EXPRESSION OF VASOPRESSIN RECEPTORS (V(2)-SUBTYPE) ON LLC-PK1 CELLS DURING CELL-CULTURE

Vasopressin receptor expression on LLC-PK1-cells (a porcine renal tubu lar cell line) during cell culture is still not fully understood. We s tudied receptor expression using a novel vasopressin analogue with hig h specific radioactivity 25][8-p-hydroxy-phenylpropionyl]-lys8-vasopre ssin, 74EBq/mol (2000 Ci/mmol)). LLC-PK1 cells were grown in monolayer s for 1 to 6 days. Scatchard analysis performed with membranes of LLC- PK1 cells revealed a single binding site with a binding constant (K(d) ) of 0.46 +/- 0.04 nmol/l. During cell culture, the binding constant ( K(d)) was not altered, but receptor density increased significantly (2 1 115 +/- 645 receptors per cell, day 2; 42 315 +/- 1512 receptors per cell, day 6). A receptor occupancy of about 30% was found to be assoc iated with a cAMP stimulation of 50%. The receptor reserve might be ev en higher because, by using a highly specific oxytocin antagonist, we found that 20% of the occupied -hydroxy-phenylpropionyl]-lys8-vasopres sin-binding sites are oxytocin receptors. For lys8 -vasopressin recept or studies, great care has to be taken to examine cells in identical c ulture phases.