LETTER TO THE EDITOR

n/

Human α6β4 nicotinic acetylcholine receptor: heterologous expression and agonist behavior provide insights into the immediate binding site.

Study of α6β4 nicotinic acetylcholine receptors (nAChRs) as a pharmacological target has recently gained interest because of their involvement in analgesia, control of catecholamine secretion, dopaminergic pathways, and aversive pathways. However, an extensive characterization of the human α6β4 nAChRs has been vitiated by technical difficulties resulting in poor receptor expression. In 2020, Knowland and collaborators identified BARP (β-anchoring and regulatory protein), a previously known voltage-gated calcium channel suppressor, as a novel human α6β4 chaperone. Here we establish that co-expression of human BARP with human α6β4 in Xenopus oocytes, resulted in the functional expression of human α6β4 receptors with ACh-elicited currents that allow an in-depth characterization of the receptor using two electrode voltage-clamp electrophysiology together with diverse agonists and receptor mutations. We report: (1) an extended pharmacological characterization of the receptor, and (2) key residues for agonist-activity located in or near the first shell of the binding pocket. Significance Statement The human α6β4 nAChR has attained increased interest because of its involvement in diverse physiological processes and diseases. Although recognized as a pharmacological target, development of specific agonists has been hampered by limited knowledge of its structural characteristics and by challenges in expressing the receptor. By including the chaperone BARP for enhanced expression and employing different ligands, we have studied the pharmacology of α6β4, providing insight into receptor residues and structural requirements for ligands important to consider for agonist-induced activation

Human α6β4 nicotinic acetylcholine receptor: heterologous expression and agonist behavior provide insights into the immediate binding site

Study of α6β4 nicotinic acetylcholine receptors (nAChRs) as a pharmacological target has recently gained interest because of their involvement in analgesia, control of catecholamine secretion, dopaminergic pathways, and aversive pathways. However, an extensive characterization of the human α6β4 nAChRs has been vitiated by technical difficulties resulting in poor receptor expression. In 2020, Knowland and collaborators identified BARP (β-anchoring and regulatory protein), a previously known voltage-gated calcium channel suppressor, as a novel human α6β4 chaperone. Here, we establish that co-expression of human BARP with human α6β4 in Xenopus oocytes, resulted in the functional expression of human α6β4 receptors with acetylcholine-elicited currents that allow an in-depth characterization of the receptor using two electrode voltage-clamp electrophysiology together with diverse agonists and receptor mutations. We report: 1) an extended pharmacological characterization of the receptor, and 2) key residues for agonist-activity located in or near the first shell of the binding pocket. SIGNIFICANCE STATEMENT: The human α6β4 nicotinic acetylcholine receptor has attained increased interest because of its involvement in diverse physiological processes and diseases. Although recognized as a pharmacological target, development of specific agonists has been hampered by limited knowledge of its structural characteristics and by challenges in expressing the receptor. By including the chaperone β-anchoring and regulatory protein for enhanced expression and employing different ligands, we have studied the pharmacology of α6β4, providing insight into receptor residues and structural requirements for ligands important to consider for agonist-induced activation

Novel positive allosteric modulators of GABAA receptors with anesthetic activity.

GABAA receptors are the main inhibitory neurotransmitter receptors in the brain and are targets for numerous clinically important drugs such as benzodiazepines, anxiolytics and anesthetics. We previously identified novel ligands of the classical benzodiazepine binding pocket in α1β2γ2 GABAA receptors using an experiment-guided virtual screening (EGVS) method. This screen also identified novel ligands for intramembrane low affinity diazepam site(s). In the current study we have further characterized compounds 31 and 132 identified with EGVS as well as 4-O-methylhonokiol. We investigated the site of action of these compounds in α1β2γ2 GABAA receptors expressed in Xenopus laevis oocytes using voltage-clamp electrophysiology combined with a benzodiazepine site antagonist and transmembrane domain mutations. All three compounds act mainly through the two β+/α- subunit transmembrane interfaces of the GABAA receptors. We then used concatenated receptors to dissect the involvement of individual β+/α- interfaces. We further demonstrated that these compounds have anesthetic activity in a small aquatic animal model, Xenopus laevis tadpoles. The newly identified compounds may serve as scaffolds for the development of novel anesthetics

Strategie di educazione a confronto. Progetto di miglioramento dell'aderenza al percorso riabilitativo rivolto a pazienti sottoposte a dissezione linfonodale ascellare

Introduzione: Le complicanze conseguenti al trattamento chirurgico del carcinoma mammario con dissezione ascellare possono, se non precocemente trattate, cronicizzare causando disabilità dell’arto superiore. Il percorso riabilitativo prevede: esecuzione autonoma di esercizi finalizzati al recupero dell’abilità motoria, attenersi a regole che riducono l’insorgenza del linfedema e saper riconoscere precocemente le complicanze, ovvero che la paziente partecipi attivamente al processo. Si ipotizza che modificando la strategia educativa che precede l’inizio di questo percorso autogestito, utilizzando tecniche di apprendimento attivo in gruppi educativi interattivi con proiezione di immagini, si possa influire sugli aspetti psicologico-motivazionali che influenzano la compliance. Metodo: Sono previste in questo studio randomizzato e controllato 200 pazienti sottoposte per la prima volta a dissezione ascellare assegnate a: educazione individuale/verbale standard (controllo) vs educazione in gruppo/immagini (intervento). Enpoint primario: incremento dell’aderenza della paziente al percorso riabilitativo autonomo. Endpoint secondari: funzionalità del braccio; costo/efficacia misurata come tempo educativo impiegato/disabilità residua (DASH); recupero articolare; comparsa linfedema; QoL; soddisfazione; dolore. Le variabili soggettive sono valutate con questionari. Sono previste 2 misurazioni degli endpoint: -basale; -a 3 mesi. Risultati preliminari: ad oggi 19 pazienti (10 gruppo/immagini; 9 individuale/verbale) hanno concluso lo studio. Non si e’ evidenziata alcuna differenza in termini di compliance. Tuttavia le pazienti del gruppo/immagini riportano: un tempo di educazione inferiore (15’ gruppo/immagini vs 20’ individuale/verbale; p=0.02), una minor assunzione di FANS/Analgesici [2(20%) pazienti gruppo/immagini vs 6(66.7%) individuale/verbale; p=0.04], e un miglior recupero funzionale da ritenersi clinicamente rilevante [delta DASH mediana(3m – T0) gruppo/immagini –2.1(IC95%: –17.5, 19.2) vs individuale/verbale 7.5 (IC95%: –21.7, 37.5); p=0.10]. Conclusioni: I risultati sono promettenti e, nonostante il numero limitato di pazienti, questa analisi preliminare ha già permesso di verificare la fattibilità dell’intervento suggerendo l’adeguatezza del programma educativo in “gruppo/immagini” perché, senza diminuire l'efficacia su tutte le variabili misurate rispetto all'intervento “individuale/verbale”, prevede un risparmio di risorse

Do Early Breast Cancer Patients with Only Sentinel Lymph Node Biopsy Experience Lower Psychological Morbidity Over Time?

n/