G-protein-coupled receptor signaling components localize in both sarcolemmal and intracellular caveolin-3-associated microdomains in adult cardiac myocytes

J Biol Chem. 2005 Sep 2;280(35):31036-44. doi: 10.1074/jbc.M502540200. Epub 2005 Jun 16.


This study tests the hypothesis that G-protein-coupled receptor (GPCR) signaling components involved in the regulation of adenylyl cyclase (AC) localize with caveolin (Cav), a protein marker for caveolae, in both cell-surface and intracellular membrane regions. Using sucrose density fractionation of adult cardiac myocytes, we detected Cav-3 in both buoyant membrane fractions (BF) and heavy/non-buoyant fractions (HF); beta2-adrenergic receptors (AR) in BF; and AC5/6, beta1-AR, M4-muscarinic acetylcholine receptors (mAChR), mu-opioid receptors, and Galpha(s) in both BF and HF. In contrast, M2-mAChR, Galpha(i3), and Galpha(i2) were found only in HF. Immunofluorescence microscopy showed co-localization of Cav-3 with AC5/6, Galpha(s), beta2-AR, and mu-opioid receptors in both sarcolemmal and intracellular membranes, whereas M2-mAChR were detected only intracellularly. Immunofluorescence of adult heart revealed a distribution of Cav-3 identical to that in isolated adult cardiac myocytes. Upon immunoelectron microscopy, Cav-3 co-localized with AC5/6 and Galpha(s) in sarcolemmal and intracellular vesicles, the latter closely allied with T-tubules. Cav-3 immunoprecipitates possessed components that were necessary and sufficient for GPCR agonist-promoted stimulation and inhibition of cAMP formation. The distribution of GPCR, G-proteins, and AC with Cav-3 in both sarcolemmal and intracellular T-tubule-associated regions indicates the existence of multiple Cav-3-localized cellular microdomains for signaling by hormones and drugs in the heart.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Adenylyl Cyclases / metabolism
  • Animals
  • Biomarkers
  • Caveolin 3
  • Caveolins / metabolism*
  • Cell Fractionation
  • Cells, Cultured
  • Cyclic AMP / metabolism
  • Fibroblasts / cytology
  • Fibroblasts / metabolism
  • GTP-Binding Protein alpha Subunits / metabolism
  • Humans
  • Intracellular Membranes / chemistry
  • Intracellular Membranes / metabolism*
  • Intracellular Membranes / ultrastructure
  • Isoenzymes / metabolism
  • Male
  • Muscle, Smooth, Vascular / cytology
  • Myocytes, Cardiac* / cytology
  • Myocytes, Cardiac* / metabolism
  • Rats
  • Rats, Sprague-Dawley
  • Receptor, Muscarinic M2 / metabolism
  • Receptors, Adrenergic, beta / metabolism
  • Receptors, G-Protein-Coupled / genetics
  • Receptors, G-Protein-Coupled / metabolism*
  • Receptors, Muscarinic / metabolism
  • Receptors, Opioid / metabolism
  • Sarcolemma / chemistry
  • Sarcolemma / metabolism*
  • Sarcolemma / ultrastructure
  • Signal Transduction / physiology*


  • Biomarkers
  • Cav3 protein, rat
  • Caveolin 3
  • Caveolins
  • GTP-Binding Protein alpha Subunits
  • Isoenzymes
  • Receptor, Muscarinic M2
  • Receptors, Adrenergic, beta
  • Receptors, G-Protein-Coupled
  • Receptors, Muscarinic
  • Receptors, Opioid
  • Cyclic AMP
  • Adenylyl Cyclases