Description: Salmeterol is a long-acting β 2-adrenergic receptor ( β 2AR) agonist that is widely used as a bronchodilator for the treatment of persistent asthma and chronic obstructive pulmonary disease in conjunction with steroids. Previous studies demonstrated that salmeterol showed weak efficacy for activation of adenylyl cyclase; however, its efficacy in the complex desensitization of the β 2AR remains poorly understood. In this work, we provide insights into the roles played by the G protein–coupled receptor kinase/arrestin and protein kinase A in salmeterol-mediated desensitization through bioluminescence resonance energy transfer (BRET) studies of liganded- β 2AR binding to arrestin and through kinetic studies of cAMP turnover. First, BRET demonstrated a much reduced efficacy for salmeterol recruitment of arrestin to β 2AR relative to isoproterenol. The ratio of BRET ISO /BRET SALM after 5-minute stimulation was 20 and decreased to 5 after 35 minutes, reflecting a progressive decline in BRET ISO and a stable BRET SALM . Second, to assess salmeterol efficacy for functional desensitization, we examined the kinetics of salmeterol-induced cAMP accumulation (0–30 minutes) in human airway smooth muscle cells in the presence and absence of phosphodiesterase inhibition. Analysis of shaping of cAMP turnover for both agonists demonstrated significant salmeterol desensitization, although it was reduced relative to isoproterenol. Using an isoproterenol rescue protocol after either short-term (10 minutes) or long-term (2 and 14 hours) salmeterol pretreatments, we found that salmeterol progressively depressed isoproterenol stimulation but did not prevent subsequent rescue by isoproterenol and additional isoproterenol-mediated desensitization. Our findings reveal a complex efficacy for functional desensitization, demonstrating that although salmeterol shows weak efficacy for adenylyl cyclase activation and G protein–coupled receptor kinase/arrestin-mediated desensitization, it acts as a strong agonist in highly amplified protein kinase A–mediated events.

Description: Designer receptors exclusively activated by a designer drug (DREADDs) are clozapine-N-oxide-sensitive designer G protein-coupled receptors (GPCRs) that have emerged as powerful novel chemogenetic tools to study the physiological relevance of GPCR signaling pathways in specific cell types or tissues. Like endogenous GPCRs, clozapine-N-oxide-activated DREADDs do not only activate heterotrimeric G proteins but can also trigger β-arrestin-dependent (G protein-independent) signaling. To dissect the relative physiological relevance of G protein-mediated versus β-arrestin-mediated signaling in different cell types or physiological processes, the availability of G protein- and β-arrestin-biased DREADDs would be highly desirable. In this study, we report the development of a mutationally modified version of a non-biased DREADD derived from the M3 muscarinic receptor that can activate Gq/11 with high efficacy but lacks the ability to interact with β-arrestins. We also demonstrate that this novel DREADD is active in vivo and that cell type-selective expression of this new designer receptor can provide novel insights into the physiological roles of G protein (Gq/11)-dependent versus β-arrestin-dependent signaling in hepatocytes. Thus, this novel Gq/11-biased DREADD represents a powerful new tool to study the physiological relevance of Gq/11-dependent signaling in distinct tissues and cell types, in the absence of β-arrestin-mediated cellular effects. Such studies should guide the development of novel classes of functionally biased ligands that show high efficacy in various pathophysiological conditions but display a reduced incidence of side effects.

Description: Adipose tissue constitutes an extremely active endocrine organ with a network of signaling pathways enabling the organism to adapt to a wide range of different metabolic challenges, such as starvation, stress, infection, and short periods of gross energy excess. The functional pleiotropism of adipose tissue relies on its ability to synthesize and release a huge variety of hormones, cytokines, complement and growth factors, extracellular matrix proteins, and vasoactive factors, collectively termed adipokines. Obesity is associated with adipose tissue dysfunction leading to the onset of several pathologies including type 2 diabetes, dyslipidemia, nonalcoholic fatty liver, or hypertension, among others. The mechanisms underlying the development of obesity and its associated comorbidities include the hypertrophy and/or hyperplasia of adipocytes, adipose tissue inflammation, impaired extracellular matrix remodeling, and fibrosis together with an altered secretion of adipokines. Recently, the potential role of brown and beige adipose tissue in the protection against obesity has been also recognized. In contrast to white adipocytes, which store energy in the form of fat, brown and beige fat cells display energy-dissipating capacity through the promotion of triacylglycerol clearance, glucose disposal, and generation of heat for thermogenesis. Identification of the morphological and molecular changes in white, beige, and brown adipose tissue during weight gain is of utmost relevance for the identification of pharmacological targets for the treatment of obesity and its associated metabolic diseases.

Description: Temperature and light are important factors affecting production in the aquaculture industry, as they can drive behaviour and physiological responses of free‐swimming larval stages. However, the influence of light on crustacean farming has received little consideration. The common spider crab Maja brachydactyla Balss, 1922 has a great potential for aquaculture because of the easy maintenance, high fecundity, and short larval development. In order to optimize larval culture techniques, we quantified the influence of temperature and light on larval survival, development, and elemental (carbon and nitrogen) body composition. Constant darkness resulted in longer developmental time as compared with daily light photoperiod (6–16 light hours). Larvae reared under constant darkness showed also reduced dry mass, carbon and nitrogen content, and C:N ratio. We also found carry‐over effects of light conditions: constant darkness experienced during the zoeal stage led to increased developmental time in the megalopa stage. Temperature and light showed additive effects. We optimized the larval culture of M. brachydactyla requiring around 14 days from hatch to first juvenile by keeping cultures under 21 ± 1°C and light sources simulating the daily light cycle.