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Effect of glycolytic inhibitors on the sinoatrial node, atrium and atrioventricular node in the rabbit heart
Journal
of Molecular
and Cellular
Effect Atrium Jochen
Cardiology
(1981)
13,
811-821
of Glycolytic Inhibitors and Atrioventricular Senges,
Johannes and
Abteilung
Innere Medizin
on the Sinoatrial Node in the Rabbit
Brachmann, Wolfgang
Dieter Kiibler
Pelzer,
III (h*urdiologie), Medizinische 69 Heidelberg, Germany
(Received 23 June 1980, accepted in revisedform
Node, Heart*
Ioannis
Rizos
C:niversitiitsklinik, 12 May 1981)
J. SENGES, J.
BRACHMANN, D. PELZER, I. Rrzos AND W. K~~BLER. Effect of Glycolytic Inhibitors on the Sinoatrial Node, Atrium and Atrioventricular Node in the Rabbit Heart. Journal of Molecular and Cellular Cardiology (1981) 13, 811-82 1. The effect of unspecific noncompetitive (iodoacetir acid ; IA:\) and of specific competitive (deoxyglucose) glycolytic inhibitors were studied under aerobic and anaerobic conditions in the isolated sinoatrial ISA) node, right atrium and atriovcntricular (A\‘) node of the rabbit heart. Trammembrane action potentials were recorded simultaneously with the His bundle rlectrogram. Under aerobic conditions (PO, 460 mmHg), 7.5 x 10-j Y IAA caused no significant alterations of sinus rate, intraatrial and AV nodal conduction. Under hypoxic conditions (PO, 40 mmHg), 7.5 ./ 10-j M IAA resulted in rapid and complete abolition of the SA and A\’ nodal electrical activity and in a more delayed atria1 inexcitability. Similar changes were never observed in the presence of hypoxia alone. Higher concentrations of IA.4 (5 Y 1O-1 M) severely depressed the sinus rate and the AV nodal conduction already under aerobic conditions. The action potential amplitude and the rate of diastolic depolarization of the SA nodal fibers was significantly reduced, the maximum diastolic potential remained unchanged. In atria1 fibers, 5 x 1OV M IAA cattsed predominantly shortening of the action potential duration but had less marked and delayed depressant effects on the action potential amplitude. Elrctrophysiologic abnormalities included MobitL type II sinoatrial block. The electrophysiological effects of deoxyglucose (50 mM) under aerobic and hypoxic conditions resembled those obtained with 7.5 x 10e5 M IAA. The results suggest that the “normal” nodal function is predominantly related to aerobic metabolism but that under hypoxic conditions glycolytic energy production can effectively contribute to the maintenance of nodal elec,triral activity. Also that the specific relation between cardiac metabolism and electrical activity of nodal and atria1 cells may at least partly be explained by the particular electrical membrane properties of the various fiber types. KEY
IVORDS:
action
potential.
Iodoacetic
acid;
Deoxyglucose;
Anoxia;
Glycolysis;
Slow
response;
Nodal
Introduction Knowledge of the relationship between cardiac muscle metabolism function has been advanced by many authors to an appreciable metabolic processes involved in the maintenance and production activity of the heart including automaticity and conduction have been studied [7, 8, 18-21, 241. The present experiments were derived results suggesting that hypoxia inhibits predominantly the sinoatrial ventricular (AV) nodal transmembrane action potentials dependent * This work was supported vascullres System”. This work fur Kreislaufforschung, Bad International Society for Heart 0022%2828/8
l/O9081
1 + 11
by the Deutsche Forschungsgemeinschaft was presented in part at the 1979 meeting Nauheim and at the 1979 meeting of Research, Dijon.
$02.00/O
0
1981
Academic
and contractile extent but the of electrical less extensively from previous (SA) and atrioon the slow
within the SFB 90 “Cardioof the Deutsche Gessellschaft the European Section of the Press
Inc.
(London)
Limited
812
J. Senges
et
al.
