Evaluation of a protocol for studying the chick chorioallantoic membrane in vitro

Explants of eggshell with and without the chorioallantoic membrane were taken from fertile chicken eggs on day 16 of incubation and exposed in vitro to inhibitors (acetazolamide and benzolamide) of carbonic anhydrase to determine if enzyme inhibition affected release of calcium from the shell. A separate experiment examined the effect of the metabolic poison dinitrophenol (DNP) on release of calcium from explants. Explants with the chorioallantois in situ released more calcium than those lacking the epithelium, but neither the enzyme inhibitors nor DNP affected release of calcium. The lack of effect of the enzyme inhibitors could indicate that activity of carbonic anhydrase is not as important to the release of calcium from the eggshell as has been assumed. However, the absence of an effect of DNP instead indicates that release of calcium mediated by the chorioallantois in vitro simply lacks physiological relevance. Thus, results of this investigation raise doubts that the mechanism underlying release of calcium from the eggshell can be assessed in vitro.