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1 Animal and Poultry Science, University of Guelph, Guelph, ON, Canada
* To whom correspondence should be addressed. E-mail: cxiao{at}uoguelph.ca.
Glucose transport kinetics were quantified in isolated bovine mammary epithelial cells using 3-O-methyl-D-glucose (3OMG). Isolated cells retained satisfactory viability and glucose uptake activity that was inhibited by cytochalasin B (CCB), phloretin, HgCl2 and low temperature. Initial rates of entry were measured over a 15-s interval at 37 °C under zero-trans, equilibrium-exchange, high-cis and high-trans concentrations of 3OMG between 0 and 20 mM. The combined set of rate measurements from all experimental conditions was fit to the fixed-site carrier model by non-linear regression to estimate parameters of transport. The regression between predicted and observed initial rates had an r2 of 0.97. Forward Vmax was estimated at 18.2 nmol min-1 mg protein-1 and the Michaelis constant was 8.29 mM. The cooperativity parameter was 1.63, trans-stimulation was 2.13-fold and asymmetry was 2.06-fold. Based on the kinetic parameters, variations in intracellular glucose concentrations are not responsible for the range of glucose uptakes by bovine mammary glands observed in vivo.
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