inward current but has only little, if any effect on atria1 and ventricular specialized conduction related to the rapid Na current [14, 151. The contribution of glycolytic energy metabolism to the maintenance of nodal electrical activity is strongly supported by recent clinical and experimental observations demonstrating a close relation between SA and AV nodal function and the extracellular glucose concentration [16, 171. The purpose of the present experiments was to study the combined effects of glycolytic inhibitors and hypoxia on various electrophysiological parameters of cardiac automaticity and conduction. Iodoacetic acid (IAA) in low concentrations produces block of the Embden-Meyerhof glycolytic chain by inhibiting triophosphate dehydrogenase and exerts general sulfhydryl inhibiting effects at higher concentrations [.5, 201. In order to demonstrate that the electrophysiologic alterations observed in the presence of IAA were due to inhibition of glycolysis rather than to unspecific blockade of SH groups, the effects of deoxyglucose were also tested. Deoxyglucose has been reported to induce block of glucose catabolism by competitive inhibition of deoxyglucose-6-PO, on glucose-6-PO, at the phosphoglucoisomerase level [22]. Materials
and
Methods
Rabbits weighing 2 to 3 kg were killed by a single blow. The heart was rapidly removed and dissected in an oxygenated Tyrode’s solution of the following composition (mM) [I]: NaCl, 107.7; KCl, 3.48; CaCl,, 1.53; MgSO,, 0.69; NaHCO,, 26.2; NaH,PO,, 1.67; Na gluconate, 9.64; glucose, 5.5; sucrose, 7.6. The preparation consisting of sinoatrial node, crista terminalis, pectinate muscles of the atria1 appendage, intraatrial septum, coronary sinus, atrioventricular node and His bundle [12] was mounted in a I5 ml tissue bath and perfused at a rate of 30 ml/min with Tyrode’s solution at 36 f 0.5% A mixture of 95O;, 0, and 5”:, CO, was admitted directly to the chamber through a fine sintered glass disc and also to the Tyrode’s solution. Transmembrane action potentials were recorded simultaneously from the SA node and the crista terminalis by means of floating microelectrodes. An extracellular His bundle electrogram was recorded through a close bipolar electrode. In the SA node, transmembrane action potentials were recorded from different SA nodal pacemaker cells within an identifiable small area in presence of different test solutions. For measuring atria1 action potential parameters and atria1 conduction time, the atrium was stimulated through bipolar electrodes placed on the right atria1 appendage at a cycle length of 330 ms. The stimulus was a square pulse 2 ms long and twice threshold and was suitably isolated from the ground. Intraatrial conduction was determined as the interval between the upstroke of the atria1 action potential recorded at the upper part of the crista teminalis and the extracellular atria1 electrogram near the AV node. For the measurements of the AV nodal conduction, different preparations were used in which the SA node was excised. Total AV nodal conduction time was determined as the interval between atria1 and His deflection in the extracellular His bundle electrogram at a constant atria1 cycle length (660 ms). This slow stimulation rate was chosen in order to avoid rate-dependent AV nodal block in the presence of hypoxia alone [ 151. The following experimental protocol was used. The preparation was allowed to equilibrate for a 45-min control period before experimentation was initiated. In the first series of experiments, iodoacetic acid was added to the perfusate under aerobic
Glycolysis
and
Nodal
813
Potentials
conditions resulting in three different concentrations of 10e5, 7.5 x 10-s and 3 x 10m3 M. Each preparation was exposed to one single iodoacetic acid concentration for 60 min after which electrophysiological measurements for statistical analysis were repeated. In the second series of experiments, each preparation was initially csposed to one single IAA concentration for a 30-min aerobic period and the combined effects of the drug plus hypoxia were studied after the following 30 min substituting 95’111 N, and 5”(, CO? for the control gas in the presence of the drug; the effects of hypoxia alone were determined using the same experimental protocol but in absence of IAA. The IAA concentrations were chosen since 10m5 M had no significant cffcct, 7.5 i, 10-s M was only effective under hypoxic conditions and 5 x 10m4 M was effective both under aerobic and hypoxic conditions. Measurements of Po2, Pco, and pH were made by withdrawing samples of Tyrode’s solution from the vicinity of the tissue preparation and analyzing them with a pH/gas analyzer (radiometer CIopenhagen). The pH (7.4 & 0.02; n = 8) and PCO, (42 & 1.5 mmHg) remained unchanged with either gas mixture. The PO, was 460 f 25 mmHg in the control solution and fell to 40 & 10 mmHg in presence of 95”,, N, and 5”,, GO?. The effects of 50 mM 2-deoxyglucose were studied in two series of experiments, i.e., under aerobic and anaerobic conditions using the same experimental protocol as with IAA but only one concentration.
Results Effect of IAA
under aerobic and anaerobic conditions on the SA node
Under aerobic conditions, concentrations of 10~~ and 7.5 x 1O-5 M IAA had no significant effect on the sinus rate. However, combination of 7.5 x 1O-5 M IAA with hypoxia caused a rapid decrease in SA nodal spontaneous activity that significantly exceeded the depressant action of hypoxia alone. The results obtained from six experiments are summarized in Figure 1 and typical recordings are shown in Figure 2 demonstrating no significant alterations in the presence of 7.5 x 10~~ M IAA alone but marked bradycardia associated with a shift of the dominant pacemaker site
1 4
5 - Lo9
FIGURE conditions. 40mmHg.
I. Relationship Values are
means
lodwcetote
between sinus rate and IAA & S.D. from six experiments.
3
( M)
concentration under aerobic and hypoxic (0)
PO,
=
460
mmHg;
(0)
PO,
=
814
J. Senges (a)
et al.
(b)
(cl
IOOmV
IS
FIGURE 2. Effect of 7.5 x 1OF M IAA under aerobic and hypoxic conditions on SA nodal and atria1 action potentials and AV nodal conduction. In each panel, the upper trace shows action potentials obtained from the same SA nodal fiber, the middle trace shows action potentials from the same atria1 fiber and the lower trace is a His bundle electrogram. (a) 45-min control conditions; (b) 30 min 7.5 x 10-s M IAA; (c) 20-min combination of 7.5 x 10e5 IAA plus hypoxia. Note marked depression of SA nodal electrical activity but relatively well maintained atria1 action potential amplitude.
