Muscle acetylcholine receptor conversion into chloride conductance at positive potentials by a single mutation.

Charge selectivity forms the basis of cellular excitation or inhibition by Cys-loop ligand-gated ion channels (LGICs), and is essential for physiological receptor function. There are no reports of naturally occurring mutations in LGICs associated with the conversion of charge selectivity. Here, we report on a CHRNA1 mutation (α1Leu251Arg) in a patient with congenital myasthenic syndrome associated with transformation of the muscle acetylcholine receptor (AChR) into an inhibitory channel. Performing patch-clamp experiments, the AChR was found to be converted into chloride conductance at positive potentials, whereas whole-cell currents at negative potentials, although markedly reduced, were still carried by sodium. Umbrella sampling molecular dynamics simulations revealed constriction of the channel pore radius to 2.4 Å as a result of the mutation, which required partial desolvation of the ions in order to permeate the pore. Ion desolvation was associated with an energetic penalty that was compensated for by the favorable electrostatic interaction of the positively charged arginines with chloride. These findings reveal a mechanism for the transformation of the muscle AChR into an inhibitory channel in a clinical context.

Structure of human CALHM1 reveals key locations for channel regulation and blockade by ruthenium red.

Calcium homeostasis modulator 1 (CALHM1) is a voltage-dependent channel involved in neuromodulation and gustatory signaling. Despite recent progress in the structural biology of CALHM1, insights into functional regulation, pore architecture, and channel blockade remain limited. Here we present the cryo-EM structure of human CALHM1, revealing an octameric assembly pattern similar to the non-mammalian CALHM1s and the lipid-binding pocket conserved across species. We demonstrate by MD simulations that this pocket preferentially binds a phospholipid over cholesterol to stabilize its structure and regulate the channel activities. Finally, we show that residues in the amino-terminal helix form the channel pore that ruthenium red binds and blocks.

Structural insights into binding of therapeutic channel blockers in NMDA receptors.

Excitatory signaling mediated by N-methyl-D-aspartate receptor (NMDAR) is critical for brain development and function, as well as for neurological diseases and disorders. Channel blockers of NMDARs are of medical interest owing to their potential for treating depression, Alzheimer's disease, and epilepsy. However, precise mechanisms underlying binding and channel blockade have remained limited owing to challenges in obtaining high-resolution structures at the binding site within the transmembrane domains. Here, we monitor the binding of three clinically important channel blockers: phencyclidine, ketamine, and memantine in GluN1-2B NMDARs at local resolutions of 2.5-3.5 Å around the binding site using single-particle electron cryo-microscopy, molecular dynamics simulations, and electrophysiology. The channel blockers form different extents of interactions with the pore-lining residues, which control mostly off-speeds but not on-speeds. Our comparative analyses of the three unique NMDAR channel blockers provide a blueprint for developing therapeutic compounds with minimal side effects.

Molecular determinants of binding of non-oxime bispyridinium nerve agent antidote compounds to the adult muscle nAChR.

Organophosphorus nerve agents (NAs) are the most lethal chemical warfare agents and have been used by state and non-state actors since their discovery in the 1930s. They covalently modify acetylcholinesterase, preventing the breakdown of acetylcholine (ACh) with subsequent loss of synaptic transmission, which can result in death. Despite the availability of several antidotes for OPNA exposure, none directly targets the nicotinic acetylcholine receptor (nAChR) mediated component of toxicity. Non-oxime bispyridinium compounds (BPDs) have been shown previously to partially counteract the effects of NAs at skeletal muscle tissue, and this has been attributed to inhibition of the muscle nAChR. Functional data indicate that, by increasing the length of the alkyl linker between the pyridinium moieties of BPDs, the antagonistic activity at nAChRs can be improved. Molecular dynamics simulations of the adult muscle nAChR in the presence of BPDs identified key residues likely to be involved in binding. Subsequent two-electrode voltage clamp recordings showed that one of the residues, εY131, acts as an allosteric determinant of BPD binding, and that longer BPDs have a greater stabilizing effect on the orthosteric loop C than shorter ones. The work reported will inform future design work on novel antidotes for treating NA exposure.

