Abstract
Seven-transmembrane receptors mediate diverse skeletal muscle responses for a wide variety of stimuli, via activation of heterotrimeric G-proteins. Herein we evaluate the expression and activation of rat diaphragm or cultured skeletal muscle G-proteins using [(35)S]GTPγS. Total membrane Gα subunit content was 4-7 times higher in rat primary cultured myotubes and L6 cell line than in diaphragm (32.6±1.2fmol/mg protein) and 7-27% of them were in the active conformational state. Immunoprecipitation assay showed equal expression of diaphragm Gαs, Gαq and Gαi/o. Addition of GDP allowed the measurement of G-protein activation by different GPCR, including adrenoceptor, adenosine, melatonin and muscarinic receptors. Diaphragm denervation resulted in a marked increase in both total and active state G-protein levels. Together, the results show that [(35)S]GTPγS binding assay is a sensitive and valuable method to evaluate GPCR activity in skeletal muscle cells, which is of particular interest for pharmacological analysis of drugs with potential use in the management of respiratory muscle failure.
Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