during the combined effects of IAA plus hypoxia. After 20 min, the SA nodal cell showed subthreshold oscillations followed by complete stand still of all spontaneous activity at r‘maximum diastolic potentials” ranging between -38 and -47 mV. Superfusion of the isolated SA node with oxygenated Tyrode’s solution containing IAA in a concentration of 5 x lO-4 M decreased the sinus rate by an average of 65% from the control values (Figure 1). The time course of this rate effect showed only moderate changes during the initial 20 min but a marked depression following 30-min exposure to the drug. Recordings of SA nodal action potentials obtained from the same cell at a constant atria1 driving rate before and after administration of 5 x 10e4 M IAA under aerobic conditions are shown in Figure 3. In six experiments, the most striking change was a decrease in action potential amplitude from 72 & 10 to 35 & 13 mV; (P < 0.001) and a slowing of the depolarization rate of the diastolic pacemaker potential that (a)
(b)
200 ms FIGURE 3. Effect of IAA under aerobic conditions (PO, 460 mmHg) on SA nodal and atria1 action potentials and AV nodal conduction at constant atria1 stimulation (180/min). In each section, the upper and middle trace show action potentials from the same respective SA nodal and adjacent atria1 fiber; the lower trace is a His bundle electrogram. (a) 45-min control conditions. (b) 30 min after administration of 5 x lo-’ M IAA; note complete AV nodal block and markedly prolonged sinoatrial conduction preceding complete SA block. Amplification of the lower trace was increasing by a factor of two because of progressive reduction of the atria1 electrogram.
Glycolysis
and Nodal
Potentials
815
was not associated with a significant alteration of the maximum diastolic potential (55 f 8 and 53 f 9 mV; P > 0.05). Retrograde sinoatrial conduction was markedly prolonged in the presence of 5 x 10~~ M IAA (Figure 3) but was difficult to evaluate because of the frequent occurrence of intermittent sinoatrial block that sometimes resulted in bizarre atria1 rhythm at regular sinus rate. A remarkable example is shown in Figure 4 exhibiting regular sinoatrial and intraatrial conduction only during the initial two beats. The third SA nodal impulse is normally conducted to the adjacent atria1 fiber but intraatria1 conduction to the lower right atrium is abruptly blocked. The fourth SA nodal action potential occurs also at a regular interval but the level of conduction block has transiently proceeded to the sinoatrial transition resulting in sinoatrial block. 50 mm
IOOmV
FIGURE 4. Sinoatrial and intraatrial block in the presence of 5 x 10Wp M IAA. Upper, middle and lower trace show action potentials of a SA nodal and an upper right atria1 fiber and the His bundle electrogram. Note regular sequence of SA and intraatrial activation during the initial two beats followed by sudden drop of intraatrial and subsequently of sinoatrial conduction.
Efects of ZAA under aerobic and anaerobic conditions on the right atrium Under aerobic conditions the most prominent effect of 5 x 10e4 M IAA on the action potential of atria1 fibers was a shortening of the repolarization phase (Figure 3). The action potential amplitude and the resting potential remained almost unchanged during the initial 30 min but were significantly decreased during the subsequent period. In six experiments, exposure to 5 x 1O-4 M IAA under aerobic conditions for 60 min resulted in a reduction of action potential duration from 135 f 21 to 83 &- 19 ms (P < 0.001). The action potential amplitude was altered from 98 -& 12 to 75 f 18 mV (P < 0.01) and the resting potential from 85 + 5 to 71 & 13 mV (P < 0.05) associated with intermittent block of the intraatrial conduction in four of the six experiments (Figures 4 and 5). Lower concentrations of IAA (1O-5 and 7.5 x 10m5 M) under aerobic conditions or hypoxia alone in the absence of IAA had no significant effect on the intraatrial
816
J. Senges et al.
conduction but the combination of 7.5 x 10m5 M IAA with hypoxia caused complete intraatrial block and atria1 inexcitability in all experiments (Figure 5). The sequence of electrophysiologic alterations following the combination of the 7.5 x 1O-5 M IAA with hypoxia is indicated in Figure 2 showing that during the initial 20 minutes extreme bradycardia and AV nodal block had already developed but the atria1 action potential amplitude was still relatively well maintained.
I
1 con
/
1
I
5
4
3
- Log
iodoocetote
(M 1
FIGURE 5. Relationship between intraatrial conduction time and IAA concentration under aerobic and hypoxic conditions. Atria1 stimulation rate 180/min. The ordinate is separated in two parts indicating intraatrial conduction time or incidence of intraatrial conduction block. Values are means h S.D. from six experiments. (0) PO, = 40 mmHg; (0) PO, = 460 mmHg.
Effect of IAA
under aerobic and anaerobic conditions on the AV nodal conduction
The alterations of the AV nodal electrical activity were not systematically studied using intracellular recording and various driving rates but some limited results were obtained determining the metabolic effects on the A-H interval in the His bundle electrogram at a constant atria1 rate (901 min). The results obtained from preparations in which the SA node was excised are summarized in Figure 6. Under aerobic conditions, 5 x 1O-4 M IAA caused complete AV nodal block in all experiments; 10~~ and 7.5 x 1O-5 M IAA had no significant effect. Hypoxia alone in the absence of IAA inhibited the AV nodal conduction but no block was induced during the 30 min observation period. During combination of 7.5 x 10e5 M IAA with hypoxia, AV nodal conduction was invariably blocked before all atria1 electrical activity was abolished. Electraphysiologic In order inhibition effects of conditions, 133 * 15
effects of deoxyglucose under aerobic and anaerobic conduction
to substantiate that the electrophysiologic effects of IAA were due to of glycolysis rather than to unspecific blockade of the SH groups, the deoxyglucose were also determined. In six experiments under aerobic deoxyglucose caused no significant change of the sinus rate (control to 125 f 19 beats/min; P > 0.05) and of the SA nodal action potential
Glycolysis
and
Nodal
Potentials
- Log iodocetote
CM i
FIGURE 6. Relationship between AV nodal conduction and IAA concentration hypoxic conditions. Atria1 stimulation rate SO/min. The ordinate is separated AV nodal conduction time or incidence of AV nodal block. Values are means ments. (0) PO, = 40 mmHg; (0) PO, = 460 mmHg.
