Abnormal response to chronic social defeat stress and fear extinction in a mouse model of cholinergic dysregulation.

Cholinergic signaling is critical for an individual to react appropriately and adaptably to salient stimuli while navigating a complex environment. The cholinergic neurotransmitter system drives attention to salient stimuli, such as stressors, and aids in orchestrating the proper neural and behavioral response. Fine-tuned regulation of the cholinergic system has been linked to appropriate stress responses and subsequent mood regulation while dysregulation has been implicated in mood disorders. Among the multiple layers of regulation are cholinergic protein modulators. Here, we use validated models of experiential-based affective disorders to investigate differences in responses to stress in a genetic mouse model of cholinergic dysregulation based on the loss of protein modulator. The lynx2 nicotinic receptor modulatory protein provides negative cholinergic regulation within the amygdala, medial prefrontal cortex, and other brain regions. We discovered here that lynx2 knockout (KO) mice demonstrate an inability to update behavior with an inability to extinguish learned fear during a fear extinction test. We also observed, under an increased stress load following exposure to chronic social defeat stress (CSDS) paradigm, there was a unified resilience phenotype in lynx2KO mice, as opposed to the wild-type cohort which was split between resilience and susceptible phenotypes. Furthermore, we provide evidence for the functional role of α7 nicotinic receptor subtypes by phenotypic rescue with MLA or crossing with an α7 null mutant mouse (e.g. lynx2/α7 double KO mice). We demonstrate a direct physical interaction between lynx2 and α7 nAChR by co-immunoprecipitation of complexes from mouse BLA extracts. The genetic predisposition to heightened basal anxiety-like behavior and altered cholinergic signaling impairs individual behavior responses stressors. Together, these data indicate that the effects of social stress can be influenced by baseline genetic factors involved in anxiety regulation.

Biophysical characterization of lynx-nicotinic receptor interactions using atomic force microscopy.

Nicotinic acetylcholine receptors (nAChRs) are broadly expressed in the central and peripheral nervous systems, playing essential roles in cholinergic neurotransmission. The lynx family proteins, a subset of the Ly6/uPAR superfamily expressed in multiple brain regions, have been shown to bind to nAChRs and modulate their function via allosteric regulation. The binding interactions between lynx and nAChRs, however, have not been systematically quantified and compared. In this work, we characterized the interactions between lynx1 or lynx2 and α3ß4- or α7-nAChRs using single-molecule atomic force microscopy (AFM). The AFM technique allows the quantification of the off-rate of lynx-nAChR binding and of the energetic barrier width between the bound state and transition state, providing a biophysical means to compare the selectivity of lynx proteins for nAChR subtypes. Results indicate that lynx1 has a marginal preference for α7- over α3ß4-nAChRs. Strikingly, lynx2 exhibits a two order of magnitude stronger affinity for α3ß4- compared to α7-nAChRs. Together, the AFM assay serves as a valuable tool for the biophysical characterization of lynx-nAChR binding affinities. Revealing the differential affinities of lynx proteins for nAChR subtypes will help elucidate how lynx regulates nAChR-dependent functions in the brain, including nicotine addiction and other critical pathways.

Highlights from the Era of Open Source Web-Based Tools.

High digital connectivity and a focus on reproducibility are contributing to an open science revolution in neuroscience. Repositories and platforms have emerged across the whole spectrum of subdisciplines, paving the way for a paradigm shift in the way we share, analyze, and reuse vast amounts of data collected across many laboratories. Here, we describe how open access web-based tools are changing the landscape and culture of neuroscience, highlighting six free resources that span subdisciplines from behavior to whole-brain mapping, circuits, neurons, and gene variants.

Dynamics and Interactions of GPI-Linked lynx1 Protein with/without Nicotinic Acetylcholine Receptor in Membrane Bilayers.

Nicotinic acetylcholine receptors (nAChRs) participate in diverse biological processes, such as mood, learning, and addiction. Glycosylphosphatidylinositol-linked lynx1 is an allosteric modulator of nAChR function, including shifts in agonist sensitivity, reduced desensitization, and slower recovery from desensitization. This modulation is thought to be achieved by lynx1's interaction with nAChR subunits, particularly at the α4:α4 interface. In this study, we used molecular modeling and simulation to study the structure, dynamics, and interactions of lynx1 when bound to nAChRs, as well as unbound, monomeric lynx1 in membranes. Though lynx1 structures are similar in both states, its dynamics is more restricted in the bound state than in the unbound one. When bound, interactions between lynx1 and nAChR are observed to be maintained throughout the simulations. Of particular note, lynx1 demonstrates prolonged interactions with the receptor C-loop in one of the nAChR α4 subunits, a region important for agonist binding and possibly the transition between open/closed states. During interactions with lynx1, an α4 C-loop tends to be restricted in either a closed or open state, whereas the C-loop state transitions are more evident when lynx1 is unbound. Interestingly, the conformational change of the C-loop is stochastic, suggesting that lynx1 can influence nAChR (critical for its multimodal action), for instance, by shifting its agonist sensitivity and recovery from desensitization.

Lynx Prototoxins: Roles of Endogenous Mammalian Neurotoxin-Like Proteins in Modulating Nicotinic Acetylcholine Receptor Function to Influence Complex Biological Processes.

