Miniature wireless recording and stimulation system for rodent behavioural testing.

OBJECTIVE: Elucidation of neural activity underpinning rodent behaviour has traditionally been hampered by the use of tethered systems and human involvement. Furthermore the combination of deep-brain stimulation (DBS) and various neural recording modalities can lead to complex and time-consuming laboratory setups. For studies of this type, novel tools are required to drive forward this research. APPROACH: A miniature wireless system weighing 8.5 g (including battery) was developed for rodent use that combined multichannel DBS and local-field potential (LFP) recordings. Its performance was verified in a working memory task that involved 4-channel fronto-hippocampal LFP recording and bilateral constant-current fimbria-fornix DBS. The system was synchronised with video-tracking for extraction of LFP at discrete task phases, and DBS was activated intermittently at discrete phases of the task. MAIN RESULTS: In addition to having a fast set-up time, the system could reliably transmit continuous LFP at over 8 hours across 3-5 m distances. During the working memory task, LFP pertaining to discrete task phases was extracted and compared with well-known neural correlates of active exploratory behaviour in rodents. DBS could be wirelessly activated/deactivated at any part of the experiment during EEG recording and transmission, allowing for a seamless integration of this modality. SIGNIFICANCE: The wireless system combines a small size with a level of robustness and versatility that can greatly simplify rodent behavioural experiments involving EEG recording and DBS. Designed for versatility and simplicity, the small size and low-cost of the system and its receiver allow for enhanced portability, fast experimental setup times, and pave the way for integration with more complex behaviour.

Exploration of the two-photon excitation spectrum of fluorescent dyes at wavelengths below the range of the Ti:Sapphire laser.

We have studied the wavelength dependence of the two-photon excitation efficiency for a number of common UV excitable fluorescent dyes; the nuclear stains DAPI, Hoechst and SYTOX Green, chitin- and cellulose-staining dye Calcofluor White and Alexa Fluor 350, in the visible and near-infrared wavelength range (540-800 nm). For several of the dyes, we observe a substantial increase in the fluorescence emission intensity for shorter excitation wavelengths than the 680 nm which is the shortest wavelength usually available for two-photon microscopy. We also find that although the rate of photo-bleaching increases at shorter wavelengths, it is still possible to acquire many images with higher fluorescence intensity. This is particularly useful for applications where the aim is to image the structure, rather than monitoring changes in emission intensity over extended periods of time. We measure the excitation spectrum when the dyes are used to stain biological specimens to get a more accurate representation of the spectrum of the dye in a cell environment as compared to solution-based measurements.

On the minimum requirements for otolaryngology clinics in National Health Service hospitals.

OBJECTIVE: This paper, a report by the Clinical Governance and Audit Committee of the Scottish Otolaryngological Society, presents a consensus view of the minimal requirements for ENT clinics in National Health Service hospitals. RESULTS AND CONCLUSION: The provision of adequate equipment and staff has gained increasing importance as the vast majority of ENT procedures can be safely performed in the out-patient or office setting.

C1 inhibitor deficiency: 2014 United Kingdom consensus document.

C1 inhibitor deficiency is a rare disorder manifesting with recurrent attacks of disabling and potentially life-threatening angioedema. Here we present an updated 2014 United Kingdom consensus document for the management of C1 inhibitor-deficient patients, representing a joint venture between the United Kingdom Primary Immunodeficiency Network and Hereditary Angioedema UK. To develop the consensus, we assembled a multi-disciplinary steering group of clinicians, nurses and a patient representative. This steering group first met in 2012, developing a total of 48 recommendations across 11 themes. The statements were distributed to relevant clinicians and a representative group of patients to be scored for agreement on a Likert scale. All 48 statements achieved a high degree of consensus, indicating strong alignment of opinion. The recommendations have evolved significantly since the 2005 document, with particularly notable developments including an improved evidence base to guide dosing and indications for acute treatment, greater emphasis on home therapy for acute attacks and a strong focus on service organization.

A UK national audit of hereditary and acquired angioedema.

Hereditary angioedema (HAE) and acquired angioedema (AAE) are rare life-threatening conditions caused by deficiency of C1 inhibitor (C1INH). Both are characterized by recurrent unpredictable episodes of mucosal swelling involving three main areas: the skin, gastrointestinal tract and larynx. Swelling in the gastrointestinal tract results in abdominal pain and vomiting, while swelling in the larynx may be fatal. There are limited UK data on these patients to help improve practice and understand more clearly the burden of disease. An audit tool was designed, informed by the published UK consensus document and clinical practice, and sent to clinicians involved in the care of HAE patients through a number of national organizations. Data sets on 376 patients were received from 14 centres in England, Scotland and Wales. There were 55 deaths from HAE in 33 families, emphasizing the potentially lethal nature of this disease. These data also show that there is a significant diagnostic delay of on average 10 years for type I HAE, 18 years for type II HAE and 5 years for AAE. For HAE the average annual frequency of swellings per patient affecting the periphery was eight, abdomen 5 and airway 0·5, with wide individual variation. The impact on quality of life was rated as moderate or severe by 37% of adult patients. The audit has helped to define the burden of disease in the UK and has aided planning new treatments for UK patients.

