JOURNAL OF EXPERIMENTAL ZOOLOGY 286:699–706 (2000)

© 2000 WILEY-LISS, INC.

Anoxic Performance of the American Eel (Anguillarostrata L.) Heart Requires Extracellular Glucose

JOHN R. BAILEY, KENNETH J. RODNICK, ROBERT MACDOUGALL,SEANA CLOWE, AND WILLIAM R. DRIEDZIC*Department of Biology, Mount Allison University, Sackville,New Brunswick, Canada

ABSTRACT The importance of extracellular glucose in the maintenance of performance of theheart of the American eel (Anguilla rostrata Le Sueur (L.) under anoxia was assessed under avariety of experimental conditions. Ventricular strips, electrically paced at 36 bpm, in N2-gassedmedium maintained the imposed pace rate and generated approximately 25% of the initial twitchforce of contraction for at least 60 min when glucose was present in the medium. But ventricularstrips challenged without glucose in the medium failed to maintain the pacing rate within 5–10min. Isolated and intact, perfused hearts maintained pressure and followed an imposed pace rateof 24 bpm for at least 2 hr, under anoxic conditions, if glucose was present in the medium. Butwithout glucose in the medium isolated hearts failed within 30 min. Endogenous glycogen storeswere utilized in hearts perfused with medium containing NaCN to impair oxidative phosphoryla-tion. The presence of glucose in the medium did not protect against glycogen mobilization. Thedata indicate that exogenous glucose is necessary to maintain performance under anoxia at highworkloads and physiological Ca2+ levels. Finally, ventricular strips treated with NaCN and forcedto contract at 24 bpm lost 70% of initial twitch force. Increasing extracellular Ca2+ concentrationstepwise from 1.5 to 9.5 mM restored twitch force to approximately 50% of the initial level andthis response was not dependent on exogenous glucose. However, glucose was required to main-tain resting tension even under normoxic conditions in the face of a Ca2+ challenge. J. Exp. Zool.286:699–706, 2000. © 2000 Wiley-Liss, Inc.

The hearts of some species of fish perform verywell in the absence of oxygen (Driedzic, ’92;Driedzic and Gesser, ’94). Under anoxic conditions,cardiac glycogen is mobilized and lactate is pro-duced through anaerobic glycolysis (e.g., Driedzicet al., ’78; van Waarde et al., ’83). Some studiessuggest but do not prove that extracellular glu-cose is also critical to the maintenance of func-tional integrity. This contention is based on thehigh activities of hexokinase generally found infish hearts (Driedzic, ’92; Driedzic and Gesser, ’94),the match between potential rates of ATP turn-over predicted from hexokinase and ATPase ac-tivities (Bailey et al., ’91), the activation of glucoseuptake in the hearts of American eel under con-ditions of hypoxic incubation (Rodnick et al., ’97),and the observation that extracellular glucose ex-tends the beat time of isolated hearts from twospecies of Amazonian fishes under conditions ofcyanide poisoning (Driedzic et al., ’78).

In this study, we directly test the hypothesis thatextracellular glucose is critical to the maintenanceof performance of the American eel heart under oxy-gen-limiting conditions and assess if glucose has a

protective effect on glycogen reserves. American eelwas selected since the intact isolated eel heart isable to tolerate extreme hypoxia in the presence ofglucose (Bailey et al., ’90). Additionally, even underanoxic conditions, force development by ventricu-lar strips from European eel can be enhancedthrough increases in extracellular calcium (Neilsonand Gesser, ’84a). Here we assess if this responseis dependent upon glucose availability. We consid-ered this to be of interest since in a marine teleost,Zoarces viviparous, ventricular strips from hypoxia-adapted individuals display a glucose-dependent,calcium enhancement of contractile force under an-oxia (Driedzic et al., ’85).

