An autoradiographic analysis of beta adrenergic receptors on immunocytochemically defined astroglia
Mccarthy, K.D.
Journal of Pharmacology and Experimental Therapeutics 226(1): 282-290
1983
ISSN/ISBN: 0022-3565 PMID: 6306225 Document Number: 221061
Previous studies demonstrated that cultured astroglia exhibit a variety of receptor systems which regulate cAMP accumulation. The method described was developed to examine whether such receptors are present on cultured rat astroglia regardless of their morphological features or intercellular contacts. Primary cultures containing predominantly polygonal astroglia and a few multipolar astroglia were fixed and permeated to permit the intracellular staining of the astroglial cell marker, glial fibrillary acidic protein (GFAP). The cells were then incubated in medium containing antiserum to GFAP, rinsed and incubated in medium containing rhodamine-conjugated goat antirabbit antiserum and the .beta.-adrenergic receptor antagonist (-)-[125I]iodopindolol ([125I]PIN). The cells were then rinsed, frozen, lyophilized and opposed to emulsion-coated coverslips. After 10-60 days, the autoradiograms were developed and the grains were visualized by reflected polarized light microscopy and GFAP-positive cells by fluorescence microscopy. Results from binding assays indicated that these methods did not alter the following: the specificity of GFAP antiserum for astroglia; the kinetics of [125I]PIN binding to astroglia; the number of [125I]PIN binding sites per cell; the stereospecific displacement of [125I]PIN by propranolol. One effect of the fixation-permeation procedure was an increase (10- to 50-fold) in the potency of both isomers of isoproterenol to displace [125I]PIN. This effect may be due to the ability of an agonist, such as isoproterenol, to induce a rapid decrease in the affinity of .beta.-adrenergic receptors for agonists on living but not fixed-permeated cells. There evidently are at least 2 populations of morphologically distinct GFAP positive astroglia which can also be distinguished via their expression of .beta.-adrenergic receptors.