The effect of parathyroid hormone and thyrocalcitonin on the accumulation of cyclic adenosine 3':5'-monophosphate in freshly isolated bone cells.

Bone cells were isolated from embryonic rat calvaria by a modtied method designed to give larger yields and undamaged membranes. Essentially it consists of perfusing bone chips with fresh collagenase, collecting the cells continuously in the efiluent, and washing off the collagenase. Cyclic adenosine 3’:5’-monophosphate (CAMP) accumulation in the freshly isolated cells was measured by equilibrating the cells with the specific antibody. The amount of CAMP detected by this method was similar to that determined in the trichloroacetic acid extract from homogenized cells. Parathyroid hormone, thyrocalcitonin, and epinephrine enhanced CAMP accumulation in this preparation. The hormone response to parathyroid hormone was directly proportional to the logarithm of the concentration between 2 X lo-* and 2 pg per ml and between 2 x lop3 and 20 pg per ml, respectively. The lowest consistently effective concentration was 1 x lo-’ pg per ml (1 x lo-l4 M). The response to thyrocalcitonin was directly proportional to the logarithm of the concentration between 0.2 to 2 pg per ml, and the lowest active thyrocalcitonin concentration was 2 pg per ml (about 10F2 M). The effect of the two hormones administered together at maximum stimulatory concentration was additive, indicating that the hormones act on separate, independent sites. These findings show that physiological concentrations of both target-specific hormones, parathyroid hormone and thyrocalcitonin, enhance CAMP accumulation in the isolated bone cells and that the two hormones act on separate sites.