Selective coupling of α₂-adrenergic receptor subtypes to cyclic AMP-dependent reporter gene expression in transiently transfected JEG-3 cells

A cAMP-dependent reporter gene has been used in transiently transfected human choriocarcinoma (JEG-3) cells to examine the second messenger coupling of the human α₂-adrenergic receptor subtypes. The reporter gene consists of a cAMP response element linked to the gene for chloramphenicol acetyltransferase (CAT). Plasmids encoding the α₂-C10 (α_2A), α₂-C2 (α_2B), or α₂-C4 (α_2C) receptor subtypes were co-transfected with a plasmid containing the reporter gene, and the ability of α₂ receptor agonists to influence forskolin-stimulated CAT expression was examined. For α₂-C10, agonists had a biphasic effect on forskolin-stimulated CAT expression. Thus, low (nanomolar) concentrations of agonist inhibited CAT expression by ∼60%, whereas high (micromolar) concentrations reversed this inhibition and could even potentiate CAT expression by as much as 140%. A significantly different pattern of coupling was observed for the other α₂ receptor subtypes. For α₂-C4, agonists only inhibited forskolin-stimulated CAT expression, whereas for α₂-C2 only potentiation of expression was seen. Each of these responses was specifically blocked by α₂- but not α₁- or β-adrenergic receptor antagonists. For α₂-C4, the inhibition of forskolin-stimulated CAT expression was prevented by pretreatment of the cells with pertussis toxin. This was also true for the inhibition obtained with α₂-C10. The potentiation of CAT expression, however, was not prevented by pertussis toxin pretreatment in cells transfected with either α₂-C2 or α₂-C10. In this transient expression system, each α₂-adrenergic receptor subtype had access to the same complement of G proteins, adenylyl cyclase, and other second messengers. It would appear, therefore, that the potential for the activation of unique intracellular responses exists even among closely related receptor subtypes.