PHARMACOLOGICAL REGULATION OF TREK1, TREK2 AND TRESK TWO PORE DOMAIN POTASSIUM CHANNELS

ABSTRACT Introduction: Two pore domain potassium (K2P) channels are responsible for background currents that regulate membrane potential and neuronal excitability. Compounds which alter the activity of these channels are predicted to have therapeutic potential in treating CNS disorders. Members of the TREK family of K2P channels (TREK1 and TREK2) have been shown to play an active role in neuroprotection, depression and pain, whilst TRESK, with high expression in sensory neurons, has a role in nociception. Sipatrigine, a neuroprotective agent and a derivative of the anticonvulsant lamotrigine, is a known antagonist of TREK channels whilst lamotrigine is thought to primarily inhibit TRESK channels. A new compound, Cen-092-C, has also been developed which is structurally similar to lamotrigine. However, its effects on K2P channels are unknown. To understand the mechanism of channel inhibition by drugs, the structure of TREK2 was solved and was co-crystallised with norfluoxetine. This showed that fenestration sites were important in channel and current inhibition. Furthermore, TRESK docking studies showed that F145 and F352 function in a similar way to TREK2 fenestration site, as the bulky phenylalanine faces into the pore, and are thought to be important for compound binding. The aim of this study is to clarify to differences in the inhibitory effect of these compounds on the selected K2P channels and to investigate the mechanism by which these compounds inhibit the channels current. Methods: Wild-type (WT) and mutated human K2P channels were transiently expressed in tsA-201 cells. The currents were measured using whole-cell patch-clamp electrophysiology. Results: Sipatrigine was shown to inhibit both TREK1 and TREK2 current. Lamotrigine was also found to inhibit TREK1 and to a lesser extent TREK2. Cen-092-C was found to be less effective on TREK1 and TRESK current compared to sipatrigine, but similar to lamotrigine results. The sipatrigine inhibitory effect, but not lamotrigine, was reduced by mutations on the M4 region at the fenestration site of TREK1 and TREK2 (L286 and L320). This sensitivity is selective at this site as other mutations in the central cavity showed no change in sipatrigine inhibition. Interestingly, the gain-of-function mutation (TREK1 E306A) on the C terminus showed a reduced sipatrigine inhibition. The effect of sipatrigine on TREK2 showed an over-recovery of current following wash-off of the compound. The wash-off current increase was not seen if the N-terminus length is forced into intermediate and short isoform. Sipatrigine inhibition was significantly decreased when the N-terminus was truncated. Sipatrigine has been shown to strongly inhibit TRESK. Lamotrigine was seen to inhibit TRESK current, however significantly less effective compared to sipatrigine. Furthermore, lamotrigine did show state dependent inhibition when TRESK is in the fixed activated state. Cen-092-C was also found to inhibit TRESK to a similar degree to lamotrigine, however there was no state dependent inhibition on TRESK current. The effects of these antagonists on TRESK has been shown to be abolished by mutations on two sites at the central cavity (F145 and F352). Conclusion: Lamotrigine was found not to be TRESK selective, contrary to other studies. Sipatrigine and lamotrigine inhibition works through binding to the channel. The fenestration site in both TREK1 and TREK2 has been found to be an important binding site of sipatrigine, differing from lamotrigine. This suggests that the structurally similar compounds bind to different regions of the TREK channels. Furthermore, the over recovery of TREK2 current after sipatrigine wash off is believed to show the compound's biphasic effect, where the underlying enhancement of current is hidden by the action of inhibition. The N-terminus is therefore believed to be important in regulating sipatrigine action on TREK2. It remains unclear whether the TRESK potential binding sites (F1452 and F352) are important in compound binding as the inserted mutation is believed to shift the channel to constant active state. The newly developed compound Cen-092-C shows a significantly greater degree of inhibition of TRESK when compared to TREK1. Cen-092-C and lamotrigine inhibition of TRESK is not significantly different. Lamotrigine inhibition of TRESK current is state dependent whereas sipatrigine and Cen-092-C inhibition of TRESK current is shown as state independent. All of this together could lead to a better understanding of how neuroprotective agents effect TREK and TRESK channels and could contribute to the design of more efficient ligands

What and How Do Students Learn in an Interprofessional Student-Run Clinic? An Educational Framework for Team-Based Care