We report that under anaerobic conditions thereis a necessity for extracellular glucose to maintainheart performance. The positive inotropic effect of

Grant sponsors: Natural Sciences and Engineering Research Coun-cil of Canada and the New Brunswick Heart and Stroke Foundation.

K.J. Rodnick’s present address: Department of Biological Sciences,Idaho State University, Pocatello, Idaho 83209-8007.

*Correspondence to: William R. Driedzic, Ocean Sciences Cen-tre, Memorial University of Newfoundland, St. John’s, NF, CanadaA1C 5S7.

Received 21 December 1998; Accepted 29 October 1999

700 J.R. BAILEY ET AL.

extracellular calcium under oxygen-limiting condi-tions is confirmed and shown to be glucose inde-pendent. A novel finding that extracellular glucoseis intimately related to the regulation of resting ten-sion in isolated ventricular strip preparations evenunder aerobic conditions is also presented.

MATERIALS AND METHODSAnimals

American eels (Anguilla rostrata L.) were pur-chased from South Shore Trading Co., Port Elgin,NB. Animals were transported to Mount Allisonand kept in flow-through aquaria at 13–15°C un-til use. In all experiments, eels were removed fromthe holding tank, rendered torpid in ice water, anddoubly pithed. Hearts were quickly excised andplaced in a beaker of ice-cold Ringer’s solution.

MediaThe bathing or perfusion medium was a basic

Ringer’s for freshwater fishes (Hoar and Hickman,’67), containing 111 mM NaCl, 2.0 mM KCl, 1.5mM CaCl2, 1.0 mM MgSO4 and 5.0 mM NaH2PO4,modified by addition of extra bicarbonate (10 mM)to aid in long-term buffering to pH 7.8 at 15°C.In addition, 5 mM glucose was added as an exog-enous fuel in some preparations. In studies in-volving ventricular strips, the medium was gassedwith either 0.5% CO2 : balance O2 or 0.5% CO2 :balance N2 for the duration of the experiment. Inisolated perfused heart studies, the medium wasgassed with either 0.5% CO2 : balance air or 0.5%CO2 : balance N2 for the duration of the experi-ment. Chemical impairment of oxidative phospho-rylation was achieved through the addition ofNaCN solution to the medium to give a final con-centration of 1 mM. Increased calcium in the bath-ing medium was achieved by the addition ofaliquots of a CaCl2 solution, giving a concentra-tion increase of 1 mM with each addition. The ex-perimental temperature was 15°C throughout.

PreparationsVentricular strips

The ventricle was dissected free of the atriumand bulbus and bisected. Each section of ventriclewas rinsed in cold bathing medium to wash outblood. Two strips were cut from each piece of ven-tricle, for a total of four strips. Each strip wasmounted in an isolated strip bath between plati-num wire electrodes and connected to a HarvardApparatus (Saint-Laurent, Quebec, Canada) iso-metric force transducer (model 60-2994), which was

interfaced to a Biotronix BL-882 strip chart re-corder. Strips were stimulated to contract via fieldstimulation with a Grass model S9 square-wavegenerator (Grass Medical Instruments, Quincy,MA) connected to the platinum wire electrodes.Oxygenated medium was added to each bath, andthe ventricular strips were stretched to an opti-mum length for peak force generation and allowedto equilibrate for 15 min at a pacing rate of 36(exp. 1) or 24 (exp. 4) beats · min–1 (bpm). Supra-threshold voltage, determined at the start of eachexperiment, was used throughout. Resting heartrate for A. anguilla has been reported as 29 bpmat 12°C (Janvier, ’97) and 26 bpm at 16°C, withadrenergic stimulation increasing rates to 45 bpm(Peyraud-Waitzenegger et al., ’80).

