chlortetracycline and Pituitary-Neoplasms

Topics: 1-Methyl-3-isobutylxanthine; Animals; Calcium; Cells, Cultured; Chlortetracycline; Cyclic AMP; Guanylyl Imidodiphosphate; Ion Channels; Pituitary Gland; Pituitary Neoplasms; Potassium; Prolactin; Protein Kinases; Sodium Fluoride; Thyrotropin; Thyrotropin-Releasing Hormone; Verapamil

We have shown that arachidonic acid stimulates 45Ca2+ efflux from prelabeled rat pituitary mammotropic (GH3) cells resuspended in "Ca2+-free" medium (Kolesnick, R. N., Mussachio, I., Thaw, C., and Gershengorn, M. C. (1984) Am. J. Physiol. 246, E458-E462). In this study, we further characterize the effects of arachidonic acid on Ca2+ homeostasis in GH3 cells and demonstrate its antagonism of changes induced by thyrotropin-releasing hormone (TRH). At below 5 microM, arachidonic acid stimulated intracellular for extracellular Ca2+ exchange without affecting cell Ca2+ content. Above 5 microM, arachidonic acid decreased membrane-bound Ca2+, as monitored by chlortetracycline, and decreased total cell 45Ca2+ content by depleting nonmitochondrial and mitochondrial pools. However, arachidonic acid did not elevate cytoplasmic free Ca2+ concentration ([Ca2+]i). Arachidonic acid inhibited TRH-induced 45Ca2+ efflux, loss of membrane-bound Ca2+, mobilization of nonmitochondrial Ca2+, and elevation of [Ca2+]i. Arachidonic acid also lowered elevated [Ca2+]i caused by release of mitochondrial Ca2+ with an uncoupler or by influx of extracellular Ca2+ stimulated with K+ depolarization. Hence, arachidonic acid stimulates Ca2+ extrusion from and depletes Ca2+ stores within GH3 cells. We suggest that arachidonic acid may be an important regulator of cellular Ca2+ homeostasis which may inhibit TRH-induced elevation of [Ca2+]i.

Topics: Animals; Arachidonic Acid; Arachidonic Acids; Calcium; Cell Line; Chlortetracycline; Culture Media; Membrane Potentials; Mitochondria; Pituitary Neoplasms; Rats; Serum Albumin, Bovine; Spectrometry, Fluorescence; Thyrotropin-Releasing Hormone

Chlortetracycline (CTC), a probe of membrane-bound divalent cations, was used to study the action of thyrotropin-releasing hormone (TRH) in mouse pituitary thyrotropic tumor (TtT) cells in culture. Cellular fluorescence of CTC was caused by both Ca2+- and Mg2+-CTC complexes and was influenced by the concentration of these cations in the incubation medium. TRH, but not other peptides, caused a rapid, transient, and concentration-dependent decrease in the CTC fluorescence intensity; half-maximal effect occurred with 10--30 nM TRH. The decrement in fluorescence intensity caused by TRH was not due to enhanced loss of CTC from the cells. The decrease in fluorescence elicited by TRH was specific for Ca2+-CTC complexes because preincubation of the cells with 1 mM EGTA or 1 mM EDTA plus 2.05 mM Mg2+ abolished the response, whereas preincubation with 1 mM EDTA plus 2.05 mM Ca2+ permitted the usual TRH response. Antimycin A and carbonyl cyanide m-chlorophenylhydrazone decreased cellular ATP content to 37 +/- 1 and 32 +/- 1% of control, respectively, and abolished the TRH-induced decrease in CTC fluorescence. We conclude that TRH displaced Ca2+ from an energy-dependent, membrane-bound pool(s) within TtT cells and that this may be one mechanism by which the concentration of intracellular free Ca2+ is raised so that is couples stimulation by TRH to TSH secretion.

Topics: Animals; Calcium; Cell Membrane; Chlortetracycline; Cobalt; Edetic Acid; Egtazic Acid; Kinetics; Magnesium; Mice; Neoplasms, Experimental; Pituitary Neoplasms; Thyrotropin; Thyrotropin-Releasing Hormone