In pursuit of excellence: an integrated care pathway for C1 inhibitor deficiency.

There are estimated to be approximately 1500 people in the United Kingdom with C1 inhibitor (C1INH) deficiency. At BartsHealth National Health Service (NHS) Trust we manage 133 patients with this condition and we believe that this represents one of the largest cohorts in the United Kingdom. C1INH deficiency may be hereditary or acquired. It is characterized by unpredictable episodic swellings, which may affect any part of the body, but are potentially fatal if they involve the larynx and cause significant morbidity if they involve the viscera. The last few years have seen a revolution in the treatment options that are available for C1 inhibitor deficiency. However, this occurs at a time when there are increased spending restraints in the NHS and the commissioning structure is being overhauled. Integrated care pathways (ICP) are a tool for disseminating best practice, for facilitating clinical audit, enabling multi-disciplinary working and for reducing health-care costs. Here we present an ICP for managing C1 inhibitor deficiency.

Increased signals from short-wavelength-excited fluorescent molecules using sub-Ti:Sapphire wavelengths.

We report the use of an all-solid-state ultrashort pulsed source specifically for two-photon microscopy at wavelengths shorter than those of the conventional Ti:Sapphire laser. Our approach involves sum-frequency mixing of the output from an optical parametric oscillator (λ= 1400-1640 nm) synchronously pumped by a Yb-doped fibre laser (λ= 1064 nm), with the residual pump radiation. This generated an fs-pulsed output tunable in the red spectral region (λ= 620-636 nm, ~150 mW, 405 fs, 80 MHz, M(2) ~ 1.3). We demonstrate the performance of our ultrashort pulsed system using fluorescently labelled and autofluorescent tissue, and compare with conventional Ti:Sapphire excitation. We observe a more than 3-fold increase in fluorescence signal intensity using our visible laser source in comparison with the Ti:Sapphire laser for two-photon excitation at equal illumination peak powers of 1.16 kW or less.

A compact instrument for adjusting laser beams to be accurately coincident and coaxial and its use in biomedical imaging using wave-mixed laser sources.

Biomedical imaging applications that involve nonlinear optical processes such as sum-frequency generation (SFG) and four-wave mixing require that the pulses are synchronized in time and the beams are coaxial to better than 400 µrad. For this reason, folding mirrors are normally used to extend the beam path over a few meters so that detectors can be put into the beams to check their overlap at the start of a long path and also at the end of it. We have made a portable instrument with a footprint of only 22 cm × 11 cm × 16 cm that uses a short focal length lens and a telephoto combination for viewing the near-field and far-field simultaneously. Our instrument is simple to build and use, and we show its application in coherent anti-Stokes Raman scattering microscopy and SFG-based two-photon fluorescence microscopy.

Transient outward K+ current reduction prolongs action potentials and promotes afterdepolarisations: a dynamic-clamp study in human and rabbit cardiac atrial myocytes.

Human atrial transient outward K(+) current (I(TO)) is decreased in a variety of cardiac pathologies, but how I(TO) reduction alters action potentials (APs) and arrhythmia mechanisms is poorly understood, owing to non-selectivity of I(TO) blockers. The aim of this study was to investigate effects of selective I(TO) changes on AP shape and duration (APD), and on afterdepolarisations or abnormal automaticity with ß-adrenergic-stimulation, using the dynamic-clamp technique in atrial cells. Human and rabbit atrial cells were isolated by enzymatic dissociation, and electrical activity recorded by whole-cell-patch clamp (35-37°C). Dynamic-clamp-simulated I(TO) reduction or block slowed AP phase 1 and elevated the plateau, significantly prolonging APD, in both species. In human atrial cells, I(TO) block (100% I(TO) subtraction) increased APD(50) by 31%, APD(90) by 17%, and APD(-61 mV) (reflecting cellular effective refractory period) by 22% (P < 0.05 for each). Interrupting I(TO) block at various time points during repolarisation revealed that the APD(90) increase resulted mainly from plateau-elevation, rather than from phase 1-slowing or any residual I(TO). In rabbit atrial cells, partial I(TO) block (∼40% I(TO) subtraction) reversibly increased the incidence of cellular arrhythmic depolarisations (CADs; afterdepolarisations and/or abnormal automaticity) in the presence of the ß-agonist isoproterenol (0.1 µm; ISO), from 0% to 64% (P < 0.05). ISO-induced CADs were significantly suppressed by dynamic-clamp increase in I(TO) (∼40% I(TO) addition). ISO+I(TO) decrease-induced CADs were abolished by ß(1)-antagonism with atenolol at therapeutic concentration (1 µm). Atrial cell action potential changes from selective I(TO) modulation, shown for the first time using dynamic-clamp, have the potential to influence reentrant and non-reentrant arrhythmia mechanisms, with implications for both the development and treatment of atrial fibrillation.

