Positive allosteric modulators of ␣7 nicotinic acetylcholine receptors (nAChRs) have attracted considerable interest as potential tools for the treatment of neurological and psychiatric disorders such as Alzheimer's disease and schizophrenia. However, despite the potential therapeutic usefulness of these compounds, little is known about their mechanism of action. Here, we have examined two allosteric potentiators of ␣7 nAChRs (PNU-120596 and LY-2087101). From studies with a series of subunit chimeras, we have identified the transmembrane regions of ␣7 as being critical in facilitating potentiation of agonist-evoked responses. Furthermore, we have identified five transmembrane amino acids that, when mutated, significantly reduce potentiation of ␣7 nAChRs. The amino acids we have identified are located within the ␣-helical transmembrane domains TM1 (S222 and A225), TM2 (M253), and TM4 (F455 and C459). Mutation of either A225 or M253 individually have particularly profound effects, reducing potentiation of EC 20 concentrations of acetylcholine to a tenth of the level seen with wild-type ␣7. Reference to homology models of the ␣7 nAChR, based on the 4Å structure of the Torpedo nAChR, indicates that the side chains of all five amino acids point toward an intrasubunit cavity located between the four ␣-helical transmembrane domains. Computer docking simulations predict that the allosteric compounds such as PNU-120596 and LY-2087101 may bind within this intrasubunit cavity, much as neurosteroids and volatile anesthetics are thought to interact with GABA A and glycine receptors. Our findings suggest that this is a conserved modulatory allosteric site within neurotransmitter-gated ion channels.allosteric modulators ͉ neurotransmitter receptor N icotinic acetylcholine receptors (nAChRs) are excitatory neurotransmitter-gated ion channels and members of a superfamily that also includes ionotropic receptors for 5-hydroxytryptamine (5-HT; serotonin), glycine and ␥-aminobutyric acid (GABA) (1, 2). Nicotinic receptors are allosteric proteins (3), which have been implicated in a variety of neurological and psychiatric disorders, including Alzheimer's disease, Parkinson's disease, epilepsy, and schizophrenia (4-6). As a consequence, nAChRs are viewed as being important targets for therapeutic drug discovery (7,8).Although most nAChR subunits assemble only into heteromeric nAChRs (9, 10), the ␣7 and ␣8 subunits are important exceptions. Both ␣7 and ␣8 are able to generate functional homomeric nAChRs (11, 12) and have much closer sequence similarity to each other than to other nAChR subunits (2, 9). Whereas there is evidence that ␣7 nAChRs are an important receptor subtype in the mammalian brain (and in other vertebrates), the ␣8 subunit has been identified only in avian species. The 5-HT receptor 3A subunit (5-HT 3A ) has close sequence similarity to nAChR subunits (2) and, like the nAChR ␣7 and ␣8 subunits, is able to generate functional homomeric cationselective ion channels (13). Indeed, the ability of ␣7 and 5-HT 3A subunits to...

This paper describes the design and synthesis of a photostable, far-red to near-infrared (NIR) platform for optical voltage sensing. We developed a new, sulfonated silicon rhodamine fluorophore and integrated it with a phenylenevinylene molecular wire to create a Berkeley Red Sensor of Transmembrane potential, or BeRST 1 (“burst”). BeRST 1 is the first member of a class of farred to NIR voltage sensitive dyes that make use of a photoinduced electron transfer (PeT) trigger for optical interrogation of membrane voltage. We show that BeRST 1 displays bright, membrane-localized fluorescence in living cells, high photostability, and excellent voltage sensitivity in neurons. Depolarization of the plasma membrane results in rapid fluorescence increases (24% ΔF/F per 100 mV). BeRST 1 can be used in conjunction with fluorescent stains for organelles, Ca2+ indicators, and voltage-sensitive fluorescent proteins. In addition, the red-shifted spectral profile of BeRST 1, relative to commonly employed optogenetic actuators like ChannelRhodopsin2 (ChR2), which require blue light, enables optical electrophysiology in neurons. The high speed, sensitivity, photostability and long-wavelength fluorescence profiles of BeRST 1 make it a useful platform for the non-invasive, optical dissection of neuronal activity.

