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The ?-arrestin-biased ?-adrenergic receptor blocker carvedilol enhances skeletal muscle contractility.

ABSTRACT: A decrease in skeletal muscle strength and functional exercise capacity due to aging, frailty, and muscle wasting poses major unmet clinical needs. These conditions are associated with numerous adverse clinical outcomes including falls, fractures, and increased hospitalization. Clenbuterol, a ?2-adrenergic receptor (?2AR) agonist enhances skeletal muscle strength and hypertrophy; however, its clinical utility is limited by side effects such as cardiac arrhythmias mediated by G protein signaling. We recently reported that clenbuterol-induced increases in contractility and skeletal muscle hypertrophy were lost in ?-arrestin 1 knockout mice, implying that arrestins, multifunctional adapter and signaling proteins, play a vital role in mediating the skeletal muscle effects of ?2AR agonists. Carvedilol, classically defined as a ?AR antagonist, is widely used for the treatment of chronic systolic heart failure and hypertension, and has been demonstrated to function as a ?-arrestin-biased ligand for the ?2AR, stimulating ?-arrestin-dependent but not G protein-dependent signaling. In this study, we investigated whether treatment with carvedilol could enhance skeletal muscle strength via ?-arrestin-dependent pathways. In a murine model, we demonstrate chronic treatment with carvedilol, but not other ?-blockers, indeed enhances contractile force in skeletal muscle and this is mediated by ?-arrestin 1. Interestingly, carvedilol enhanced skeletal muscle contractility despite a lack of effect on skeletal muscle hypertrophy. Our findings suggest a potential unique clinical role of carvedilol to stimulate skeletal muscle contractility while avoiding the adverse effects with ?AR agonists. This distinctive signaling profile could present an innovative approach to treating sarcopenia, frailty, and secondary muscle wasting.

SUBMITTER: Kim J

PROVIDER: S-EPMC7275696 | biostudies-literature | 2020 Jun

REPOSITORIES: biostudies-literature

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The β-arrestin-biased β-adrenergic receptor blocker carvedilol enhances skeletal muscle contractility.

Kim Jihee J Grotegut Chad A CA Wisler James W JW Mao Lan L Rosenberg Paul B PB Rockman Howard A HA Lefkowitz Robert J RJ

Proceedings of the National Academy of Sciences of the United States of America 20200515 22

A decrease in skeletal muscle strength and functional exercise capacity due to aging, frailty, and muscle wasting poses major unmet clinical needs. These conditions are associated with numerous adverse clinical outcomes including falls, fractures, and increased hospitalization. Clenbuterol, a β<sub>2</sub>-adrenergic receptor (β<sub>2</sub>AR) agonist enhances skeletal muscle strength and hypertrophy; however, its clinical utility is limited by side effects such as cardiac arrhythmias mediated b ...[more]

PMID: 32414934

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Project description:BACKGROUND:?2-adrenergic receptors (?2ARs) are the target of catecholamines and play fundamental roles in cardiovascular, pulmonary, and skeletal muscle physiology. An important action of ?2AR stimulation on skeletal muscle is anabolic growth, which has led to the use of agonists such as clenbuterol by athletes to enhance muscle performance. While previous work has demonstrated that ?2ARs can engage distinct signaling and functional cascades mediated by either G proteins or the multifunctional adaptor protein, ?-arrestin, the precise role of ?-arrestin in skeletal muscle physiology is not known. Here, we tested the hypothesis that agonist activation of the ?2AR by clenbuterol would engage ?-arrestin as a key transducer of anabolic skeletal muscle growth. METHODS:The contractile force of isolated extensor digitorum longus muscle (EDL) and calcium signaling in isolated flexor digitorum brevis (FDB) fibers were examined from the wild-type (WT) and ?-arrestin 1 knockout mice (?arr1KO) followed by chronic administration of clenbuterol (1?mg/kg/d). Hypertrophic responses including fiber composition and fiber size were examined by immunohistochemical imaging. We performed a targeted phosphoproteomic analysis on clenbuterol stimulated primary cultured myoblasts from WT and ?arr1KO mice. Statistical significance was determined by using a two-way analysis with Sidak's or Tukey's multiple comparison test and the Student's t test. RESULTS:Chronic administration of clenbuterol to WT mice enhanced the contractile force of EDL muscle and calcium signaling in isolated FDB fibers. In contrast, when administered to ?arr1KO mice, the effect of clenbuterol on contractile force and calcium influx was blunted. While clenbuterol-induced hypertrophic responses were observed in WT mice, this response was abrogated in mice lacking ?-arrestin 1. In primary cultured myoblasts, clenbuterol-stimulated phosphorylation of multiple pro-hypertrophy proteins required the presence of ?-arrestin 1. CONCLUSIONS:We have identified a previously unappreciated role for ?-arrestin 1 in mediating ?2AR-stimulated skeletal muscle growth and strength. We propose these findings could have important implications in the design of future pharmacologic agents aimed at reversing pathological conditions associated with skeletal muscle wasting.