817
under aerobic and in two parts indicating i SD. from six experi-
amplitude (78 + 11 to 76 & 12 mV; P > 0.05). Typical recordings obtained from the same respective SA nodal and atria1 fibers before and after deoxyglucose are shown in Figure 7. The only significant alteration was a small reduction of the atria1 action potential duration (126 & 19 to 108 f 20 ms; P < 0.01) associated with no significant change of the atria1 action potential amplitude (102 f 5.2 to 100 i 8.1 mV; P > 0.05). The AV nodal conduction time remained unchanged (65 5 15 to 68 & 2 1 ms; P > 0.05). The effects of deoxyglucose on atria1 and lower AV nodal action potential and on the A-H interval in the His bundle electrogram are demonstrated in Figure 8. Combination of deoxyglucose with hypoxia in six additional experiments resulted in complete inexcitability of the preparation within 30 min [Figure 8(c)]. The sequence of events resembled those obtained with the combined effects of 7.5 x 1O-5 M IAA plus hypoxia: during an initial period, complete AV nodal block rapidly
IS FIGURE 7. Effect of 50 rnM Z-deoxyglucose under aerobic and hypoxic conditions on SA nodal and atria1 action potentials and AV nodal conduction. In each panel, the upper trace shows action potentials obtained from the same SA nodal fiber, the middle trace shows action potentials obtained from the same atria1 fiber and the lower trace is a His bundle electrogram. (a) 45-min control conditions; (b) 30-min deoxyglucose; (c) 18-min combination of deoxyglucose and hypoxia. Note complete AV nodal block and almost abolished SA nodal potentials but still preserved atria1 action potentials. M.C.C.
21
818
J. Senges
et
al.
developed associated with marked atria1 bradycardia and depression of the SA nodal action potential amplitude. Simultaneously, the atria1 action potential duration was shortened but the atria1 action potential amplitude remained almost unchanged (Figure 7). Subsequently, the atria1 action potential amplitude was also reduced resulting in abolition of all electrical activity. (0 1
(b)
FIGURE 8. Effect of 50 rn~ Z-deoxyglucose under aerobic and hypoxic conditions on atria1 and lower (NH) AV nodal action potentials and on AV nodal conduction at constant atria1 stimulation (90/min). In each panel, the upper and middle trace shows action potentials from the same respective atria1 and lower AV nodal fiber; the lower trace is a His bundle electrogram. (a) 45-min control conditions; (b) 30-min deoxyglucose; (c) (slower time base), 30-min combination of deoxyglucose plus hypoxia. Note that under aerobic conditions deoxyglucose induces a small reduction of the plateau phase of both the atria1 and the nodal action potential and that complete inexcitability develops following combination with hypoxia (marked stimulus artefacts due to increased stimulation strength).
Discussion The present results show that under aerobic conditions the glycolytic inhibitors iodoacetate and deoxyglucose at respective concentrations of 7.5 x 10e5 and 5 x 10~~ M have no significant effect on cardiac automaticity and conduction. Under hypoxic conditions, iodoacetate and deoxyglucose at the respective concentrations induce (1) early electrophysiological changes including complete depression of SA or AV nodal function and marked shortening of the atria1 action potential duration and (2) more delayed effects including reduction of atria1 action potential amplitude and block of intraatrial conduction. The effects of higher IAA concentrations (5 x lo-* M) under aerobic conditions resemble those obtained with lower concentrations (7.5 x 10-s M IAA) combined with hypoxia.
Glycolysis
and
Nodal
Potentials
81’1
Iodoacetate is well known to inhibit the formation of p)-ruvate from gly-cogen by non-competitive inhibition of triophosphate but this drug is not a specific inhibitor of glycolysis in mammalian cardiac muscle and even in low concentrations it is likely that other enzyme systems are affcctcd [20]. Although in the present expcrimerits rather low concentrations of IAA were used, some unspecific inhibition of various metabolic pathway particularly in the presence of 5 x lo-” M IAA cannot be excluded. In contrast, deoxyglucose appears to be a specific and competitive inhibitor interacting with glycolysis at the phosphoglucoisomerase level 12’2 1. ‘T‘llc possibility that the depressant action of 50 nlM deoxyglucose was due to incrrascd osmolarity rather than to inhibition of glycolysis can be excluded since rrcc‘nt studies have shown that in the presence of hypoxia 50 mM glucose markedly improved the nodal electrical activity and 50 IYIM sucrose had no significant effect [17]. l’hat the upstroke of the action potential in SA and A\’ nodal fibers depends on a transmembrane slow inward current has been suggested by several authors [!/, 2.7, 251. Recent experiments have demonstrated a close relationship betbvccn hyposic nodal function and the extracellular glucose concentration supporting the hypoth& that the SA and AV nodal cells can utilize energy for production and maintenance of slow inward current-dependent electrical activity from both aerobic or anaerobic glycolytic metabolism [17]. The relevance of these experimental results to the human situation was strongly supported by clinical observations in patients with SA and/or AV nodal dysfunction who developed marked improvement of nodal aulomaticity and conduction after increase in extracellular glucose concentration [Ih‘]. The present data are in general agreement with this hypothesis suggesting that in the normoxic heart the nodal electrical activity is predominantly related to aerobic metabolism and is little or not affected by moderate inhibition of glycolysis. However, during selective administration of hypoxia some more or less depressed nodal function was still preserved and complete deterioration of all nodal electrical activity occurred exclusively following inhibition of both aerobic plus glycolytic pathways. The results obtained under hypoxic conditions support the suggestion that glycolytic energy production may effectively contribute to the maintenance of nodal electrical activity. The fast Na system-dependent atria1 action potential amplitude and the atria1 conduction was relatively long maintained even during IAA at high concentrations (5 :< 10m4 M) or during the combined effects of hypoxia plus 7.5 x 1OP” M lA.4. For comparison, the atria1 action potential plateau was much more sensitive to such metabolic interventions. In papillary muscle fibers, the hypoxic alterations of membrane properties have been proposed to result from an increased potassium conductance leading to an increased rate of repolarization [4, 18, 211. Howevcsr, more recent results have suggested that anaerobic glycolysis is involved in the rrgulation of the slow inward current rather than of the potassium conductance [3, 10, II, 131. In the present experiments, metabolically-induced inhibition of the slow inward current could well explain not only the depression of the nodal electrical activity but also the reduction of the atria1 action potential plateau. The contrast between maintained atria1 action potential amplitude but depressed nodal electrical activity can be explained either by differences in metabolic energy production of the various cell types or by differences in the metabolic regulation of the various ionic mechanisms. Since the margin of safety for generation and conduction of the fast Na-dependent electrical activity is very high as compared with slog inward current-dependent electrical activity [?I, this discrepancy in intrinsic generator properties of the local membrane could result in a different sensitivity of
