Intracellular Transport and Packaging of Prolactin: a Quantitative Electron Microscope Autoradiographic Study of Mammotrophs Dissociated from Rat Pituitaries

Dispersed pituitary cells prepared from estrogen-treated female rats were subjected to pulse labeling with [3H]leucine (5 min) followed by a chase incubation (up to 3 h) in order to study intracellular transport of PRL in mammotrophs. Sites of synthesis, rates of transport, and sites of packaging and storage of PRL were determined by quantitative electron microscopic autoradiography. Results of grain counts show that label is initially (end of pulse) distributed randomly over the rough endoplasmic reticulum (ER), but rapidly (5--15 min of chase) moves to the stacked Golgi cisternae where concentration into secretion granules takes place. The label moves successively from small (Type I) immature granules (15--55 min of chase) to large (Types II and III) polymorphic granules (55--115 min) in the Golgi region, to rounded or ovoid mature (Type IV) granules (55--185 min) usually found in the peripheral cytoplasm, indicating that these types of granules represent successive stages in granule concentration and assembly. Analysis of the relative grain density (percentage of total grains/percentage of total area) confirmed that there was progressive concentration (up to 20--150 times) along the transport route with the concentration lowest in the ER, higher in the Golgi, and highest in immature and mature secretion granules. These data indicate that synthesis of PRL occurs randomly in the ER, transport to the Golgi occurs rapidly (within 5--10 min), and is completed rapidly (90% within 15--20 min), and concentration into granules and aggregation of small granules into larger forms also occurs rapidly (by 15--20 min), but goes on over a prolonged period of time (up to 3 h). Use of dispersed cells has allowed a more precise determination of the location and kinetics of steps in the intracellular processing of PRL than has been possible previously using other systems.