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Chronic adrenaline treatment fails to down‐regulate the Del 301–303 ‐α 2B ‐adrenoceptor in neuronal cells
Author(s) -
Salim S,
Desai AN,
Taneja M,
Eikenburg DC
Publication year - 2009
Publication title -
british journal of pharmacology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.432
H-Index - 211
eISSN - 1476-5381
pISSN - 0007-1188
DOI - 10.1111/j.1476-5381.2009.00294.x
Subject(s) - adrenergic receptor , neuroscience , medicine , endocrinology , receptor , biology
Background and purpose: A polymorphism of the human α 2B ‐adrenoceptor (Del 301–303 ‐α 2B ‐adrenoceptor) has been described, and this receptor exhibits reduced G‐protein‐coupled receptor kinase (GRK) phosphorylation and impaired short‐term desensitization. Expression of the Del 301–303 ‐α 2B ‐adrenoceptor also is associated with an increased risk for myocardial infarction in humans. Recent evidence from our laboratory suggests a quantitative relationship between cellular GRK3 expression levels and the sensitivity of the α 2B ‐adrenoceptor to agonist‐induced down‐regulation. Therefore, the present study was undertaken to study agonist‐induced down‐regulation of the wild‐type (WT)‐ and Del 301–303 ‐α 2B ‐adrenoceptor in a neuronal cell model. Experimental approach: Haemagglutinin (HA) epitope‐tagged WT‐ and Del 301–303 ‐α 2B ‐adrenoceptor containing plasmids were constructed and the receptors were stably or transiently transfected in neuroblastoma/glioma hybrid NG108 cells. The expression levels in stable transfects were ∼50 fmol·mg −1 . These cells were used to examine agonist‐induced down‐regulation and phosphorylation of the WT‐ and Del 301–303 ‐α 2B ‐adrenoceptor. Key results: The Del 301–303 ‐α 2B ‐adrenoceptor, compared with the WT‐α 2B‐ adrenoceptor, displayed reduced adrenaline‐stimulated (20 µM) phosphorylation and did not down‐regulate in response to adrenaline (20–1000 µM). Using immunofluorescence labelling, we observed that transiently transfected WT‐α 2B ‐adrenoceptors internalized upon adrenaline treatment whereas the Del 301–303 ‐α 2B ‐adrenoceptor did not. Finally, we determined the effect of adrenaline on the Del 301–303 ‐α 2B ‐adrenoceptor in cells stably over‐expressing GRK3 3‐fold. In spite of the GRK3 over‐expression, 20–1000 µM ADR failed to down‐regulate or to increase phosphorylation of the Del 301–303 ‐α 2B ‐adrenoceptor in these cells. Conclusions and implications: The results suggest that the 301–303 deletion mutation of the α 2B ‐adrenoceptor eliminates agonist‐induced down‐regulation, an effect that cannot be overcome by increasing agonist concentration or by modest GRK3 over‐expression.