The long-term objective is to determine the cellular and extracellular factors that influence adaptation of the small intestine's absorptive processes to changed circumstances, especially diabetes. The two specific transport proteins to be studied are Na,K-ATPase and the Na-glucose cotransporter, both of which are functionally altered in experimental diabetes. Studies will be performed on rat small intestine and, in parallel, on 2 other model systems - differentiated Caco-2 colon cancer cells and brine shrimp intestine.
The specific aims are to (1) determine Na and K affinities of intestinal Na, K-ATPase under control conditions, and under conditions which modify ouabain sensitivity (measurements will be made in both intact cells and isolated membranes); (2) purify rat intestinal Na,K-ATPase and determine at which stage differences in affinities for ouabain, Na or K present in intact cells are lost; (3) then analyze solubilized membrane-attached proteins to determine if any regulate Na,K- ATPase; (4) establish whether PGE rapidly alters the ouabain- sensitivity of Na,K-ATPase; (5) determine cell Na with NMR and relate to changes in the Na pump; (6) measure also the effect of glucagon, diabetes and absorptive stimuli on the number of ouabain binding sites on intact cells; (7) using 3 cDNA probes for isoforms of the catalytic subunit of Na,K-ATPase, determine the presence and prevalence of the specific mRNA's in rat intestine, Caco-2 cells and brine shrimp intestine; (8) determine if the diabetes-induced increase in number of Na-glucose cotransporters is due to a decrease in epithelial cell turnover; (9) determine effects of diabetes on other brush border enzymes and transport processes; (10) determine if glucagon alone or in combination with insulin deficiency stimulates Na-glucose cotransport; determine also effects of other growth factors; (11) establish Na-glucose cotransporter function in Caco-2 cells and explore means for its enhancement; and (12) determine if alterations in cell Na influence proliferation of Na-glucose cotransporters.
| Minhas, B S; Sullivan, S K; Field, M (1993) Bicarbonate secretion in rabbit ileum: electrogenicity, ion dependence, and effects of cyclic nucleotides. Gastroenterology 105:1617-29 |
| Fedorak, R N; Cortas, N; Field, M (1991) Diabetes mellitus and glucagon alter ouabain-sensitive Na(+)-K(+)-ATPase in rat small intestine. Diabetes 40:1603-10 |
