REGULATION OF ADENYLYL-CYCLASE IN POLARIZED RENAL EPITHELIAL-CELLS BYG-PROTEIN-COUPLED RECEPTORS

We employed two guanine nucleotide binding protein (G protein)-coupled receptors known to be targeted to opposite domains in renal epithelia l cells to test the hypothesis that the polarized receptor expression of receptors regulates the activity of the receptor's effector molecul e, adenylyl cyclase. We used LLC-PK1 cells stably transfected with cDN A encoding the alpha(2B)-adrenergic receptor (alpha(2B)-AR) or A(1)-ad enosine receptor (A(1)-AdR). Immunohistochemistry and Western blot ana lysis confirmed the basolateral and apical expression of alpha(2B)-ARs and A(1)-AdRs, respectively. Adenylyl cyclase activity was assessed b y measuring cAMP accumulation following the addition of forskolin (10 mu M) in the presence of 3-isobutyl-1-methylxanthine to apical or baso lateral chambers of confluent monolayers. A five- to sixfold increase in cAMP accumulation occurred following apical (or basolateral) stimul ation of LLC-PK1 cells expressing apical (or basolateral) receptors in comparison to forskolin stimulation of corresponding domains of untra nsfected cells. We conclude 1) adenylyl cyclase activity is present at or near the apical and basolateral domains of LLC-PK1 cells, and 2) f actors that regulate the polarized expression of inhibitory G protein- coupled receptors may also regulate local adenylyl cyclase activity.