PROSTAGLANDIN-E2 AND F2-ALPHA BINDING-SITES IN THE BOVINE IRIS-CILIARY BODY

Purpose. To determine the regional distribution and selectivity of pro staglandin E2 (PGE2) and prostaglandin F2alpha (PGF2alpha) specific bi nding sites in membrane preparations from bovine iris ciliary body. Me thods. Bovine eyes were obtained fresh from a local abattoir. Whole ir ides were separated into sphincter muscle, ciliary body, and remaining iris tissue then homogenized in 50 mM tris buffer, pH 7.5, containing cyclooxygenase, protease, and soybean trypsin inhibitors. Membranes w ere obtained after three-stage high-speed centrifugation then reconsti tuted in buffer. Aliquot portions were incubated with H-3-PGE2 or H-3- PGF2alpha at 37-degrees-C for 30 min in a final volume of 500 mul. Com petition studies were performed in the presence of up to 1000-fold exc ess unlabeled ligand. At the end of a 30-min incubation period, free a nd membrane bound ligand were separated by rapid filtration through a type HA millipore filter preequilibrated with buffer. The radioactivit y bound to the membranes retained on the filter was quantitated in a s cintillation counter. Results. The equilibrium dissociation constant a nd maximum number 3H-PGE2 saturable binding sites were determined by S catchard analysis. The data best fit to a single binding site model fo r all three membrane preparations. The majority of the H-3-PGE2 specif ic binding sites were in sphincter muscle (65%), followed by iris (I 7 %), and ciliary body (I 8%). H-3-PGF2alpha binding sites were not meas urable in either iris or ciliary body and PGF2alpha was less competiti ve than PGE2 in all tissues. The rank order of potency for agonist dis placement by both H-3-PGE2 and H-3-PGF2 in sphincter muscle membrane w as PGE2 > PGF2alpha > PGD2 > Iloprost. It was determined that H-3-PGE2 and H-3-PGF2alpha binding in the bovine sphincter was primarily to EP not FP, DP, or IP prostaglandin receptor types. Conclusions. The char acteristics of PGE2 and PGF2alpha specific binding sites in the bovine sphincter correlate with its in vitro contractile response to these p rostanoids. The inability of PGF2alpha to effectively compete with PGE 2 for specific binding sites in the sphincter and lack of high affinit y PGF2alpha specific binding sites in the iris and ciliary body sugges ts that this prostaglandin may exert its in vivo effects through PGE2 specific binding sites.