820
J. Senges et al.
the two inward current systems to inhibition of aerobic or glycolytic metabolism. However, the delayed abolition of atria1 excitability during the combined effects of hypoxia plus glycolytic inhibitors suggests that the fast Na system was also related to the metabolic state of atria1 fibers. Sinoatrial block resembling type Mobitz II was exclusively obtained with 5 x 1O-4 M IAA but never with hypoxia and merits particular comment. This discrepancy could be explained by the fact that 5 x 1O-4 M IAA resulted in depression of both nodal and atria1 electrical activity whereas anoxia inhibited rather selectively the SA nodal function and failed to significantly affect the atria1 conduction. Recently it has been speculated that “all or none” behaviour of Mobitz type II SA block suggests that atria1 excitation may be occurring via a limited narrow path [6]. It is interesting to note that IAA-induced SA block type Mobitz was closely related to the simultaneous occurrence of conduction block within the atrium. Acknowledgement
We are grateful
to Mrs Katharina
Marquard
for technical
assistance.
REFERENCES 1.
A. Synthetic interstitial fluid for isolated mammalian tissue. Life Sciences 8, (1969) VAN CAPELLE, F. J. L. & JANSE, M. J. Influence of geometry on the shape of the propagated action potential. In The Conduction System of the Heart. H. J. J, Wellens, Ki Lie & M. J. Janse, Eds, pp. 316-335. H. E. Stanfert Kroese: Bv Leiden (1976). CARMELIET, E. Cardiac transmembrane potentials and metabolism. Circulation Research 42, 577-587 (1978). DE MELLO, W. C. Metabolism and electrical activity of the heart: action of 2,4-dinitrophenol and ATP. American Journal of Physiology 196, 377-380 (1959). DIXON, M. Action of iodoacetate in dehydrogenase and alcoholic fermation. Nature 140, 806 (1937). GOLDREYER, B. N. Sinus node dysfunction: A physiologic consideration of arrhythmias involving the sinus node. In Complex Electrocardiography 2. G. Fisch, Ed. Philadelphia: F. A. Davis (1974). HAAS, H. G., KERN, R. & EINW~~CHTER, H. M. Electrical activity and metabolism in cardiac tissue: An experimental and theoretical study. Journal of Membrane Biology 3, 180-209 (1970). KOHLHARDT, M., MNICH, Z. & MAIER, G. Alterations of the excitation process of the sionatrial pacemaker cell in the presence of anoxia and metabolic inhibitors. Journal of Molecular and Cellular Cardiology 9, 477-488 (1977). Lu, H. H. & BROOKS, C. MC. C. Role of calcium in cardiac pacemaker cell action. (Abstr.) Bulletin of the New York Academy Medicine 45, 100 (1969). MCDONALD, T. F. & MACLEOD, D. P. Effects of manganese, glucose and isoprenaline on the action potential of anoxia ventricular muscle. Naunyn-Schmiedebergs Archiv fiir Pharmakologie und Experimentelle Pathologie 275, 169-181 (1972). MCDONALD, T. F. & MACLEOD, D. P. Metabolism and electrical activity of anoxic ventricular muscle. Journal of Physiology 229, 559-582 (1973). PAES DE CARVALHO, A., DE MELLO, W. C. & HOFFMAN, B. F. Electrophysiological evidence for specialized fiber types in atrium. American Journal of Physiology 196, 483-488 (1959). PRASAD, M. D. & MACLEOD, D. P. Influence of glucose on the transmembrane action potential of guinea-pig papillary muscle. Metabolic inhibitors, ouabain, and calcium chloride, and their interaction with glucose, sympathomimetic amines, and aminophylline. Circulation Research 14, 939-950 (1969). BRETAG,
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SENGES, J., BRACHMANN, J., PELZER, D., MIZUTANI, T. & K~~BLER, W. Effects of some components of ischemia on electrical activity and re-entry in the ventricular conducting system. Circulation Research 44, 864-872 (1979). SENGES, J., MIZUTANI, T., PELZER, D., BRACHMANN, J., SONNHOF, U. & K~~BLER, \V. Effect of hypoxia on the sinoatrial node, atrium and atrioventricular node in the rabbit heart. Circulation Research 44, 856-863 (1979). SENGES, J., PELZER, D., BRACHMANN, J., KRAMER, B. & K~BLER, W. Glucose-insulin potassium in bradyarrhythmias (abstr.). Transactions of the Eurobean SocieQ of Cardiolog, 1, 69 (1978). SENGES, J., BRACHMANN, J., PELZER, D., KRAMER, B. & K~BLER, W. Combined effects of glucose and hypoxia on cardiac automaticity and conduction. Journal of Alolecular and Cellular Cardiology 12, 3 1 l-323 ( 1980). TRAUTWEIN, W. & DC’DEL, J. Aktionspotential und Kontraktion des Herzmuskels in Sauerstoffmangel. PJliigers Archio 263, 23-32 (1956). WEBB, J. L. Actions of metabolic substrates