Purpose The COVID-19 pandemic is placing unprecedented demand upon critical care services for invasive mechanical ventilation. There is current uncertainty regarding the role of tracheostomy for weaning ventilated patients with COVID-19 pneumonia. This is due to a number of factors including prognosis, optimal healthcare resource utilisation, and safety of healthcare workers when performing such a high-risk aerosol-generating procedure. Methods Literature review and proposed practical guideline based on the experience of a tertiary healthcare institution with 195 critical care admissions for COVID-19 up until 4th April 2020. Results A synthesis of the current international literature and reported experience is presented with respect to prognosis, viral load and staff safety, thus leading to a pragmatic recommendation that tracheostomy is not performed until at least 14 days after endotracheal intubation in COVID-19 patients. Practical steps to minimise aerosol generation in percutaneous tracheostomy are outlined and we describe the process and framework for setting up a dedicated tracheostomy team. Conclusion In selected COVID-19 patients, there is a role for tracheostomy to aid in weaning and optimise healthcare resource utilisation. Both percutaneous and open techniques can be performed safely with careful modifications to technique and appropriate enhanced personal protective equipment. ORL-HNS surgeons can play a valuable role in forming tracheostomy teams to support critical care teams during this global pandemic.

BackgroundInitial reports describing COVID-19 were dominated by the presence of cough, breathlessness, and fever, anecdotal reports suggested anosmia may also be a manifestation. We sought to use Google Trends (GT) to investigate whether there was a surge in individuals searching for information related . Spearman rank analysis was performed to correlate loss of smell relative search volumes (RSV) with the increases of daily confirmed cases of COVID-19 and deaths attributed to disease. As a control event, we also performed analysis of smell-related searches during the last UK Influenza epidemic of 2009. ResultsIn all three countries, we observed strong correlations between daily RSVs related to loss of smell, increases of daily COVID-19+ cases and deaths ranging from 0.633 to 0.952. All correlations were statistically significant (p<0.05). ConclusionThere is a strong correlation between the frequency of searches for smell-related information and the onset of COVID-19 infection in Italy, Spain, UK, USA, Germany, France, Iran and Netherlands. We would hypothesise this may relate to a previously under-recognised symptom.

How features of the visual scene are encoded in the population activity of retinal ganglion cells (RGCs) targeting specific regions of the brain is not well understood. To address this, we have used a genetically encoded reporter of presynaptic function (SyGCaMP3) to record visually evoked activity in the population of RGC axons innervating the zebrafish tectum. Using unbiased voxel-wise analysis of SyGCaMP3 signals, we identify three subtypes of direction-selective and two subtypes of orientation-selective retinal input. Composite parametric functional maps generated across many larvae show laminar segregation of direction- and orientation-selective responses and unexpected retinotopic biases in the distribution of functional subtypes. These findings provide a systematic description of the form, organization, and dimensionality of visual inputs to the brain and will serve as a platform for understanding emergent properties in tectal circuits associated with visually driven behavior.

An understanding of an individual's level of grit may be used to identify doctors at a greater risk of burnout. As a high level of grit is associated with less burnout, interventions to improve grit through resilience training should be examined. Further research is needed to understand how grit levels change during a doctor's career and why GPs experience higher levels of burnout.

BackgroundTraining in patient and public involvement (PPI) is recommended, yet little is known about what training is needed. We explored researchers’ and PPI contributors’ accounts of PPI activity and training to inform the design of PPI training for both parties.MethodsWe used semi-structured qualitative interviews with researchers (chief investigators and trial managers) and PPI contributors, accessed through a cohort of clinical trials, which had been funded between 2006 and 2010. An analysis of transcripts of audio-recorded interviews drew on the constant comparative method.ResultsWe interviewed 31 researchers and 17 PPI contributors from 28 trials. Most researchers could see some value in PPI training for researchers, although just under half had received such training themselves, and some had concerns about the purpose and evidence base for PPI training. PPI contributors were evenly split in their perceptions of whether researchers needed training in PPI. Few PPI contributors had themselves received training for their roles. Many informants across all groups felt that training PPI contributors was unnecessary because they already possessed the skills needed. Informants were also concerned that training would professionalise PPI contributors, limiting their ability to provide an authentic patient perspective. However, informants welcomed informal induction ‘conversations’ to help contributors understand their roles and support them in voicing their opinions. Informants believed that PPI contributors should be confident, motivated, intelligent, focussed on helping others and have relevant experience. Researchers looked for these qualities when selecting contributors, and spoke of how finding ‘the right’ contributor was more important than accessing ‘the right’ training.ConclusionsWhile informants were broadly receptive to PPI training for researchers, they expressed considerable reluctance to training PPI contributors. Providers of training will need to address these reservations. Our findings point to the importance of reconsidering how training is conceptualised, designed and promoted and of providing flexible, learning opportunities in ways that flow from researchers’ and contributors’ needs and preferences. We also identify some areas of training content and the need for further consideration to be given to the selection of PPI contributors and models for implementing PPI to ensure clinical trials benefit from a diversity of patient perspectives.

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