Engineering, applications, and future perspectives of GPCR-based genetically encoded fluorescent indicators for neuromodulators.

This review explores the evolving landscape of G-protein-coupled receptor (GPCR)-based genetically encoded fluorescent indicators (GEFIs), with a focus on their development, structural components, engineering strategies, and applications. We highlight the unique features of this indicator class, emphasizing the importance of both the sensing domain (GPCR structure and activation mechanism) and the reporting domain (circularly permuted fluorescent protein (cpFP) structure and fluorescence modulation). Further, we discuss indicator engineering approaches, including the selection of suitable cpFPs and expression systems. Additionally, we showcase the diversity and flexibility of their application by presenting a summary of studies where such indicators were used. Along with all the advantages, we also focus on the current limitations as well as common misconceptions that arise when using these indicators. Finally, we discuss future directions in indicator engineering, including strategies for screening with increased throughput, optimization of the ligand-binding properties, structural insights, and spectral diversity.

Innovative moments in low-intensity, telephone-based cognitive-behavioral therapy for depression.

Background: Innovative moments (IMs), defined as moments in psychotherapy when patients' problematic patterns change toward more elaborated and adaptive patterns, have been shown to be associated with a clinical change in patients with depression. Thus, far IMs have been studied in face-to-face settings but not in telephone-based cognitive-behavioral therapy (t-CBT). This study investigates whether IMs occur in t-CBT and examines the association between IMs and symptom improvement, and reconceptualization and symptom improvement. Methods: The therapy transcripts of n = 10 patients with mild to moderate depression (range: 7-11 sessions, in total 94 sessions) undergoing t-CBT were qualitatively and quantitatively analyzed. Symptom severity (Patient Health Questionnaire-9) and IMs (levels and proportions) were assessed for each therapy session. Hierarchical linear models were used to test the prediction models. Results: The rating of IMs was shown to be feasible and reliable using the Innovative Moments Coding System (IMCS) (84.04% agreement in words coded), which is indicative of the applicability of the concept of IMs in t-CBT. Only reconceptualization IMs were shown to have a predictive value for treatment success (R2 = 0.05, p = 0.01). Discussion: The results should be interpreted with caution due to the exploratory nature of this study. Due to the telephone setting, it was necessary to adapt the IMCS. Nonetheless, the extent of IMs identified in the low-intensity t-CBT investigated was comparable to IMs in face-to-face therapy. Further studies are needed to clarify the association between IMs and treatment success as a change process, especially for low-intensity treatments.

Sensitive multicolor indicators for monitoring norepinephrine in vivo.

Genetically encoded indicators engineered from G-protein-coupled receptors are important tools that enable high-resolution in vivo neuromodulator imaging. Here, we introduce a family of sensitive multicolor norepinephrine (NE) indicators, which includes nLightG (green) and nLightR (red). These tools report endogenous NE release in vitro, ex vivo and in vivo with improved sensitivity, ligand selectivity and kinetics, as well as a distinct pharmacological profile compared with previous state-of-the-art GRABNE indicators. Using in vivo multisite fiber photometry recordings of nLightG, we could simultaneously monitor optogenetically evoked NE release in the mouse locus coeruleus and hippocampus. Two-photon imaging of nLightG revealed locomotion and reward-related NE transients in the dorsal CA1 area of the hippocampus. Thus, the sensitive NE indicators introduced here represent an important addition to the current repertoire of indicators and provide the means for a thorough investigation of the NE system.