Stability of Secretory Granules Containing Growth Hormone and Prolactin from Bovine Anterior Pituitary Gland

Abstract

Bovine adenohypophyseal secretory granules were purified by a technique giving a high yield of large granules containing 70–90% of prolactin (PRL) and 10–20% of growth hormone (GH). Purity of the preparation was checked by electron microscopy and enzymatic markers. Stability of the isolated secretory granules was studied by measurement of the optical density of the granule suspensions and by measurement of GH and PRL release by disc gel electrophoresis at pH 8.9. At 0 °C, granules suspended in an hypotonic medium are stable at pH 7.4. At room temperature, they are particularly stable at acidic pH but are almost completely solubilized at a pH ranging from 7.0 to 10.0, a half-maximal effect on PRL release being observed at pH 7.5. Lysolecithin has a marked solubilizing effect at a pH of 3.0–6.0 whereas 0.15% sodium dodecyl sulfate does not affect granule stability at a pH below 5. ATP (0.5 mM) inhibits GH and PRL release to, respectively, 65 and 27% of the control rates during a 60 min incubation at 37 °C and pH 7.4. A concentration of 2 mM Ca2+ has a marked inhibitory effect on both GH and PRL release whereas 2 mM Mg2+ inhibits PRL release but does not affect the release of GH. EGTA increases PRL release by 30% but does not significantly affect GH release. As evidenced by polyacrylamide gel electrophoresis, no degradation of GH or PRL occurred during a 1 h incubation of secretory granules at 37 °C and pH 7.4. These data suggest a role of Ca2+- and Mg2+-ATPase activities on the stability and possibly the formation of GH- and PRL-containing secretory granules.