Perfused heartsIsolated hearts were perfused in a manner pre-

viously described (Driedzic and Bailey, ’94). In per-formance studies (exp. 2), the atrium was fittedwith a stainless steel cannula and connected to awater-jacketed filling head 3 cm above the heart,which in turn was filled from a water-jacketed res-ervoir of perfusion medium. The ventricle was alsocannulated via the bulbus arteriosus with a stain-less steel cannula, which lead to an imposedafterload of 25 cm H2O. Pressure developed by theventricle was monitored with a Biotronix BL 630pressure transducer interfaced to a Biotronix BL882 strip chart recorder. Flow rates were moni-tored with a Biotronix BL 610 pulsed logic flow-meter. Mean pressure development by the isolatedventricle was approximately 40 cm H2O in con-trol preparations. This was somewhat lower thanthe ventral aortic pressure in A. anguilla, whichapproximates 50 cm H2O (Janvier, ’97). Initial car-diac output was approximately 30 mL · kg–1 · min–1

which is higher than the 20 mL · kg–1 · min–1 re-ported for A. anguilla. In glycogen determinationstudies (exp. 3), there was no imposed afterload.The hearts emptied to the atmosphere and wereforce-filled at a flow rate of 25 mL · kg–1 body mass· min–1, with flow rate being monitored via aGilmont Instruments (Great Neck, NY) flowme-ter. In both cases, hearts were stimulated to con-tract with a Grass model S9 square-wave generatorset at 5 V and 200 msec duration.

Experimental protocolsExperiment 1

The capacity of ventricular strips to maintainperformance at rates of contraction typical for in-

ANOXIC PERFORMANCE OF THE EEL HEART 701

tact normoxic animals was assessed. Following theequilibration period, the oxygenated medium wasreplaced with anoxic medium in two of the fourbaths. Glucose was added to one oxygenated bathand one anoxic bath. All strips were then pacedat 36 bpm for 120 min or until contractile failure.

Experiment 2The findings with ventricular strips were ex-

tended to monitor the performance of intact iso-lated hearts. Hearts were perfused with normoxicRinger’s plus glucose for 10 min to clear the bloodand to establish baseline performance. Prepara-tions were paced at 24 bpm, although under aer-ated conditions hearts assumed an inherent ratehigher than the set pacing regime. Contractionwas regular with respect to rates of change inpressure development and relaxation with no evi-dence of extra-systolic events or other abnormali-ties. Following the equilibration period, the heartswere then perfused with either normoxic mediumplus 5 mM glucose, anoxic medium plus 5 mMglucose, or anoxic medium without glucose. Ex-periments continued for 120 min or until contrac-tions were no longer evident.

Experiment 3Further experiments involved measurements of

glycogen content in perfused hearts. Hearts werestimulated at a frequency of 30 bpm and perfusedwith normoxic Ringer’s for 15 min to clear theblood. These hearts did not have an imposedafterload and emptied to atmosphere. They wereperfused under four conditions: with glucose inthe perfusion medium; without glucose in the me-dium; with glucose plus NaCN; or without glu-cose but with NaCN. Hearts were perfused for30 min with the media listed above, the experi-ment terminated, and the ventricles were freeze-clamped for analysis of glycogen content.

Experiment 4Finally, the capacity of ventricular strips to re-

spond to a calcium challenge, under conditions ofsimulated anoxia, was assessed with or withoutglucose in the medium. Oxidative phosphorylationwas inhibited with NaCN in order to achieve arapid setting to a lower level of performance. Thebathing solution consisted of oxygenated mediumwith either no external glucose or 5 mM glucoseadded. Strips were paced at 24 bpm, since pre-liminary work revealed that preparations fail tomaintain pacing rates at high frequencies (>30bpm) and high calcium loads. Following the equili-

bration period, NaCN was added to two of the fourbaths to a final concentration of 1 mM, and all ofthe strips were then allowed to equilibrate for anadditional 15 min. Then 100 µl of a 200 mM CaCl2solution were immediately added to each tissuebath. This step was repeated at 3-min intervalsuntil the calcium concentration reached 9.5 mM.

