The avian eggshell acts as the major source of calcium for embryogenesis in bird. Using scanning electron microscopy, the inner eggshell surface was analysed before and after embryonic development and its accompanying calcium removal. This was done in eggs from two bird species with different growth rates and modes of development, the precocial Japanese quail Coturnix japonica and the altricial starling Sturnus vulgaris. Next, enzyme histochemistry was used to localize carbonic anhydrase in the extra-embryonic chorioallantoic membrane to provide support for the hypothesis that calcium may be released from the eggshell by means of acidification through the action of carbonic anhydrase. However, neither in the precocial quail nor in the altricial starling could any staining for carbonic anhydrase be detected. These results lead us to the conclusion that the role of carbonic anhydrase in embryonic calcium mobilization from the avian eggshell remains unclear.

Data are presented suggesting that birds have evolved eggs with shells containing different structures (numbers of mammillae per unit of inner eggshell surface area, i.e., mammillary densities) to cope up with different calcium requirements imposed by different growth rates and modes of development. Precocial bird species grow slowly, but have high mammillary density, while altricial bird species grow rapidly, but have low mammillary density. These results suggest an adaptation associated with growth rate and mode of development and show, moreover, that the mammillary layer is indicative of the breeding biology of the bird.

The avian eggshell contributes to successful embryogenesis by acting as the major source of calcium. It has been suggested, moreover, that birds have evolved eggshells with different structures adapted to cope with different calcium requirements imposed by different growth rates and modes of development. Yet the mechanism of calcium removal from the avian eggshell by the embryo remains unknown. Using enzyme histochemistry, we here test an old hypothesis stating that calcium is released from the eggshell by means of acidification by the action of carbonic anhydrase (CA) in the extra-embryonic chorio-allantoic membrane (CAM). This enzyme catalyses the reversible reaction CO2+H2O <-> H+HCO3 and is known to take significant part in acid-base regulation in the body. We examined CAMs from two bird species, Coturnix japonica and Sturnus vulgaris, with very different growth rates and modes of development. The CAMs were examined for CA activity at the later stages of embryonic development. However, in all CAMs examined, staining for CA was absent, leading us to the conclusion that existing hypothesis about a link between CA and embryonic calcium mobilization from the avian eggshell is at present premature.