The cholinergic system modulates many biological functions, due to the widespread distribution of cholinergic neuronal terminals, and the diffuse release of its neurotransmitter, acetylcholine. Several layers of regulation help to refine and control the scope of this excitatory neurotransmitter system. One such regulatory mechanism is imparted through endogenous toxin-like proteins, prototoxins, which largely control the function of nicotinic receptors of the cholinergic system. Prototoxins and neurotoxins share the distinct three finger toxin fold, highly effective as a receptor binding protein, and the former are expressed in the mammalian brain, immune system, epithelium, etc. Prototoxins and elapid snake neurotoxins appear to be related through gene duplication and divergence from a common ancestral gene. Protein modulators can provide a graded response of the cholinergic system, and within the brain, stabilize neural circuitry through direct interaction with nicotinic receptors. Understanding the roles of each prototoxin (e.g., lynx1, lynx2/lypd1, PSCA, SLURP1, SLURP2, Lypd6, lypd6b, lypdg6e, PATE-M, PATE-B, etc.), their binding specificity and unique expression profile, has the potential to uncover many fascinating cholinergic-dependent mechanisms in the brain. Each family member can provide a spatially restricted level of control over nAChR function based on its expression in the brain. Due to the difficulty in the pharmacological targeting of nicotinic receptors in the brain as a result of widespread expression patterns and similarities in receptor sequences, unique interfaces between prototoxin and nicotinic receptor could provide more specific targeting than nicotinic receptors alone. As such, this family is intriguing from a long-term therapeutic perspective.

Augmenting the antinociceptive effects of nicotinic acetylcholine receptor activity through lynx1 modulation.

Neuronal nicotinic acetylcholine receptors (nAChRs) of the cholinergic system have been linked to antinociception, and therefore could be an alternative target for pain alleviation. nAChR activity has been shown to be regulated by the nicotinic modulator, lynx1, which forms stable complexes with nAChRs and has a negative allosteric action on their function. The objective in this study was to investigate the contribution of lynx1 to nicotine-mediated antinociception. Lynx1 contribution was investigated by mRNA expression analysis and electrophysiological responses to nicotine in the dorsal raphe nucleus (DRN), a part of the pain signaling pathway. In vivo antinociception was investigated in a test of nociception, the hot-plate analgesia assay with behavioral pharmacology. Lynx1/α4ß2 nAChR interactions were investigated using molecular dynamics computational modeling. Nicotine evoked responses in serotonergic and GABAergic neurons in the DRN are augmented in slices lacking lynx1 (lynx1KO). The antinociceptive effect of nicotine and epibatidine is enhanced in lynx1KO mice and blocked by mecamylamine and DHßE. Computer simulations predict preferential binding affinity of lynx1 to the α:α interface that exists in the stoichiometry of the low sensitivity (α4)3(ß2)2 nAChRs. Taken together, these data point to a role of lynx1 in mediating pain signaling in the DRN through preferential affinity to the low sensitivity α4ß2 nAChRs. This study suggests that lynx1 is a possible alternative avenue for nociceptive modulation outside of opioid-based strategies.

Medication adherence among adults prescribed imatinib, dasatinib, or nilotinib for the treatment of chronic myeloid leukemia.

BACKGROUND: Oral tyrosine kinase inhibitors are the standard of care for chronic myeloid leukemia. Tyrosine kinase inhibitors are administered in an outpatient setting for an indefinite period which may negatively impact adherence. Non-adherence to tyrosine kinase inhibitors is associated with disease progression. OBJECTIVES: To evaluate the need for adherence-enhancing interventions, this study was designed to determine the proportion of chronic myeloid leukemia patients non-adherent to their tyrosine kinase inhibitor regimen. The secondary objective was to identify the influence of patient characteristics on tyrosine kinase inhibitor adherence. METHODS: Cross-sectional retrospective chart and dispensing record reviews were performed to identify patients receiving a tyrosine kinase inhibitor from 1 June 2010 to 31 January 2012. Adherence was evaluated using the medication possession ratio. RESULTS: A total of 124 patients were included. Thirty-eight (31%) patients were non-adherent to their tyrosine kinase inhibitor regimen. Patients not receiving concurrent medications were more likely to be non-adherent (odds ratio (OR) 2.33, 95% confidence interval (CI) 1.05-5.13, p=0.04). The median medication possession ratio was 0.95 (IQR=0.83-1.07). Median medication possession ratio was lower in patients receiving imatinib compared to dasatinib or nilotinib (0.95 vs. 1.00, p=0.01) and in those less than 50 years old compared to those greater than 50 years old (0.92 vs. 0.97, p=0.02). CONCLUSIONS: Optimal tyrosine kinase inhibitor adherence in chronic myeloid leukemia patients poses a significant obstacle in achieving best possible outcomes while reducing healthcare costs. In this study, one in three chronic myeloid leukemia patients treated with a tyrosine kinase inhibitor were non-adherent to their regimen. Those at higher risk of non-adherence were on no concurrent medications, less than 50 years old, and those treated with imatinib. Active intervention to improve tyrosine kinase inhibitor adherence should be developed, implemented, and evaluated to improve patient outcomes at our center.