Project description:Reperfusion as a therapeutic intervention for acute myocardial infarction-induced cardiac injury itself induces further cardiomyocyte death. ?-arrestin (?arr)-biased ?-adrenergic receptor (?AR) activation promotes survival signaling responses in vitro; thus, we hypothesize that this pathway can mitigate cardiomyocyte death at the time of reperfusion to better preserve function. However, a lack of efficacious ?arr-biased orthosteric small molecules has prevented investigation into whether this pathway relays protection against ischemic injury in vivo. We recently demonstrated that the pepducin ICL1-9, a small lipidated peptide fragment designed from the first intracellular loop of ?2AR, allosterically engaged pro-survival signaling cascades in a ?arr-dependent manner in vitro. Thus, in this study we tested whether ICL1-9 relays cardioprotection against ischemia/reperfusion (I/R)-induced injury in vivo. Methods: Wild-type (WT) C57BL/6, ?2AR knockout (KO), ?arr1KO and ?arr2KO mice received intracardiac injections of either ICL1-9 or a scrambled control pepducin (Scr) at the time of ischemia (30 min) followed by reperfusion for either 24 h, to assess infarct size and cardiomyocyte death, or 4 weeks, to monitor the impact of ICL1-9 on long-term cardiac structure and function. Neonatal rat ventricular myocytes (NRVM) were used to assess the impact of ICL1-9 versus Scr pepducin on cardiomyocyte survival and mitochondrial superoxide formation in response to either serum deprivation or hypoxia/reoxygenation (H/R) in vitro and to investigate the associated mechanism(s). Results: Intramyocardial injection of ICL1-9 at the time of I/R reduced infarct size, cardiomyocyte death and improved cardiac function in a ?2AR- and ?arr-dependent manner, which led to improved contractile function early and less fibrotic remodeling over time. Mechanistically, ICL1-9 attenuated mitochondrial superoxide production and promoted cardiomyocyte survival in a RhoA/ROCK-dependent manner. RhoA activation could be detected in cardiomyocytes and whole heart up to 24 h post-treatment, demonstrating the stability of ICL1-9 effects on ?arr-dependent ?2AR signaling. Conclusion: Pepducin-based allosteric modulation of ?arr-dependent ?2AR signaling represents a novel therapeutic approach to reduce reperfusion-induced cardiac injury and relay long-term cardiac remodeling benefits.

Project description:RATIONALE:MicroRNAs (miRs) are small, non-coding RNAs that function to post-transcriptionally regulate target genes. First transcribed as primary miR transcripts (pri-miRs), they are enzymatically processed by Drosha into premature miRs (pre-miRs) and further cleaved by Dicer into mature miRs. Initially discovered to desensitize ?-adrenergic receptor (?AR) signaling, ?-arrestins are now well-appreciated to modulate multiple pathways independent of G protein signaling, a concept known as biased signaling. Using the ?-arrestin-biased ?AR ligand carvedilol, we previously showed that ?-arrestin1 (not ?-arrestin2)-biased ?1AR (not ?2AR) cardioprotective signaling stimulates Drosha-mediated processing of six miRs by forming a multi-protein nuclear complex, which includes ?-arrestin1, the Drosha microprocessor complex and a single-stranded RNA binding protein hnRNPA1. OBJECTIVE:Here, we investigate whether ?-arrestin-mediated ?AR signaling induced by carvedilol could regulate Dicer-mediated miR maturation in the cytoplasm and whether this novel mechanism promotes cardioprotective signaling. METHODS AND RESULTS:In mouse hearts, carvedilol indeed upregulates three mature miRs, but not their pre-miRs and pri-miRs, in a ?-arrestin 1- or 2-dependent manner. Interestingly, carvedilol-mediated activation of miR-466g or miR-532-5p, and miR-674 is dependent on ?2ARs and ?1ARs, respectively. Mechanistically, ?-arrestin 1 or 2 regulates maturation of three newly identified ?AR/?-arrestin-responsive miRs (?-miRs) by associating with the Dicer maturation RNase III enzyme on three pre-miRs of ?-miRs. Myocardial cell approaches uncover that despite their distinct roles in different cell types, ?-miRs act as gatekeepers of cardiac cell functions by repressing deleterious targets. CONCLUSIONS:Our findings indicate a novel role for ?AR-mediated ?-arrestin signaling activated by carvedilol in Dicer-mediated miR maturation, which may be linked to its protective mechanisms.

Dataset Information

The ?-arrestin-biased ?-adrenergic receptor blocker carvedilol enhances skeletal muscle contractility.

Publications

The β-arrestin-biased β-adrenergic receptor blocker carvedilol enhances skeletal muscle contractility.

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