Lateral Motion of β Receptors in Membranes of Cultured Liver Cells

We have studied the lateral mobility and distribution of β receptors on Chang human liver cells by fluorescence photobleaching recovery and video intensification microscopy. The β receptors were labeled with the fluorescent antagonist 7-(2-allylphenoxy)-2,2-dimethyl-6-hydroxy- 1-(4-nitrobenzo-2-oxa-1,3-diazolyl)-1,4-diazaheptane (Alp-NBD). Sixty to 75% of the staining was specific (displaceable by unlabeled antagonists). Most of the antagonist-occupied β receptors were immobile, because only 15-25% of their fluorescence recovered on the experimental time scale at 23 degrees C. This immobility correlates with the clustered distribution of Alp-NBD-β -receptor complexes at 4 degrees C and 37 degrees C. The β receptors appear to be aggregated prior to antagonist binding, because visible patches were observed immediately after labeling for 30 sec at 4 degrees C. Preincubation at 37 degrees C with (-)-isoproterenol, a β agonist, prior to Alp-NBD labeling induced a time-dependent release of the β receptors to a more homogeneous distribution and increased the mobile fraction to 70-80% (lateral diffusion coefficient = 1.4× 10-9cm2/sec at 23 degrees C). This is not due to an effect on membrane fluidity, because the diffusion coefficient of a lipid probe was not altered. The time course of agonist-induced β -receptor mobilization correlates with receptor loss and adenylate cyclase desensitization but is much slower than adenylate cyclase activation. This indicates that adenylate cyclase activation by β receptors does not require macroscopic lateral mobility of the majority of the β receptors.