VIP receptor—Effector system in rat Harderian gland and its coupling to activation of type II thyroxine 5′-deiodinase

Vasoactive intestinal peptide (VIP) receptors were investigated in rat Harderian gland membranes using [125I]VIP as ligand. The receptor binding was rapid, reversible, saturable, specific, and dependent on time, temperature, and membrane concentration. At 30 degrees C, the stoichiometric data suggested the presence of two classes of VIP receptors with Kd values of 0.36 +/- 0.06 and 65.37 +/- 8.08 nM and binding capacities of 323 +/- 54 and 39,537 +/- 3100 fmol VIP/mg protein, respectively. The interaction showed a high degree of specificity, as suggested by competitive displacement experiments with several peptides structurally or not structurally related to VIP. The binding of [125I]VIP to membranes was sensitive to guanine nucleotides in a dose-dependent manner. The molecular characterization of VIP receptors was realized by chemical cross-linking; sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the solubilized membrane proteins revealed the presence of two specific [125I]VIP-protein complexes of M(r) 57 and 35 kDa as estimated in denaturing conditions. VIP stimulated adenylate cyclase activity in rat Harderian gland membranes in a dose-dependent manner. Finally, VIP stimulated in vivo the type II thyroxine 5'-deiodinase activity. These results demonstrate the presence of specific and functional VIP receptors in Harderian gland and suggest a role for VIP in the physiology of this gland.