Cysteamine, zinc, and thiols modify detectability of rat pituitary prolactin: A comparison with effects on bovine prolactin suggests differences in hormone storage

Little is known about the structure of prolactin (PRL) within secretory granules. Evidence from our previous studies in bovine tissue preparations suggests that control of secretion may reside, in part, in the conversion of storage hormone to releasable PRL. The conversion can be monitored by measuring changes in immunodetectability since the oligomeric, storage form is poorly recognized by antisera raised against monomeric PRL. Since many investigators use rats to study the secretory process and changes in detectability of rat pituitary PRL occur during lactation (“depletion-transformation”), we undertook the present immunodetectability studies to gain insight into the storage structure of rat (r) PRL. Cysteamine and zinc inhibited tissue PRL immunoassayability in male rat pituitary homogenates and also in partially purified secretory granules as they had inhibited bovine (b) PRL; however, zinc inhibited the rodent hormone less potently than the bovine. In vitro incubation of rat tissue samples without additions resulted in increases in rPRL detectability of up to 84% after 180 minutes; such incubation of bovine samples had no significant effect. A striking additional difference between the species was that exposure to reduced glutathione (GSH), cysteine, homocysteine, mercaptoethanol, and dithiothreitol inhibited rPRL by up to 44%. This compared to thiol stimulation of bPRL by as much as 450%. The inhibitory GSH effect on rPRL was abolished when 0.5% sodium dodecyl sulfate (SDS) was included; in contrast, the stimulatory GSH effect on bPRL did not change with added SDS. SDS alone had no effect on rat homogenate PRL, and only increased rat granule rPRL by 23% compared to its ability to increase bPRL assayability by 44%. High performance liquid chromatography demonstrated a high molecular weight peak of immunoreactive rat PRL. This was largely abolished, with a simultaneous increase in monomer and intermediate zone material, by treatment of extracts with either GSH or 4 mol/L urea. The immunoreactivity of the high molecular weight peak was increased nearly 50% by treatment with GSH. The striking qualitative discrepancy between rPRL and bPRL responses to GSH and other thiols cannot be accounted for by dosage of exposure, the use of fresh v frozen tissue, or old v young donor animals. The mode of hormone packaging within secretory granules or differences in nonhormonal granule components (eg, enzymes) may explain the differential responses. Alternatively, si