Imaging of intracellular calcium in rat anterior pituitary cells in response to growth hormone releasing factor.

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Abstract
1. Changes in intracellular ionized calcium [Ca2+]i induced by human growth hormone releasing factor (hGRF) were analysed by quantitative fluorescent microscopy using a dual‐wavelength, ratiometric video imaging system and low light level charge‐coupled device (CCD) camera visualizing Fura‐2 in dispersed male rat anterior pituitary cells. 2. In cells responding to hGRF, spontaneous basal oscillations in [Ca2+]i were frequently observed, and these were usually characterized by a gradient of [Ca2+]i localized in the subplasmalemmal region of the cell. 3. Of the cells which responded to hGRF, the peptide evoked a rise in [Ca2+]i, especially in the region of the subplasmalemma. Continuous application of 10 nM‐hGRF produced several different temporal patterns of the [Ca2+]i response which were not attributable to spatial response profiles. A sustained rise in [Ca2+]i was the most common type of response to hGRF (44% of the cells examined). 4. One‐third of the cells responding to 10 nM‐hGRF showed spontaneous basal [Ca2+]i oscillations ranging from 100 to 500 nM. Mean values of basal and 10 nM‐hGRF‐induced [Ca2+]i of these cells were 81 +/‐ 11 nM (mean +/‐ S.E.M., n = 27) and 560 +/‐ 47 nM (n = 27) respectively. There was no significant correlation between basal [Ca2+]i and the hGRF‐induced [Ca2+]i increase, nor was there any consistent correlation with regard to the spatial response profile. 5. Application of 2 mM‐Co2+ abolished the hGRF‐induced rise in [Ca2+]i. Quantitative analysis of this effect, performed by comparing the mean [Ca2+]i evoked during the application of hGRF with and without Co2+, respectively, also showed significant inhibition of the hGRF‐induced rise in [Ca2+]i by the application of Co2+ (P less than 0.001). 6. The hGRF‐induced rise in [Ca2+]i was completely suppressed by replacing extracellular Na+ with impermeant molecules such as mannitol. The onset and offset of suppression was as rapid as that induced by Co2+. Quantitative analysis showed significant inhibition of the hGRF‐induced rise in [Ca2+]i by Na+ replacement (P less than 0.01). 7. Tetrodotoxin, a potent blocker of voltage‐sensitive Na+ channels (5 and 20 microM), did not affect the hGRF‐induced rise in [Ca2+]i. 8. Extracellular application of the membrane permeable dibutyryl cyclic AMP (DBcAMP) to elevate intracellular levels of cyclic AMP caused a large rise in [Ca2+]i, which was dependent on extracellular Na+ and was abolished by 2 mM‐Co2+ applied in the bath.(ABSTRACT TRUNCATED AT 400 WORDS)