Glycogen analysisGlycogen was extracted from ventricles by first

digesting the tissue in 0.3 mL of 30% KOH (w/v)for 10 min in a boiling water bath. Glycogen wasprecipitated out of the digestate by adding 0.2 mLof 2% aqueous Na2SO4 and 2.0 mL absolute etha-nol. The resultant mixture was centrifuged at1,500g for 10 min and the supernatant decanted.The pellet was washed in 66% ethanol and thenredissolved in 2 mL of warm distilled water. An ali-quot of the solution was acid hydrolyzed by the ad-dition of an equal volume of 1.2 M HCl and heatingin a boiling water bath for 2 hr (Sephton andDriedzic, ’94). Hydrolysates were neutralized withthe addition of 1 M NaOH to pH 7.0 and analyzedfor glucose content using a commercial colorimetrickit (Sigma no. 115-A, Oakville, Ontario, Canada).

Data analysisStrip chart recordings obtained during the ven-

tricular strip experiments were analyzed forchanges in isometric twitch force development andin resting tension, which are expressed as a per-centage value of twitch force development at T =0 min. Recordings obtained during the isolatedperfused heart experiments were analyzed forpressure development, heart rate and flow rate.Pressure values are expressed as mm Hg pres-sure, heart rate as bpm, and flow rates as mL ·kg–1 body mass · min–1. Glycogen content is ex-pressed as mg glycogen · g–1 tissue mass. All dataare expressed as means ± SEM. Differencesbetween means were analyzed using the Mann-Whitney “U” nonparametric test for the ventric-ular strip experiments and the Student’s t-testfor the perfused heart studies. A P value less than0.05 was considered significant.

RESULTSExperiment 1

Under oxygenated conditions, ventricular stripsmaintained the imposed pacing regime of 36 bpm(Fig. 1). The presence or absence of glucose did notmake a significant difference in performance un-der oxygenated conditions, as force fell to about

702 J.R. BAILEY ET AL.

75% of the initial level after 120 min in both cases.Ventricular strips subjected to anoxic conditions,with glucose in the medium, could not maintainthe initial level of performance. Twitch force felloff within 15 min of onset of the anoxic period, andafter 60 min of treatment was about 25% of theinitial level. Then five of seven strips subsequentlyfailed to maintain the pacing regime at T + 60 min,while two strips maintained the pacing regime untilT + 90 min. Under anoxic conditions, strips with-out exogenous glucose failed to follow the pacingregime within 5–10 min of the onset of anoxia. Inmost cases, these preparations continued to con-tract, albeit in either an irregular fashion or a lowerthan imposed frequency for at least 75 min. It isnot valid to relate either irregular contractions orcontractions at a slower than imposed rate to ini-tial twitch-force development at a fixed frequency.Consequently, there are no quantitative data thatmay be shown for these strips. The qualitative find-ing, though, is that the ventricular strips, underanoxia, performed better with glucose than with-out glucose in the medium.

Experiment 2Hearts perfused under normoxic conditions

maintained a contraction rate substantially higherthan the imposed rate, presumably by followingan intrinsic pattern (Fig. 2A). Hearts perfusedunder anoxic conditions with glucose in the me-dium over the perfusion period showed a signifi-cant decrease to the imposed rate of 24 bpm (Fig.2B). The decrease in contraction rate was evenmore precipitous under perfusion conditions ofanoxia without glucose (Fig. 3C), and all heartsfailed to contract after 30 min of anoxic treatment.Hearts perfused with the normoxic medium (plusglucose) maintained contractility throughout theperfusion period without any change in outputpressure (Fig. 3). Hearts perfused with the an-oxic medium (plus glucose) also maintained con-tractility throughout the perfusion period, albeitat a lower level when compared to the normoxichearts (Fig. 3). If glucose was not present in theperfusion medium, output pressure decreased forhearts under anoxic conditions. When hearts wereperfused with the anoxic medium plus glucose flowthrough, the hearts decreased by 33%, from 30 ±6 to 20 ± 5 ml · kg–1 · min–1, during the first 15min of the anoxic period but thereafter stabilizedat the lower level. However, when there was noglucose in the anoxic medium, flow through thesehearts decreased rapidly, from 33 ± 3 (T = 0) to 6± 0.2 ml · kg–1 · min–1 (T = 30), until the point offailure shortly after T = 30 min. Unfortunately,flow was not measured under normoxic conditionsplus glucose, but numerous other studies haveshown that when pressure and rate are main-tained flow rate is maintained as well.Experiment 3

