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 [Ca2+]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 [Ca2+]i was calculated based on an ionomycin and Mn2+ quenching technique. As expected, TRH (1 mumol/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 [Ca2+]i, suggesting that changes of [Ca2+]i is not the sole final pathway of intracellular transduction systems for prolactin release.
Shin
S H
SH
Department of Physiology, Queen's University, Kingston, Ontario, Canada.
Soukup
C
C
Pang
S C
SC
Kubiseski
T J
TJ
Flynn
T G
TG
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002478:Cells, Cultured; D003470:Culture Media; D003600:Cytosol / Q000378:metabolism; D004958:Estradiol / Q000494:pharmacology*; D005110:Extracellular Space / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007211:Indoles; D015759:Ionomycin / Q000494:pharmacology; D008297:Male; D008345:Manganese / Q000494:pharmacology; D010637:Phenolsulfonphthalein; D010903:Pituitary Gland, Anterior / Q000378:metabolism / Q000502:physiology*; D011388:Prolactin / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D015398:Signal Transduction / Q000502:physiology; D013973:Thyrotropin-Releasing Hormone / Q000378:metabolism
D003470:Culture Media; D007211:Indoles; D008345:Manganese; D004958:Estradiol; D015759:Ionomycin; D013973:Thyrotropin-Releasing Hormone; D011388:Prolactin; D010637:Phenolsulfonphthalein; C048960:indo-1; D002118:Calcium