The effect of extracellular polyvalent cations on bovine anterior pituitary cells. Evidence for a Ca(2+)-sensing receptor coupled to release of intracellular calcium stores.
We have investigated the effects of extracellular cations ([ION]ex) on cytosolic free calcium levels ([Ca2+]i) in bovine anterior pituitary (bAP) cells, using single-cell microfluorimetry. Increasing the [Ca2+]ex from 1 mM to 20 mM caused [Ca2+]i to increase in 64 +/- 14% of bAP cells. The [Ca2+]ex-induced [Ca2+]i increase was observed when cells were maintained in the presence of the voltage-gated-calcium-channel antagonist nitrendipine, but not when cells were treated with thapsigargin. Addition of [La3+]ex (5-15 microM) decreased [Ca2+]i, whereas 30 microM-1 mM caused a [Ca2+]i rise in 60.9 +/- 8.8% of bAP cells. [La3+]ex-induced [Ca2+]i changes were abolished by treating bAP-cells with either thapsigargin or ionomycin, but not nitrendipine. [La3+]ex at 15 microM did not increase [Ca2+]i in any cells tested, but when cells were treated with thimerosal, [La3+]ex (15 microM) caused a [Ca2+]i increase in 62.5 +/- 12.2% of bAP cells. In the presence of 1 mM [Ca2+]ex, successive additions of La3+ caused successive [Ca2+]i rises, but in nominally [Ca2+]ex-free medium only the first addition of [La3+]ex caused a [Ca2+]i rise. Addition of thyroliberin (TRH) in the presence of 1 mM [Ca2+]ex, caused [Ca2+]i to increase in 70% of bAP cells; subsequent addition of [La3+]ex (1 mM) only caused [Ca2+]i increases in 75% of those cells which had already responded to TRH. However, all cells which responded to 1 mM [La3+]ex also responded subsequently to TRH. After treatment with TRH in medium that was nominally [Ca2+]ex free, addition of La3+ (0.5-1 mM) did not increase [Ca2+]i in any cells tested. The number of cells which showed [La3+]ex-induced [Ca2+]i increases decreased in culture: only 21.75 +/- 2.2% cells responded after 7-11 days. When cells were cultured for 7-11 days in the presence of tunicamycin, [La3+]ex failed to increase [Ca2+]i in any cells tested. [Mn2+]ex rapidly quenched the Fura-2 signal measured from all bAP cells, but at 10 mM it also triggered a [Ca2+]i rise in about 60% of bAP cells. The Mn(2+)-induced [Ca2+]i rise was specifically abolished in cells cultured in the presence of tunicamycin although quenching was still observed. From these data we suggest that bAP cells may express a polyvalent cation receptor coupled to the release of calcium from intracellular stores.
Shorte
S L
SL
Department of Biochemistry, School of Medical Science, University of Bristol, UK.
Schofield
J G
JG
D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism / Q000494:pharmacology; D002135:Calcium-Binding Proteins / Q000378:metabolism*; D002413:Cations, Divalent / Q000494:pharmacology*; D002417:Cattle; D002478:Cells, Cultured / Q000187:drug effects / Q000502:physiology; D004594:Electrophysiology; D016257:Fura-2; D007811:Lanthanum / Q000494:pharmacology; D008345:Manganese / Q000494:pharmacology; D010903:Pituitary Gland, Anterior / Q000166:cytology* / Q000201:enzymology; D011493:Protein Kinase C / Q000378:metabolism; D013973:Thyrotropin-Releasing Hormone / Q000494:pharmacology; D013997:Time Factors; D014415:Tunicamycin / Q000494:pharmacology
D000900:Anti-Bacterial Agents; D002135:Calcium-Binding Proteins; D002413:Cations, Divalent; D014415:Tunicamycin; D008345:Manganese; D013973:Thyrotropin-Releasing Hormone; D007811:Lanthanum; D011493:Protein Kinase C; D002118:Calcium; D016257:Fura-2