and inhibitors on the rabbit auricle. Britirh Journal of Pharmacology 5, 87-l 18 (1950). WEBB, J. L. Enzyme and Metabolic Inhibitors, 3, pp. 18-51. New York: Academic Press (1966). WEBB, J. L. & HOLLANDER, P. B. Metabolic aspects of the relationship between the contractility and membrane potentials of the rat atrium. Circulation Research 4, 618-626 (1956). WICII, A. N., DRURY, D. R., NAKADA, H. I. & WOLFE, J. B. Localization of the primary metabolic block produced by 2-deoxyglucose. Journal of Biological Chemistry 224, 963-969 (1957). WIT, A. L. & CRANEFIELD, P. F. Effect of verapamil on the sinoatrial and atrioventricular nodes of the rabbit and the mechanism by which it arrests reentrant atrioventricular nodal tachycardia. C’irculation Research 35, 4 13-425 (1974). YANG, W. C. Anaerobic functional activity of isolated rabbit atria. American Journai of Physiolop 205, 78 l-784 ( 1963). ZIPES, D. P., BESCH, H. R. & FYATANABE, A. hf. Role of the slow current in cardiac elrctrophysiology. Circulation 51, 761-766 (1975).
of Molecular
and Cellular
Effect Atrium Jochen
Cardiology
(1981)
13,
811-821
of Glycolytic Inhibitors and Atrioventricular Senges,
Johannes and
Abteilung
Innere Medizin
on the Sinoatrial Node in the Rabbit
Brachmann, Wolfgang
Dieter Kiibler
Pelzer,
III (h*urdiologie), Medizinische 69 Heidelberg, Germany
(Received 23 June 1980, accepted in revisedform
Node, Heart*
Ioannis
Rizos
C:niversitiitsklinik, 12 May 1981)
J. SENGES, J.
BRACHMANN, D. PELZER, I. Rrzos AND W. K~~BLER. Effect of Glycolytic Inhibitors on the Sinoatrial Node, Atrium and Atrioventricular Node in the Rabbit Heart. Journal of Molecular and Cellular Cardiology (1981) 13, 811-82 1. The effect of unspecific noncompetitive (iodoacetir acid ; IA:\) and of specific competitive (deoxyglucose) glycolytic inhibitors were studied under aerobic and anaerobic conditions in the isolated sinoatrial ISA) node, right atrium and atriovcntricular (A\‘) node of the rabbit heart. Trammembrane action potentials were recorded simultaneously with the His bundle rlectrogram. Under aerobic conditions (PO, 460 mmHg), 7.5 x 10-j Y IAA caused no significant alterations of sinus rate, intraatrial and AV nodal conduction. Under hypoxic conditions (PO, 40 mmHg), 7.5 ./ 10-j M IAA resulted in rapid and complete abolition of the SA and A\’ nodal electrical activity and in a more delayed atria1 inexcitability. Similar changes were never observed in the presence of hypoxia alone. Higher concentrations of IA.4 (5 Y 1O-1 M) severely depressed the sinus rate and the AV nodal conduction already under aerobic conditions. The action potential amplitude and the rate of diastolic depolarization of the SA nodal fibers was significantly reduced, the maximum diastolic potential remained unchanged. In atria1 fibers, 5 x 1OV M IAA cattsed predominantly shortening of the action potential duration but had less marked and delayed depressant effects on the action potential amplitude. Elrctrophysiologic abnormalities included MobitL type II sinoatrial block. The electrophysiological effects of deoxyglucose (50 mM) under aerobic and hypoxic conditions resembled those obtained with 7.5 x 10e5 M IAA. The results suggest that the “normal” nodal function is predominantly related to aerobic metabolism but that under hypoxic conditions glycolytic energy production can effectively contribute to the maintenance of nodal elec,triral activity. Also that the specific relation between cardiac metabolism and electrical activity of nodal and atria1 cells may at least partly be explained by the particular electrical membrane properties of the various fiber types. KEY
IVORDS:
action
potential.
Iodoacetic
acid;
Deoxyglucose;
Anoxia;
Glycolysis;
Slow
response;
Nodal
Introduction Knowledge of the relationship between cardiac muscle metabolism function has been advanced by many authors to an appreciable metabolic processes involved in the maintenance and production activity of the heart including automaticity and conduction have been studied [7, 8, 18-21, 241. The present experiments were derived results suggesting that hypoxia inhibits predominantly the sinoatrial ventricular (AV) nodal transmembrane action potentials dependent * This work was supported vascullres System”. This work fur Kreislaufforschung, Bad International Society for Heart 0022%2828/8
l/O9081
1 + 11
by the Deutsche Forschungsgemeinschaft was presented in part at the 1979 meeting Nauheim and at the 1979 meeting of Research, Dijon.
