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First published online April 23, 2004
Journal of Experimental Biology 207, 1865-1874 (2004)
Published by The Company of Biologists 2004
doi: 10.1242/jeb.00965

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The regulation and importance of glucose uptake in the isolated Atlantic cod heart: rate-limiting steps and effects of hypoxia

Kathy A. Clow¹, Kenneth J. Rodnick², Tyson J. MacCormack¹ and William R. Driedzic^1,*

¹ Ocean Sciences Centre, Memorial University of Newfoundland, St John's, Newfoundland, Canada, A1C 5S7
² Department of Biological Sciences, Idaho State University, Pocatello, ID 83209-8007, USA

View larger version (21K): [in a new window]   Fig. 1. Power output (%) of isolated cod (Gadus morhua) hearts perfused under either normoxic or hypoxic conditions, with or without 5 mmol l-1 glucose, or hypoxic conditions with 5 mmol l-1 glucose and 25 µmol l-1 cytochalasin B in the medium. All values are means ± S.E.M. Numbers under the first data point represent the total number of hearts perfused in that group. Numbers after this initial point correspond to the number of hearts still functioning at that time.

View larger version (17K): [in a new window]   Fig. 2. 2-Deoxyglucose (2-DG) uptake in perfused cod (Gadus morhua) hearts. All hearts were perfused for 15 min with 5 mmol l-1 glucose in the medium under normoxic or hypoxic conditions. One hypoxic group was also perfused with 25 µmol l-1 cytochalasin B in the medium. All values are means ± S.E.M. and N=8 for all groups. *Significantly different from normoxia + glucose group (P<0.001). Significantly different from both the normoxia + glucose group (P<0.001) and hypoxia + glucose group (P<0.01).

View larger version (20K): [in a new window]   Fig. 3. (A) Glycogen and (B) free glucose levels in the cod (Gadus morhua) perfused heart subjected to either normoxic or hypoxic conditions, with or without 5 mmol l-1 glucose, or hypoxic conditions with 5 mmol l-1 glucose and 25 µmol l-1 cytochalasin B in the medium. These levels were taken after 120 min or immediately after the heart failed. All values are means ± S.E.M. and N=6–11 fish. *Significantly different from normoxia + glucose group (P<0.05). In the normoxic without glucose group, free glucose levels were not significantly different when compared with the normoxic with glucose group (P=0.054).

View larger version (15K): [in a new window]   Fig. 4. Effects of Na+-free medium or phloridzin (1 mmol l-1) on 2-deoxyglucose (2-DG) uptake in cod (Gadus morhua) ventricle strips. After dissection, strips were incubated for 60 min in either regular medium (basal condition), Na+-free medium containing equimolar choline, or regular medium containing 1 mmol l-1 phloridzin. All media were supplemented with 5 mmol l-1 glucose, 35 mmol l-1 mannitol and 0.1% bovine serum albumin (BSA). Ventricle strips were rinsed in glucose-free medium and then assayed for glucose uptake activity in the presence of 1 mmol l-1 2-deoxy-D-[3H(G)]glucose. Values are means ± S.E.M. for six strips per bar. *P<0.01 versus basal condition.

View larger version (13K): [in a new window]   Fig. 5. (A) Time course of 2-deoxyglucose (2-DG) uptake in cod (Gadus morhua) ventricle strips incubated with 1 mmol l-1 2-DG. Ventricle strips were incubated at 8°C for 60 min in medium containing 5 mmol l-1 glucose, 35 mmol l-1 mannitol and 0.1% bovine serum albumin (BSA). Strips were rinsed in glucose-free medium for 10 min and then incubated in medium containing 2 mmol l-1 pyruvate, 1 mmol l-1 2-deoxy-D-[3H(G)]glucose, 37 mmol l-1 [U-14C]mannitol and 0.1% BSA to assay uptake for the indicated times. ICS indicates intracellular space. Values are means ± S.E.M. for six strips per point. (B) Relationship between 2-DG uptake and 2-deoxyglucose-6-phosphate (2-DG-6-P) accumulation in cod ventricle strips under basal conditions using 1 mmol l-1 2-deoxy-D-[3H(G)]glucose. Strips were frozen after increasing periods of incubation (10–60 min) at 8°C. Intracellular 2-DG and 2-DG-6-P were measured after separation by ion-exchange chromatography into neutral and anionic fractions, respectively. The accumulation of 2-DG-6-P mirrored the rate of 2-DG uptake, whereas free 2-DG represented a minor component. The results are shown for 24 strips from six animals.