Reduced postsynaptic GABAA receptor number and enhanced gaboxadol induced change in holding currents in Purkinje cells of the dystrophin-deficient mdx mouse.

UNLABELLED: Duchenne muscular dystrophy (DMD) is caused by the absence of a functional transcript of the protein dystrophin. DMD is associated with a range of cognitive deficits that are thought to result from a lack of the protein dystrophin in brain structures involved in cognitive functions. The CNS involvement extends to an impairment of cognitive abilities, with many DMD boys having significant reduction in IQ. In the cerebellum, dystrophin is normally localized at the postsynaptic membrane of GABAergic synapses on Purkinje cells. Here, we investigate the effect of an absence of dystrophin on the number of GABA(A) channels located at the synapse in cerebellar Purkinje cells of the dystrophin-deficient mdx mouse. Whole-cell patch-clamp recordings of spontaneous miniature inhibitory postsynaptic currents (mIPSCs) were performed in cerebellar slices from mdx and littermate control mice. Our results showed that the number of receptors at GABAergic synapses in the cerebellar Purkinje cell was significantly reduced in mdx mice (38.38 ± 2.95) compared to littermate controls (53.03 ± 4.11). Furthermore, when gaboxadol was added to the bath, the change in holding current in mdx mice was significantly enhanced (65.01 ± 5.89pA) compared to littermate controls (37.36 ± 3.82pA). The single channel unitary conductance and the rise and decay time of mIPSCs were not significantly different in these two groups of mice, indicating that those GABA(A) channels located at the postsynaptic sites in the mdx mice function normally. CONCLUSION: There is a reduction in the number of functional receptors localized at GABAergic synapses in the cerebellar Purkinje cells of dystrophin-deficient mdx mice and an increase in a gaboxadol induced holding current, which is evidence for an increase in extrasynaptic GABA(A) receptors in mdx mice. We hypothesize that the absence of dystrophin, from mdx Purkinje cells, reduces the number of post-synaptic GABA(A) receptors and as a result there is an increase in extrasynaptic receptors. If similar changes occur in the CNS in boys with DMD, it will impact on the function of neural networks and may contribute to some of the motor, behavioral and cognitive impairment apparent in many boys with DMD.

Assessment of sarcoplasmic reticulum Ca2+ depletion during spontaneous Ca2+ waves in isolated permeabilized rabbit ventricular cardiomyocytes.

In this study, Ca2+ release due to spontaneous Ca2+ waves was measured both from inside the sarcoplasmic reticulum (SR) and from the cytosol of rabbit cardiomyocytes. These measurements utilized Fluo5N-AM for intra-SR Ca2+ from intact cells and Fluo5F in the cytosol of permeabilized cells. Restricted subcellular volumes were resolved with the use of laser-scanning confocal microscopy. Local Ca2+ signals during spontaneous Ca2+ release were compared with those induced by rapid caffeine application. The free cytoplasmic [Ca2+] increase during a Ca2+ wave was 98.1% +/- 0.3% of that observed during caffeine application. Conversion to total Ca2+ release suggested that Ca2+ release from a Ca2+ wave was not significantly different from that released during caffeine application (104% +/- 6%). In contrast, the maximum decrease in intra-SR Fluo-5N fluorescence during a Ca2+ wave was 82.5% +/- 2.6% of that observed during caffeine application. Assuming a maximum free [Ca2+] of 1.1 mM, this translates to a 96.2% +/- 0.8% change in intra-SR free [Ca2+] and a 91.7% +/- 1.6% depletion of the total Ca2+. This equates to a minimum intra-SR free Ca2+ of 46 +/- 7 microM during a Ca2+ wave. Reduction of RyR2 Ca2+ sensitivity by tetracaine (50 microM) reduced the spontaneous Ca2+ release frequency while increasing the Ca2+ wave amplitude. This did not significantly change the total depletion of the SR (94.5% +/- 1.1%). The calculated minimum [Ca2+] during these Ca2+ waves (87 +/- 19 microM) was significantly higher than control (p < 0.05). A computational model incorporating this level of Ca2+ depletion during a Ca2+ wave mimicked the transient and sustained effects of tetracaine on spontaneous Ca2+ release. In conclusion, spontaneous Ca2+ release results in substantial but not complete local Ca2+ depletion of the SR. Furthermore, measurements suggest that Ca2+ release terminates when luminal [Ca2+] reaches approximately 50 microM.