Novel assessment of leukocyte-rich platelet-rich plasma on functional and patient-reported outcomes in knee osteoarthritis: a pilot study.

Background: Leukocyte-rich platelet-rich plasma (LR-PRP) has demonstrated to be beneficial for patients with knee osteoarthritis (KOA); however, reliable objective end points to accurately assess its therapeutic effects is lacking. Aim: To investigate the efficacy of LR-PRP as assessed by functional and patient-reported outcomes at early time points (6 weeks). Materials & methods: We conducted a prospective cohort study in 12 patients with diagnosed KOA (Kellgren Lawrence score of II-III), who underwent a single ultrasound-guided LR-PRP injection. Results: There was significant improvement in timed up and go, pain and quality of life scales and balance parameters. There were nonsignificant improvements in range of motion and gait parameters. Conclusion: LR-PRP demonstrates efficacy in meaningful end points for functional and patient-reported outcomes at early time points in patients with KOA.

Dynamic Ultrasound Uniquely Reveals Anterior Capsular Tear Communicating With Vastus Intermedius Fibers.

We present a case of an anterior capsular tear of the right knee in a previously healthy, active individual. The patient was a 31-year-old male seen one month after the onset of throbbing anterior right knee pain, which progressed to sharp suprapatellar pain over the next three months. Both dynamic ultrasound (US) and MRI were obtained. Dynamic US revealed a tear of the anterior capsule of the right knee complicated by a suprapatellar effusion communicating into the vastus intermedius fibers of the quadriceps tendon. However, these findings were not evident on MRI. In addition to discussing this unique pathology, we highlight the utility of both standard and dynamic US in establishing diagnoses for capsular pathologies.

Ehlers-Danlos Syndrome: An Analysis of the Current Treatment Options.

BACKGROUND: Ehlers-Danlos syndrome (EDS) is a multifaceted disease that can present with a variety of types of pain. Unfortunately, both the mechanisms and treatments for pain are poorly understood. The proposed treatments for the various musculoskeletal pain syndromes in EDS have had variable success, and it becomes much more imperative to better define and evaluate the current treatment modalities in treating this debilitating disease. OBJECTIVES: The purpose of this study was to investigate the currently available treatment modalities for patients with EDS and their efficacies in pain and symptom relief. STUDY DESIGN: Retrospective cohort study. SETTING: Institutional physical medicine and rehabilitation primary care clinic. METHODS: All patients were seen between January 2015 and April 2019, in which 98 patients with EDS were identified through retrospective chart review. Institutional review board approval was obtained, and all patients provided written consent to be included in the study. We reviewed various treatment modalities, including complimentary/alternative treatments, opioids/opioid-like medications, nonsteroidal antiinflammatory drugs, physical therapy, occupational therapy, muscle relaxants, neuropathic modulators, steroids, surgery/procedures, and acetaminophen. Treatment methods were extracted from individual patient charts, and efficacy was grouped into 3 categories: improvement, no effect, or worsened symptoms. RESULTS: The most common treatments used were complimentary/alternative treatments (n = 88). Occupational therapy and bracing were the most effective options with 70% of patients reporting improvement. Neuropathic modulators were the least well tolerated with 47% of patients reporting adverse effects. LIMITATIONS: Men were a small percentage of the study. Patients were not randomized, and pain score reporting was subjective. Patient data were extracted from a single practice setting. Timing and symptom onset were not measured. CONCLUSIONS: There is a relative paucity of published literature regarding the various treatment methods for EDS. Although our study is able to identify positive and negative trends with certain modalities, it is vital to understand that EDS is not a uniform diagnosis among patients, and that a combination of several different treatments usually is needed for optimal symptom control. Further research and investigation are necessary to develop a comprehensive treatment database for this complex condition. KEY WORDS: Ehlers-Danlos syndrome, pain, hypermobility, arthralgia, subluxation, genetic, physical therapy, interventional pain.