Capacitative Ca2+ entry contributes to the Ca2+ influx induced by thyrotropin-releasing hormone (TRH) in GH3 pituitary cells.
Treatment of GH3 cells with either hypothalamic peptide thyrotropin-releasing hormone (TRH), the endomembrane Ca2+-ATPase inhibitor thapsigargin or the Ca2+ ionophore ionomycin mobilized, with different kinetics, essentially all of the Ca2+ pool from the intracellular Ca2+ stores. Any of the above- described treatments induced a sustained increase in intracellular Ca2+ concentration ([Ca2+]i), which was dependent on extracellular Ca2+ and was prevented by Ni2+ but not by dihydropyridines (DHPs), suggesting that it was due to capacitative Ca2+ entry via activation of a plasma membrane pathway which opened upon the emptying of the intracellular Ca2+ stores. The increase of the plasma membrane permeability to Ca2+ correlated negatively with the filling degree of the intracellular Ca2+ stores and was reversed by refilling of the stores. The mechanism of capacitative Ca2+ entry into GH3 cells differed from similar mechanisms described in several types of blood cells in that the pathway was poorly permeable to Mn2+ and not sensitive to cytochrome P450 inhibitors. In GH3 cells, TRH induced a transient [Ca2+]i increase due to Ca2+ release from the stores (phase 1) followed by a sustained [Ca2+]i increase due to Ca2+ entry (phase 2). At the single-cell level, phase 2 was composed of a DHP-insensitive sustained [Ca2+]i increase, due to activation of capacitative Ca2+ entry, superimposed upon which DHP- sensitive [Ca2+]i oscillations took place. The two components of the TRH-induced Ca2+ entry differed also in that [Ca2+]i oscillations remained for several minutes after TRH removal, whereas the sustained [Ca2+]i increase dropped quickly to prestimulatory levels, following the same time course as the refilling of the stores. The drop was prevented when the refilling was inhibited by thapsigargin. It is concluded that, even though the mechanisms of capacitative Ca2+ entry may show differences from cell to cell, it is also present and may contribute to the regulation of physiological functions in excitable cells such as GH3. There, capacitative Ca2+ entry cooperates with voltage-gated Ca2+ channels to generate the [Ca2+]i increase seen during phase 2 of TRH action. This contribution of capacitative Ca2+ entry may be relevant to the enhancement of prolactin secretion induced by TRH.
Villalobos
C
C
Instituto de Biología y Genética Molecular (IBGM), Universidad de Valladolid, E-47005 Valladolid, Spain.
García-Sancho
J
J
D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels / Q000187:drug effects*; D000252:Calcium-Transporting ATPases / Q000037:antagonists & inhibitors; D002460:Cell Line; D002463:Cell Membrane Permeability; D065607:Cytochrome P-450 Enzyme Inhibitors; D004095:Dihydropyridines / Q000494:pharmacology; D007425:Intracellular Membranes / Q000378:metabolism; D015759:Ionomycin / Q000494:pharmacology; D008345:Manganese / Q000493:pharmacokinetics; D010903:Pituitary Gland, Anterior / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D015398:Signal Transduction; D013729:Terpenes / Q000494:pharmacology; D019284:Thapsigargin; D013973:Thyrotropin-Releasing Hormone / Q000494:pharmacology*
D015220:Calcium Channels; D065607:Cytochrome P-450 Enzyme Inhibitors; D004095:Dihydropyridines; D013729:Terpenes; D008345:Manganese; D015759:Ionomycin; D013973:Thyrotropin-Releasing Hormone; D019284:Thapsigargin; D000252:Calcium-Transporting ATPases; D002118:Calcium