INTRACELLULAR CALCIUM-CONCENTRATION AND HORMONE-SECRETION ARE CONTROLLED DIFFERENTLY BY TRH IN RAT NEONATAL LACTOTROPHS AND SOMATOTROPHS

We studied the effects of TRH on the cytosolic free calcium concentrat ion ([Ca2+](i)) of female rat pituitary prolactin-secreting (lactotrop h) and GH-secreting (somatotroph) cells in the early postnatal period, i.e. at postnatal days 5 and 10. [Ca2+](i) of single identified lacto trophs and somatotrophs was recorded by dual-emission microspectrofluo rimetry using the intracellular fluorescent calcium probe indo 1. An a pplication of TRH (100 nM, 10 s) induced a marked [Ca2+](i) increase i n 65% of neonatal lactotrophs and 34% of neonatal somatotrophs while t he remaining cells were unaffected. Most of the responsive cells, both lactotrophs and somatotrophs, exhibited a similar biphasic Ca2+ respo nse, made up of an initial rapid large increase in [Ca2+](i) followed by sustained [Ca2+](i) fluctuations. In both cell types, removal of Ca 2+ from the extracellular medium or addition of the Ca2+ channel block er, cadmium chloride (500 mu M), inhibited the second phase whereas th e first phase persisted. Furthermore, in both cell types, protein kina se C (PKC) depletion by incubation in phorbol myristate acetate (1 mu M) for 24 h abolished the second phase but did not inhibit the first p hase. Conversely, when cells were pretreated with the Ca2+-ATPase inhi bitor, thapsigargin (100 nM), all TRH-induced [Ca2+](i) changes in bot h cell types disappeared. TRH therefore induces a biphasic increase in [Ca2+](i) involving intra-and extracellular Ca2+ in neonatal lactotro phs and somatotrophs as it does in adult lactotrophs. The first phase is presumably due to mobilization of Ca2+ from intracellular stores wh ereas the second phase presumably results from a PKC-sensitive influx of Ca2+. TRH action on membrane potential was then investigated using the patch-clamp technique in the whole-cell mode. TRH-induced changes in membrane potential consisted of an initial hyperpolarization follow ed by depolarization and action potential firing. We also investigated TRH action on prolactin and GH secretion by neonatal pituitary cells using RIA. Surprisingly, static assays of prolactin and GH revealed on ly stimulation of prolactin release by TRH but no effect on GH secreti on, although, as expected, GH-releasing factor was a potent agonist of GH secretion. Our results suggest that TRH regulates neonatal lactotr ophs and somatotrophs differently, in that the [Ca2+](i) changes do no t correlate with stimulation of exocytosis in the latter cell type.