RADIOIMMUNOASSAY OF RAT PROLACTIN AND ITS USE IN MEASURING PROLACTIN PRODUCTION BY CULTURED PITUITARY CELLS

Abstract

A sensitive and specific radioimmunoassay was developed for rat prolactin (rPRL), employing the double antibody solid phase technique for the separation of free and antibody-bound [125I]rPRL. The antiserum was raised in rabbits and showed no cross-reaction with rat growth hormone (rGH), follicle stimulating hormone (rFSH), luteinizing hormone (rLH) and thyrotropin (rTSH). The immunosorbent (sheep anti-rabbit IgG [immunoglobulin G] bound to cellulose) showed a surprisingly high binding of [125I]rPRL, but not of the other iodinated anterior pituitary hormones. Addition of serum to the incubation mixtures prevented the binding between [125I]rPRL and the immunosorbent. Three different clonal strains of pituitary cells were examined for production of rPRL, rLH and rTSH, in the basal state and after treatment with thyrotropin releasing hormone (TRH) and gonadotropin releasing hormone (LH/FSH-RH). Monolayer cultures of 2 of the cell strains produced and secreted rPRL spontaneously, and they showed a 2-fold increase in rPRL production after treatment with TRH (3 .cntdot. 10-7 mol/l). The 3rd cell strain did not produce rPRL spontaneously, or after treatment with TRH. None of these cell strains could be stimulated to produce rTSH by treatment with TRH. Treatment of the same 3 cell strains with LH/FSH-RH (1.2 .cntdot. 10-6 mol/l) failed to induce production of rLH, and there were no changes in production of rPRL. Prostaglandins E1 and E2 (3 .cntdot. 10-8 mol/l) and estradiol-17.beta. (10-7-10-10 mol/l) stimulated the production of rPRL. The effects of TRH and prostaglandins E1 and E2 were observed within 24 h of treatment, while the first effect of estradiol-17.beta. was seen after 3 days. The stimulatory effect of estradiol-17.beta. on rPRL production may differ from that of TRH and prostaglandins.