$02.00/O
0
1981
Academic
and contractile extent but the of electrical less extensively from previous (SA) and atrioon the slow
within the SFB 90 “Cardioof the Deutsche Gessellschaft the European Section of the Press
Inc.
(London)
Limited
812
J. Senges
et
al.
inward current but has only little, if any effect on atria1 and ventricular specialized conduction related to the rapid Na current [14, 151. The contribution of glycolytic energy metabolism to the maintenance of nodal electrical activity is strongly supported by recent clinical and experimental observations demonstrating a close relation between SA and AV nodal function and the extracellular glucose concentration [16, 171. The purpose of the present experiments was to study the combined effects of glycolytic inhibitors and hypoxia on various electrophysiological parameters of cardiac automaticity and conduction. Iodoacetic acid (IAA) in low concentrations produces block of the Embden-Meyerhof glycolytic chain by inhibiting triophosphate dehydrogenase and exerts general sulfhydryl inhibiting effects at higher concentrations [.5, 201. In order to demonstrate that the electrophysiologic alterations observed in the presence of IAA were due to inhibition of glycolysis rather than to unspecific blockade of SH groups, the effects of deoxyglucose were also tested. Deoxyglucose has been reported to induce block of glucose catabolism by competitive inhibition of deoxyglucose-6-PO, on glucose-6-PO, at the phosphoglucoisomerase level [22]. Materials
and
Methods
Rabbits weighing 2 to 3 kg were killed by a single blow. The heart was rapidly removed and dissected in an oxygenated Tyrode’s solution of the following composition (mM) [I]: NaCl, 107.7; KCl, 3.48; CaCl,, 1.53; MgSO,, 0.69; NaHCO,, 26.2; NaH,PO,, 1.67; Na gluconate, 9.64; glucose, 5.5; sucrose, 7.6. The preparation consisting of sinoatrial node, crista terminalis, pectinate muscles of the atria1 appendage, intraatrial septum, coronary sinus, atrioventricular node and His bundle [12] was mounted in a I5 ml tissue bath and perfused at a rate of 30 ml/min with Tyrode’s solution at 36 f 0.5% A mixture of 95O;, 0, and 5”:, CO, was admitted directly to the chamber through a fine sintered glass disc and also to the Tyrode’s solution. Transmembrane action potentials were recorded simultaneously from the SA node and the crista terminalis by means of floating microelectrodes. An extracellular His bundle electrogram was recorded through a close bipolar electrode. In the SA node, transmembrane action potentials were recorded from different SA nodal pacemaker cells within an identifiable small area in presence of different test solutions. For measuring atria1 action potential parameters and atria1 conduction time, the atrium was stimulated through bipolar electrodes placed on the right atria1 appendage at a cycle length of 330 ms. The stimulus was a square pulse 2 ms long and twice threshold and was suitably isolated from the ground. Intraatrial conduction was determined as the interval between the upstroke of the atria1 action potential recorded at the upper part of the crista teminalis and the extracellular atria1 electrogram near the AV node. For the measurements of the AV nodal conduction, different preparations were used in which the SA node was excised. Total AV nodal conduction time was determined as the interval between atria1 and His deflection in the extracellular His bundle electrogram at a constant atria1 cycle length (660 ms). This slow stimulation rate was chosen in order to avoid rate-dependent AV nodal block in the presence of hypoxia alone [ 151. The following experimental protocol was used. The preparation was allowed to equilibrate for a 45-min control period before experimentation was initiated. In the first series of experiments, iodoacetic acid was added to the perfusate under aerobic
Glycolysis
and
Nodal
813
Potentials
conditions resulting in three different concentrations of 10e5, 7.5 x 10-s and 3 x 10m3 M. Each preparation was exposed to one single iodoacetic acid concentration for 60 min after which electrophysiological measurements for statistical analysis were repeated. In the second series of experiments, each preparation was initially csposed to one single IAA concentration for a 30-min aerobic period and the combined effects of the drug plus hypoxia were studied after the following 30 min substituting 95’111 N, and 5”(, CO? for the control gas in the presence of the drug; the effects of hypoxia alone were determined using the same experimental protocol but in absence of IAA. The IAA concentrations were chosen since 10m5 M had no significant cffcct, 7.5 i, 10-s M was only effective under hypoxic conditions and 5 x 10m4 M was effective both under aerobic and hypoxic conditions. Measurements of Po2, Pco, and pH were made by withdrawing samples of Tyrode’s solution from the vicinity of the tissue preparation and analyzing them with a pH/gas analyzer (radiometer CIopenhagen). The pH (7.4 & 0.02; n = 8) and PCO, (42 & 1.5 mmHg) remained unchanged with either gas mixture. The PO, was 460 f 25 mmHg in the control solution and fell to 40 & 10 mmHg in presence of 95”,, N, and 5”,, GO?. The effects of 50 mM 2-deoxyglucose were studied in two series of experiments, i.e., under aerobic and anaerobic conditions using the same experimental protocol as with IAA but only one concentration.
