depressed mothers had greater relative right frontal EEG asymmetry, a pattern that typically accompanies greater negative affect and showed less empathic responses to a crying infant as well as to their own mothers' simulated distress. Children of depressed mothers were slower in completing the teaching task (involving mutual cooperation with their mother) and they spent more time asking for help than children of non-depressed mothers. Further, the depressed mothers stated their approval less often and spent less time helping their child complete the task.

The regulation of AMP-activated protein kinase by phosphorylation.

The AMP-activated protein kinase (AMPK) cascade is activated by an increase in the AMP/ATP ratio within the cell. AMPK is regulated allosterically by AMP and by reversible phosphorylation. Threonine-172 within the catalytic subunit (alpha) of AMPK (Thr(172)) was identified as the major site phosphorylated by the AMP-activated protein kinase kinase (AMPKK) in vitro. We have used site-directed mutagenesis to study the role of phosphorylation of Thr(172) on AMPK activity. Mutation of Thr(172) to an aspartic acid residue (T172D) in either alpha1 or alpha2 resulted in a kinase complex with approx. 50% the activity of the corresponding wild-type complex. The activity of wild-type AMPK decreased by greater than 90% following treatment with protein phosphatases, whereas the activity of the T172D mutant complex fell by only 10-15%. Mutation of Thr(172) to an alanine residue (T172A) almost completely abolished kinase activity. These results indicate that phosphorylation of Thr(172) accounts for most of the activation by AMPKK, but that other sites are involved. In support of this we have shown that AMPKK phosphorylates at least two other sites on the alpha subunit and one site on the beta subunit. Furthermore, we provide evidence that phosphorylation of Thr(172) may be involved in the sensitivity of the AMPK complex to AMP.