Hearts perfused under control conditions (i.e., noNaCN) had a significantly greater glycogen contentat the time of sampling than those perfused underconditions of impaired oxidative phosphorylation byNaCN (Table 1), regardless of whether glucose waspresent or absent in the medium. A comparison ofglycogen content between hearts perfused undercontrol conditions with and without glucose yieldedno significant difference. Likewise, there was no sig-nificant difference when the glycogen content ofhearts perfused with and without glucose in the me-dium containing NaCN were compared.Experiment 4

Control ventricular strips (i.e., no NaCN), witheither 5 mM glucose in the medium or no glucosein the medium, responded to the serial addition

Fig. 1. Twitch-force development in ventricular stripsfrom eel heart under oxygenated and anoxic conditions. Allvalues are means ± SEM, N = 7 in all cases except wherenoted. In these cases the strips continued to maintain thepacing regime of 36 bpm, although the majority of similarlytreated strips had failed at this point. Squares representstrips under oxygenated conditions with glucose in the me-dium; triangles represent strips under oxygenated conditionswith no glucose in the medium, and circles represent stripsunder anoxic conditions with glucose in the medium. Aster-isks indicate that force development of preparations sub-jected to anoxia without glucose in the medium wassignificantly different from the other conditions.

ANOXIC PERFORMANCE OF THE EEL HEART 703

of calcium with a twitch-force increase to a maxi-mum of approximately 120% in the first case and115% in the second case (Fig. 4A,C). Ventricularstrips treated with NaCN and with 5 mM glucosein the medium showed an initial decrease intwitch force generated to approximately 30% ofthe initial force (Fig. 4A). The addition of extra-cellular calcium to the bathing medium restoreda percentage of the twitch force of the strips, suchthat at 9.5 mM Ca2+, force was 60% of the origi-nal (Fig. 4A). When extracellular glucose was notpresent, NaCN-treated ventricular strips showeda similar initial decrease in twitch force, and theaddition of extracellular calcium restored twitchforce to approximately 50% of initial (Fig. 4C). Thepresence of glucose in the medium did not have a

Fig. 2. Heart rate in isolated perfused eel hearts. All valuesare means ± SEM, and N = 6 in all cases. A: Hearts perfusedunder normoxic conditions with glucose in the medium. B:Hearts perfused under anoxic conditions with glucose in themedium. C: Hearts perfused under anoxic conditions withoutglucose in the medium. The initial point on this panel repre-sents an equilibration point prior to the change to anoxic me-dium without glucose. There are no error bars on the last pointsince the heart rate was following the externally imposed rate.

Fig. 3. Pressure development in isolated perfused eelhearts. All values are means ± SEM, and N = 6 in all cases.Squares represent hearts perfused under normoxic conditionswith glucose in the medium; circles represent hearts perfusedunder anoxic conditions with glucose in the medium; and tri-angles represent hearts perfused under anoxic conditionswithout glucose in the medium. The initial point for heartsperfused under anoxic conditions without glucose in the me-dium represents an equilibration point prior to the change toanoxic medium without glucose.

