Glucose inhibits the high-affinity (Ca²⁺ + Mg²⁺)-ATPase in the plasma membrane of a glucose-responsive insulinoma

(Ca²⁺ + Mg²⁺)-ATPase enzyme activity of a purified plasma membrane preparation from a glucose responsive rat insulinoma, was characterized as Ca²⁺-dependent dephosphorylation of [γ-³²P]ATP. A high-affinity enzyme with a K_m(ATP) raging from 20 to 30 μM and a K_m(Ca²⁺) of 1 μM was identified. Glucose inhibited this high-affinity enzyme in a dose-dependent manner, with no significant inhibition at a concentration between 0 and 5 mM, 50% inhibition at 13.3 mM and 94.5% inhibition at 30 mM. The inhibitory effect of glucose was immediate and rapidly reversible. The effect of stereospecific for the α-anomer. These findings support the concept that glucose acts directly at the β-cell plasma membrane and is involved in the maintenance of elevated intracellular free calcium concentrations associated with insulin release by directly or indirectly inhibiting energy-dependent calcium efflux. Glyceraldehyde (20 mM) increased enzyme activity 3-fold, while other metabolic fuels had no effect. This suggests that inhibition of the enzyme is not an obligatory requirement for insulin release. Calmodulin stimulated the enzyme activity in calmodulin-depleted but not in undepleted membranes. Trifluoperazine (30-100 μM) inhibited (Ca¹⁺ + Mg²⁺)-ATPase in a dose-dependent manner (14-61% activity) and the activity was also inhibited by vanadate (0.1-1.0 mM) and NaCl (150 mM).