Importance of vagally mediated bradycardia for the induction of torsade de pointes in an in vivo model.

BACKGROUND AND PURPOSE: Bradycardia is a risk factor for the development of torsade de pointes (TdP). The aim of this work was to compare the importance of changes in heart rate and arterial blood pressure in the development of drug-induced TdP and to investigate the role of vagal influences. EXPERIMENTAL APPROACH: Experiments were performed in open-chest, pentobarbital-anaesthetized, male rabbits which were given clofilium (20, 60 and 200 nmol kg(-1) min(-1)) with rising doses of either phenylephrine (75, 150, 225 and 300 nmol kg(-1) min(-1)), angiotensin II (0.25, 0.5, 0.75 and 1 nmol kg(-1) min(-1)) or saline. A fourth group received phenylephrine and cloflium after bilateral vagotomy. ECGs, haemodynamics and epicardial monophasic action potentials were recorded. KEY RESULTS: TdP occurred in 57% of rabbits given phenylephrine and clofilium. Replacement of phenylephrine with saline or angiotensin II reduced the incidence of TdP to 0 and 17%, respectively. Vagotomy prevented TdP in rabbits given phenylephrine and clofilium. Increases in blood pressure induced by phenylephrine and angiotensin II were similar. Bradycardia only occurred with phenylephrine and was reduced but not abolished by vagotomy. Neither short-term variability of repolarization nor action potential triangulation could predict TdP. CONCLUSIONS AND IMPLICATIONS: These results indicate that reflex activation of vagal nerve activity is essential for the induction of drug-induced TdP in alpha1-adrenoceptor-stimulated anaesthetized rabbits. This implies that alterations in vagal activity may also precipitate episodes of drug-induced TdP in man and that this should be considered in selecting models used in drug development.

Potentiation of E-4031-induced torsade de pointes by HMR1556 or ATX-II is not predicted by action potential short-term variability or triangulation.

BACKGROUND AND PURPOSE: Torsade de pointes (TdP) can be induced by a reduction in cardiac repolarizing capacity. The aim of this study was to assess whether IKs blockade or enhancement of INa could potentiate TdP induced by IKr blockade and to investigate whether short-term variability (STV) or triangulation of action potentials preceded TdP. EXPERIMENTAL APPROACH: Experiments were performed in open-chest, pentobarbital-anaesthetized, alpha 1-adrenoceptor-stimulated, male New Zealand White rabbits, which received three consecutive i.v. infusions of either the IKr blocker E-4031 (1, 3 and 10 nmol kg(-1) min(-1)), the IKs blocker HMR1556 (25, 75 and 250 nmol kg(-1) min(-1)) or E-4031 and HMR1556 combined. In a second study rabbits received either the same doses of E-4031, the INa enhancer, ATX-II (0.4, 1.2 and 4.0 nmol kg(-1)) or both of these drugs. ECGs and epicardial monophasic action potentials were recorded. KEY RESULTS: HMR1556 alone did not cause TdP but increased E-4031-induced TdP from 25 to 80%. ATX-II alone caused TdP in 38% of rabbits, as did E-4031; 75% of rabbits receiving both drugs had TdP. QT intervals were prolonged by all drugs but the extent of QT prolongation was not related to the occurrence of TdP. No changes in STV were detected and triangulation was only increased after TdP occurred. CONCLUSIONS AND IMPLICATIONS: Giving modulators of ion channels in combination substantially increased TdP but, in this model, neither STV nor triangulation of action potentials could predict TdP.

Measurement and modeling of Ca2+ waves in isolated rabbit ventricular cardiomyocytes.