TABLE 1. Glycogen content of isolated eel hearts perfusedwith and without NaCN1

Medium Control NaCN

Ringer’s solution 3.7 ± 0.31 2.5 ± 0.57*without glucose (5) (6)

Ringer’s solution 3.3 ± 0.63 1.4 ± 0.44*plus glucose (6) (6)

1All values are means ± SEM and are expressed as mg glycogen g–1

tissue mass.*P < 0.05 comparisons being made between the control media andthe media with NaCN.

704 J.R. BAILEY ET AL.

significant effect on the contractile response to thecalcium challenge in either untreated or NaCN-treated ventricular strips.

There was an immediate increase in restingtension in ventricular strips following treatmentwith NaCN and with glucose in the medium(Fig. 4B), but the resting tension did not changewith increasing calcium concentration. The con-trol strips (i.e., glucose without NaCN) did notshow any change in resting tension over the ex-perimental period (Fig. 4B). NaCN-treatment

of ventricular strips in glucose-free medium re-sulted in an immediate 35% increase in restingtension, and there was a slight increase to ap-proximately 40% with increasing calcium con-centration (Fig. 4D). When glucose was absentfrom the medium (without NaCN) an increas-ing calcium concentration resulted in a 20% in-crease in resting tension of the ventricularstrips (Fig. 4D). This was in marked contrastto the situation with glucose in the medium (4Blower curve vs. 4D lower curve).

Fig. 4. Twitch-force generation and resting tension in iso-lated ventricular strips from eel heart with increasing cal-cium concentration. All values are means ± SEM. Squaresrepresent strips under control conditions and circles repre-sent strips under NaCN treatment. In all panels, the stripsunder NaCN conditions are all significantly different fromthe strips under control conditions. In all cases the mediumwas gassed with 99.5% O2 : 0.5% CO2, and the frequency ofcontraction was 24 bpm. A: Force generation by strips incu-bated with glucose in the medium (N = 9); asterisks indicatethat preparations treated with CN showed a significant in-crease in force development at Ca2+ levels 5.5 mM and higher,

relative to force development at 1.5 mM Ca2+. B: Resting ten-sion in strips incubated with glucose in the medium (N = 9).C: Force generation by strips incubated without glucose inthe medium (N = 8); asterisks indicate that preparationstreated with CN showed a significant increase in force devel-opment at Ca2+ levels 5.5 mM and higher, relative to forcedevelopment at 1.5 mM Ca2+. D: Resting tension by stripsincubated without glucose in the medium (N = 8); asterisksindicate that resting tension in control preparations (i.e., with-out glucose or NaCN) increased at Ca2+ levels of 2.5 mM andhigher, relative to resting tension at 1.5 mM Ca2+.

ANOXIC PERFORMANCE OF THE EEL HEART 705

DISCUSSIONGlucose is required for the American eel heart

to maintain performance at high rates of energydemand under oxygen-limiting conditions. Ven-tricular strips, paced at 36 bpm, maintained con-tractility under anoxia for 60 min with a forcedevelopment of about 25% of the initial level, butin the absence of glucose, contractility failedwithin 10 min. Isolated perfused hearts, undernormoxic conditions, have an intrinsic rate ofabout 60 bpm. Under anoxic conditions, with glu-cose in the medium, these hearts followed the im-posed pacing rate of 24 bpm and maintainedpressure development over the 2 hr of perfusion.Flow rate decreased by approximately 30% underthese conditions, but in the absence of glucose,intact hearts perfused with the anoxic mediumfailed within 30 min. The dependency upon ex-tracellular glucose is consistent with high levelsof hexokinase, the first enzyme-catalyzed step inglucose utilization, and the activation of glucosetransporter mechanisms under anoxia in the eelheart (Rodnick et al., ’97). Although dependentupon exogenous glucose, glycogen is also utilizedunder conditions of impaired oxidative phospho-rylation. Glycogen levels in perfused hearts werelower under cyanide-poisoning than control con-ditions regardless of the presence or absence ofglucose in the medium. Thus, exogenous glucosedoes not have a protective effect on the glycogenpool. The utilization of glucose and/or glycogenunder anoxia is similar in American eel and turtlehearts, both of which display high resistance tooxygen limitation. Under anaerobic conditions,mechanical work by the isolated perfused heartof the turtle (Pseudemys scripta) is compromisedwithout exogenous glucose. At high workloads theturtle heart calls upon both glycogen and glucose,and at low workloads glycogen stores are utilizedeven in the presence of glucose (Reeves, ’63).

