Label-free dynamic mass redistribution analysis of endogenous adrenergic receptor signaling in primary preadipocytes and differentiated adipocytes.

INTRODUCTION: Adipose tissues release adipokines, which regulate energy intake and expenditure. G protein-coupled receptors (GPCRs) and associated signaling pathways in adipocytes are potentially important drug targets for conditions with disturbed energy metabolism.

METHODS: The aim of the current study was to compare signaling of endogenously expressed GPCRs between primary preadipocytes and differentiated adipocytes using a novel state-of-the-art unbiased method that measures dynamic mass redistribution (DMR) in real-time. Adrenergic agonists were chosen since they control adipocyte functions such as lipolysis and glyconeogenesis.

RESULTS: Isoprenaline (ISO) and phenylephrine (PE) elicited concentration-dependent responses in preadipocytes and differentiated adipocytes. The effect of ISO was cholera toxin (CTX)-sensitive, indicating it is Gs -dependent. The effect could also be blocked by propranolol proving the signal is mediated through β-adrenergic receptors. The signaling resulting from PE stimulation was completely abolished by a Gq -selective inhibitor and CTX in preadipocytes but surprisingly became CTX-sensitive in differentiated adipocytes. The use of prazosin and propranolol revealed that the PE-response in differentiated adipocytes had a β-adrenergic receptor component to it. In addition, we tested the bone-derived peptide osteocalcin, which did not result in DMR changes in preadipocytes or differentiated adipocytes.

DISCUSSION: In conclusion, this study for the first time demonstrates that DMR assays can be used to assess signaling in differentiated adipocytes. This platform can serve as a tool for future drug screening in primary adipocytes. Furthermore, this study illustrates that PE-induced effects on adipocytes vary by developmental stage and are not as selective as originally thought.