The time course and magnitude of the Ca(2+) fluxes underlying spontaneous Ca(2+) waves in single permeabilized ventricular cardiomyocytes were derived from confocal Fluo-5F fluorescence signals. Peak flux rates via the sarcoplasmic reticulum (SR) release channel (RyR2) and the SR Ca(2+) ATPase (SERCA) were not constant across a range of cellular [Ca(2+)] values. The Ca(2+) affinity (K(mf)) and maximum turnover rate (V(max)) of SERCA and the peak permeability of the RyR2-mediated Ca(2+) release pathway increased at higher cellular [Ca(2+)] loads. This information was used to create a computational model of the Ca(2+) wave, which predicted the time course and frequency dependence of Ca(2+) waves over a range of cellular Ca(2+) loads. Incubation of cardiomyocytes with the Ca(2+) calmodulin (CaM) kinase inhibitor autocamtide-2-related inhibitory peptide (300 nM, 30 mins) significantly reduced the frequency of the Ca(2+) waves at high Ca(2+) loads. Analysis of the Ca(2+) fluxes suggests that inhibition of CaM kinase prevented the increases in SERCA V(max) and peak RyR2 release flux observed at high cellular [Ca(2+)]. These data support the view that modification of activity of SERCA and RyR2 via a CaM kinase sensitive process occurs at higher cellular Ca(2+) loads to increase the maximum frequency of spontaneous Ca(2+) waves.

BAHNO surgical specialities: same patients, different practices?

The British Association of Head and Neck Oncologists (BAHNO) is 'a multidisciplinary society for healthcare professionals involved in the study and treatment of head and neck cancer'. Surgical members of this organization are from three specialities (otolaryngology, maxillo-facial and plastic surgery). Although the overall impression is that the management of UK head and neck cancer patients is consensus based, there are appreciable differences in each surgical speciality's practice. Anecdotally, this can lead to variation in the management of very similar patients. To identify some of these variations BAHNO surgeons were surveyed regarding their current head and neck cancer practices from the perspectives of surgical activity and post-operative care. Some unexpected differences were identified, particularly in relation to post-operative care with plastic and maxillo-facial surgeons demonstrating different patterns of high dependency unit (HDU) and intensive care unit (ICU) use for the same patients. The implications for future consensus in the light of these variations are discussed.

The use of hyaluronidase in nasal infiltration: prospective randomized controlled pilot study.

A double-blind randomized prospective case-control pilot study was performed to assess tissue distortion caused by the infiltration of local anaesthetic to the dorsum of the nose and to see if this was altered by the addition of hyaluronidase. Forty patients undergoing nasal manipulation for fractured nasal bones were randomized to receive either 4 ml of two per cent lignocaine and adrenaline 1:200000 or 4 ml of two per cent lignocaine and adrenaline 1:200000 with 1500 IU hyaluronidase, which was infiltrated subcutaneously over the nasal dorsum. One surgeon using a standardized technique performed the nasal infiltration. Other outcome measures were ease of manipulation, adequacy of the reduction, patient satisfaction with cosmesis and patient analgesia requirements. There were trends for decreased tissue distortion and improved ease of manipulation in the hyaluronidase group. Larger trials are required to confirm these results.

Wegener's granulomatosis: an analysis of 33 patients seen over a 10-year period.

This retrospective study examined 33 patients with Wegener's granuloma seen between 1990 and 1999 in the Ayrshire and Arran region of Scotland. There was an estimated annual incidence of 10/million/year. The diagnosis in this series was based on the presence of one or more of the following: a positive histology, a positive c-ANCA or a typical clinical course. Twenty-eight patients were diagnosed based on either a positive histology and/or c-ANCA, whereas the remaining five were diagnosed based on the clinical course. c-ANCA was positive in 79.3% but correlated poorly with disease activity. Nasal biopsies were positive in 40%, whereas 94% of renal biopsies were positive, thereby making nasal biopsies unreliable in the diagnosis. Significantly elevated levels of erythrocyte sedimentation rate (ESR) averaging 77 mm/h were found in 32 patients at diagnosis. This showed fluctuation with disease activity. Thirteen patients died, 12 within 5 years. The best prognostic indicator statistically was age.

Adenoid cystic carcinoma of the nasal septum.

Adenoid cystic carcinoma is a malignant tumour frequently described arising from seromucinous salivary tissue in the major and minor salivary glands. Within the nasal cavity, it is uncommon and usually involves the lateral wall. A rare case of adenoid cystic carcinoma of the nasal septum is presented along with a review of the literature. The presentation and management of this uncommon condition is discussed.

Tamoxifen therapy in steroid-resistant Riedels disease.

Riedels thyroiditis is a rare chronic inflammatory disorder characterised by extensive fibrosis of the thyroid gland and sometimes the surrounding tissues. We report a case of Riedels Thyroiditis in a middle aged female presenting with goitre, stridor and dyspnoea. She initially responded to corticosteroid treatment and subsequently to tamoxifen. The rationale for these treatments are discussed.