CHARACTERIZATION OF BIOCHEMICAL RESPONSES OF ANGIOTENSIN-II (AT2) BINDING-SITES IN THE RAT PHEOCHROMOCYTOMA PC12W CELLS

Rat pheochromocytoma PC12W cell membranes have previously been shown to exclusively contain the AT2 receptor subtype. The present study extended these binding data and explored the functional expression of these binding sites. Our binding competition studies show a potency series of Ang II = Ang III > saralasin > Ang I = PD123177 much greater than Ang II(1-7) >>> losartan. PD123177 (1-mu-M) completely eliminated [I-125]Ang II binding to PC12W cells. Competitive displacement of [I-125]Ang II with Ang II shows a dissociation equilibrium constant (K(d)) of 1.79 nM and a binding site maximum (B(max)) of 3.97 fmol/mg protein. Investigating several Ang II signal transduction pathways on these cells, we found that Ang II (10(-8) to 10(-6) M) docs not affect basal cAMP, cGMP, arachidonic acid release, prostacyclin release, intracellular Ca2+ mobilization or thymidine incorporation in the PC12W cells. Nerve growth factor, cAMP, 5-fluorouridine deoxyriboside modulation of the number of AT2 receptor sites in PC12W cells failed to unmask any Ang II effects on basal cAMP, cGMP and intracellular Ca2+ mobilization. In conclusion, the present study confirms the exclusive presence of AT2 binding sites in the PC12W cells. However, these binding sites are not functionally coupled to common signal transduction pathways.