Measurement of prolactin release and cytosolic calcium in estradiol-primed lactotrophs

We have developed a perifusion system that can measure both changes of cytosolic free calcium concentration [Ca 2+] i and prolactin release simultaneously from cultured lactotrophs. This model incorporated a commonly-used perifusion system to a spectrofluorometer. Indo-1 loaded cells were injected into Sephadex G-150 matrix in the cuvette at a site where the emitting light of the fluorometer projects. During perifusion periods, the perifusate was collected in a fraction collector, while optical density of the emitting light at 405 nm was recorded. The [Ca 2+] i was calculated based on an ionomycin and Mn 2+ quenching technique. As expected, TRH (1 μmol/l) stimulated prolactin release from cultured lactotrophs in this system. We further observed that prolactin releases as induced by TRH and ionomycin were not proportional with changes of the [Ca 2+] i, suggesting that changes of [Ca 2+] i is not the sole final pathway of intracellular transduction systems for prolactin release.