Background: The student-run clinic (SRC) has the potential to address interprofessional learning among health professions students. Purpose: To derive a framework for understanding student learning during team-based care provided in an interprofessional SRC serving underserved patients. Methods: The authors recruited students for a focus group study by purposive sampling and snowballing. They constructed two sets of semi-structured questions for uniprofessional and multiprofessional groups. Sessions were audiotaped, and transcripts were independently coded and adjudicated. Major themes about learning content and processes were extracted. Grounded theory was followed after data synthesis and interpretation to establish a framework for interprofessional learning. Results: Thirty-six students from four professions (medicine, physician assistant, occupational therapy, and pharmacy) participated in eight uniprofessional groups; 14 students participated in three multiprofessional groups (N50). Theme saturation was achieved. Six common themes about learning content from uniprofessional groups were role recognition, team-based care appreciation, patient experience, advocacy-/systemsbased models, personal skills, and career choices. Occupational therapy students expressed self-advocacy, and medical students expressed humility and self-discovery. Synthesis of themes from all groups suggests a learning continuum that begins with the team huddle and continues with shared patient care and social interactions. Opportunity to observe and interact with other professions in action is key to the learning process. Discussion: Interprofessional SRC participation promotes learning ‘with, from, and about’ each other. Participation challenges misconceptions and sensitizes students to patient experiences, health systems, advocacy, and social responsibility. Learning involves interprofessional interactions in the patient encounter, reinforced by formal and informal communications. Participation is associated with interest in serving the underserved and in primary care careers. The authors proposed a framework for interprofessional learning with implications for optimal learning environments to promote team-based care. Future research is suggested to identify core faculty functions and best settings to advance and enhance student preparation for future collaborative team practice

Fourteen Draft Genome Sequences for the First Reported Cases of Azithromycin-Resistant Neisseria gonorrhoeae in Ireland

Here, we report the draft genome assemblies of 14 azithromycin-resistantNeisseria gonorrhoeaeclinical isolates, representing the first such strains identified in Ireland. Among these isolates are the first reported highly resistant strains (MIC >256mg/liter), which both belonged to the ST1580 sequence type

BUDGET PERSPECTIVES 2007

1. DISABILITY BENEFIT – CONTROLLED OR UNDER-CONTROLLED? Brenda Gannon p. 3 2. CHILD POVERTY AND CHILD INCOME SUPPORTS: IRELAND IN COMPARATIVE PERSPECTIVE Tim Callan, Kieran Coleman, Brian Nolan and John Walsh p. 23 3. STATE FINANCIAL SUPPORT FOR HORSE RACING IN IRELAND Tony Fahey and Liam Delaney p. 3

Crystal structure of an HD-GYP domain cyclic-di-GMP phosphodiesterase reveals an enzyme with a novel trinuclear catalytic iron centre

Bis-(3′,5′) cyclic di-guanylate (c-di-GMP) is a key bacterial second messenger that is implicated in the regulation of many crucial processes that include biofilm formation, motility and virulence. Cellular levels of c-di-GMP are controlled through synthesis by GGDEF domain diguanylate cyclases and degradation by two classes of phosphodiesterase with EAL or HD-GYP domains. Here, we have determined the structure of an enzymatically active HD-GYP domain protein from Persephonella marina (PmGH) alone, in complex with substrate (c-di-GMP) and final reaction product (GMP). The structures reveal a novel trinuclear iron binding site, which is implicated in catalysis and identify residues involved in recognition of c-di-GMP. This structure completes the picture of all domains involved in c-di-GMP metabolism and reveals that the HD-GYP family splits into two distinct subgroups containing bi- and trinuclear metal centres.</p

Educating professionals who will work with children in the early years: an evidence-informed interdisciplinary framework

© 2018 TACTYC “This is an Accepted Manuscript of an article published by Taylor & Francis in Early Years on 6 July 2018, available online: http://www.tandfonline.com/10.1080/09575146.2018.1488819” This author accepted manuscript is made available following 12 month embargo from date of publication (July 2018) in accordance with the publisher’s archiving policyThe first five years of a child’s life are irrefutably important, establishing life-long health, social and economic outcomes. To optimize these outcomes, global policy is directing professionals from a range of disciplinary backgrounds to work more collaboratively than ever before with children in the early years. Such collaborations have proven problematic as individual disciplines and pre-service education requirements vary widely. Using Community-Based Participatory Research and Diffusion of Innovation approaches, this study aimed to develop an educational framework for professionals working with children in the early years and their families, to begin a cultural change for interdisciplinary collaboration and participation across the early years. Systematic reviews, modified Delphi rounds and focus groups identified the diverse demands of multiple professions, qualification levels and workforce agendas, as well as highlighting shared outcomes, knowledge and intentions across disciplines

Interfacing Relational Frame Theory with Cognitive Neuroscience: Semantic Priming, The Implicit Association Test, and Event Related Potentials

The current article argues that an important component of the research agenda for Relational Frame Theory will involve studying the functional relations that obtain between environmental events and the physiological activity that takes place inside the brain and central nervous system, with a particular focus on human language and cognition. In support of this view, five separate experiments are outlined. The first three experiments replicate and extend previous research reported by Hayes and Bisset (1998). Specifically, the research, using both reaction time and neurophysiological measures, supports the argument that there is a clear functional overlap between semantic and derived stimulus relations. Specifically, an evoked potential waveform typically associated with semantic processing (N400) is shown to be sensitive to equivalence versus non-equivalence relations. Experiments 4 and 5 indicate that these reaction time and evoked potential effects are not restricted to traditional lexical decision tasks, but can also be observed using the implicit association test. Furthermore, preliminary evidence suggests that evoked potentials might constitute a more sensitive measure of derived stimulus relations than response time. The results obtained across all five experiments support the view that the study of derived stimulus relations, combined with some of the procedures and measures of cognitive psychology and cognitive neuroscience, may provide an important inroad into the experimental analysis of semantic relations in human language