Hormone responses of in vitro bone nodule cells: studies on changes of intracellular calcium and membrane potential in response to parathyroid hormone and calcitonin

Abstract

We used two techniques to study the responses of individual in vitro bone nodule cells to parathyroid hormone (PTH) and calcitonin (CT). These techniques are laser scanning confocal imaging with a fluorescent indicator to measure intracellular free [Ca2+], and microelectrode impalement to measure the electrical potential difference across the cell membrane. We applied these measurement techniques to cells in the top cellular layer of nodules that form in vitro in cultures of cells obtained from fetal rat calvaria. Our measurements showed a transient increase in intracellular free [Ca2+] following application of PTH or CT. The duration of the increase in fluorescent intensity following PTH application varied from about 100 to more than 300 s, and the duration following CT application was from 30 to 80 s. In some measurements we applied both hormones in sequence, and observed that some cells showed an intracellular [Ca2+] response to both hormones, while other cells apparently responded to only one or the other of the hormones, or to neither. We also observed membrane potential changes in response to PTH and to CT. The membrane potential response to CT was quite small. The time courses of these membrane potential changes consisted of a depolarizing phase lasting about 100 s (with both hormones) followed by a hyperpolarizing phase (with PTH). Control measurements using only the vehicle solutions were carried out with both techniques, producing negligible responses.