Ventricular strips from American eel are similarto European eel, rainbow trout, and Zoarces vivipa-rous (Nielsen and Gesser, ’84a,b; Driedzic et al., ’85;Hansen and Gesser, ’87) in their ability to increaseforce generation in response to a calcium challenge,even under conditions of impaired oxidative phos-phorylation. This response in the American eel isnot dependent on exogenous glucose, as the ven-tricular strips returned to 50–60% of their initialforce regardless of the presence or absence of glu-cose. Therefore, the possibility that the mechanisminvolves a calcium-activated glucose transport, ashas been shown for mammalian skeletal muscle

(Nolte et al., ’95) and was suggested for hypoxia-adapted Zoarces viviparous (Driedzic et al., ’85),may be ruled out in this case. The increase inforce of contraction caused by extracellular cal-cium could be due to the effect of increased in-tracellular calcium activity on the contractilefibrils, or alternatively, an activation of glycogenphosphorylase, enhancing ATP production (Frie-sen et al., ’69).

Ventricular strips treated with NaCN showedabout a 40% increase in resting tension with orwithout glucose in the medium. The inability ofthe heart to relax following impairment of oxida-tive metabolism is a well-recognized response andis considered to be the result of an increase incytoplasmic calcium activity (Hansen and Gesser,’87; Gesser and Hoglund, ’88; Hartmund andGesser, ’96). Increases in extracellular calciumwere without further effect, and glucose could notameliorate the impact of NaCN. The relationshipbetween glucose and resting tension under con-trol conditions, (i.e., without NaCN) warrants spe-cial mention. Ventricular strips subjected toincreases in extracellular calcium concentrationface the challenge of managing cytoplasmic cal-cium activity against an increased calcium gradi-ent. Ventricular strips which received glucose inthe medium maintained resting tension in asso-ciation with a calcium challenge. These prepara-tions were able to relax fully, implying an abilityto decrease cytoplasmic calcium activity. Ventricu-lar strips which were incubated without glucosein the medium, under control conditions, showedan approximately 20% increase in resting tensionat the highest calcium concentration. The reasonfor this is not known, but in rat heart, relaxationis dependent upon exogenous glucose under hy-poxic conditions, and this effect is independent oftotal ATP concentration (Anderson and Morris,’78). Additionally, calcium uptake by rat heart sar-coplasmic reticulum under oxygen-limiting condi-tions is dependent upon glycolytically generatedATP, as a number of glycolytic enzymes includinghexokinase are associated with the sarcoplasmicreticulum membrane (Xu et al., ’95). Although thefunction of the sarcoplasmic reticulum in calciummanagement in fish hearts is generally consid-ered to be minimal (Driedzic and Gesser, ’85, ’88,’94), there are cases in which it may play an ac-tive role, especially at high contraction frequen-cies (Keen et al., ’92) or high temperatures (Shielsand Farrel, ’97). The role of the sarcoplasmicreticulum in eel heart is not known, but currentstudies are under way to address this question.

706 J.R. BAILEY ET AL.

In summary, the presence of an exogenous glu-cose supply is necessary to maintain force/pres-sure development of the eel heart under anoxia.Although glucose is required, endogenous glyco-gen stores are also utilized. Ventricular strips areable to increase twitch-force generation under an-oxic conditions in the face of a calcium challenge.This response is not dependent on an exogenousglucose supply. It appears that a component of thecapacity to decrease cytosolic calcium activity evenunder oxygenated conditions is glucose dependent.

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