Characterization of 1,25-dihydroxyvitamin D3-dependent calcium uptake in cultured embryonic chick duodenum.

To confirm that embryonic chick duodenum in organ culture is a suitable model for studing the mechanism by which la,25-dihydroxyvitamin D3 stimulates intestinal calcium transport, we have further characterized this system using the following criteria: calcium uptake properties, responsiveness to vitamin D3 analogs, and properties of a la,25-dihydroxyvitamin D3 receptor-like binding protein.Our studies suggest that this in vitro system is quite similar to the rachitic chick system by all criteria examined.The calcium uptake response is saturable ( K , for calcium = 0.45 mM) and is optimal in low sodium-mannitol buffers but is not blocked by respiratory and glycolytic inhibitors.Treatment with la.25-dihydroxyvitamin D3 stimulates calcium uptake by increasing the maximal velocity approximately 2.2fold without altering K,.Significant increases in calcium uptake could be detected after culturing with la,25-dihydroxyvitamin D3 for 4.5 to 6 h with the response continuing to increase for up to 24 h.The ability of vitamin D3 analogs to stimulate calcium uptake generally parallels their in vivo biological activities, la,25dihydroxyvitamin D3 > la-hydroxyvitamin D3 ;> 25-hydroxyvitamin D3 > 24R,25-dihydroxyvitamin D3 > vitamin D3.A specific 1,25-dihydroxyvitamin D3 receptorlike binding protein is present in embryonic duodenum that is identical with the receptor from chick intestine.It sediments at 3.7 S, has an equilibrium dissociation constant for la,25-dihydroxyvitamin D3 of 8.5 X lo-" M, and binds vitamin DI analogs in order of their biological activity.These studies provide further support for the use of embryonic chick duodenum as a model for studies on the mechanism of action of la,25-dihydroxyvitamin D3 in intestine.Cultured embryonic chick duodenum has been used extensively to bioassay vitamin D metabolites (1-4), and as a tool for studying the mechanism of action of la,25-dihydroxyvitamin D R (5-9).Much evidence has been presented (1-3, 5-9) that suggests that the hormonal response in this system is generally similar to that of rachitic chicks.