Results Effect of IAA
under aerobic and anaerobic conditions on the SA node
Under aerobic conditions, concentrations of 10~~ and 7.5 x 1O-5 M IAA had no significant effect on the sinus rate. However, combination of 7.5 x 1O-5 M IAA with hypoxia caused a rapid decrease in SA nodal spontaneous activity that significantly exceeded the depressant action of hypoxia alone. The results obtained from six experiments are summarized in Figure 1 and typical recordings are shown in Figure 2 demonstrating no significant alterations in the presence of 7.5 x 10~~ M IAA alone but marked bradycardia associated with a shift of the dominant pacemaker site
1 4
5 - Lo9
FIGURE conditions. 40mmHg.
I. Relationship Values are
means
lodwcetote
between sinus rate and IAA & S.D. from six experiments.
3
( M)
concentration under aerobic and hypoxic (0)
PO,
=
460
mmHg;
(0)
PO,
=
814
J. Senges (a)
et al.
(b)
(cl
IOOmV
IS
FIGURE 2. Effect of 7.5 x 1OF M IAA under aerobic and hypoxic conditions on SA nodal and atria1 action potentials and AV nodal conduction. In each panel, the upper trace shows action potentials obtained from the same SA nodal fiber, the middle trace shows action potentials from the same atria1 fiber and the lower trace is a His bundle electrogram. (a) 45-min control conditions; (b) 30 min 7.5 x 10-s M IAA; (c) 20-min combination of 7.5 x 10e5 IAA plus hypoxia. Note marked depression of SA nodal electrical activity but relatively well maintained atria1 action potential amplitude.
during the combined effects of IAA plus hypoxia. After 20 min, the SA nodal cell showed subthreshold oscillations followed by complete stand still of all spontaneous activity at r‘maximum diastolic potentials” ranging between -38 and -47 mV. Superfusion of the isolated SA node with oxygenated Tyrode’s solution containing IAA in a concentration of 5 x lO-4 M decreased the sinus rate by an average of 65% from the control values (Figure 1). The time course of this rate effect showed only moderate changes during the initial 20 min but a marked depression following 30-min exposure to the drug. Recordings of SA nodal action potentials obtained from the same cell at a constant atria1 driving rate before and after administration of 5 x 10e4 M IAA under aerobic conditions are shown in Figure 3. In six experiments, the most striking change was a decrease in action potential amplitude from 72 & 10 to 35 & 13 mV; (P < 0.001) and a slowing of the depolarization rate of the diastolic pacemaker potential that (a)
(b)
200 ms FIGURE 3. Effect of IAA under aerobic conditions (PO, 460 mmHg) on SA nodal and atria1 action potentials and AV nodal conduction at constant atria1 stimulation (180/min). In each section, the upper and middle trace show action potentials from the same respective SA nodal and adjacent atria1 fiber; the lower trace is a His bundle electrogram. (a) 45-min control conditions. (b) 30 min after administration of 5 x lo-’ M IAA; note complete AV nodal block and markedly prolonged sinoatrial conduction preceding complete SA block. Amplification of the lower trace was increasing by a factor of two because of progressive reduction of the atria1 electrogram.
Glycolysis
and Nodal
Potentials
815
was not associated with a significant alteration of the maximum diastolic potential (55 f 8 and 53 f 9 mV; P > 0.05). Retrograde sinoatrial conduction was markedly prolonged in the presence of 5 x 10~~ M IAA (Figure 3) but was difficult to evaluate because of the frequent occurrence of intermittent sinoatrial block that sometimes resulted in bizarre atria1 rhythm at regular sinus rate. A remarkable example is shown in Figure 4 exhibiting regular sinoatrial and intraatrial conduction only during the initial two beats. The third SA nodal impulse is normally conducted to the adjacent atria1 fiber but intraatria1 conduction to the lower right atrium is abruptly blocked. The fourth SA nodal action potential occurs also at a regular interval but the level of conduction block has transiently proceeded to the sinoatrial transition resulting in sinoatrial block. 50 mm
IOOmV
FIGURE 4. Sinoatrial and intraatrial block in the presence of 5 x 10Wp M IAA. Upper, middle and lower trace show action potentials of a SA nodal and an upper right atria1 fiber and the His bundle electrogram. Note regular sequence of SA and intraatrial activation during the initial two beats followed by sudden drop of intraatrial and subsequently of sinoatrial conduction.
Efects of ZAA under aerobic and anaerobic conditions on the right atrium Under aerobic conditions the most prominent effect of 5 x 10e4 M IAA on the action potential of atria1 fibers was a shortening of the repolarization phase (Figure 3). The action potential amplitude and the resting potential remained almost unchanged during the initial 30 min but were significantly decreased during the subsequent period. In six experiments, exposure to 5 x 1O-4 M IAA under aerobic conditions for 60 min resulted in a reduction of action potential duration from 135 f 21 to 83 &- 19 ms (P < 0.001). The action potential amplitude was altered from 98 -& 12 to 75 f 18 mV (P < 0.01) and the resting potential from 85 + 5 to 71 & 13 mV (P < 0.05) associated with intermittent block of the intraatrial conduction in four of the six experiments (Figures 4 and 5). Lower concentrations of IAA (1O-5 and 7.5 x 10m5 M) under aerobic conditions or hypoxia alone in the absence of IAA had no significant effect on the intraatrial
816
J. Senges et al.
conduction but the combination of 7.5 x 10m5 M IAA with hypoxia caused complete intraatrial block and atria1 inexcitability in all experiments (Figure 5). The sequence of electrophysiologic alterations following the combination of the 7.5 x 1O-5 M IAA with hypoxia is indicated in Figure 2 showing that during the initial 20 minutes extreme bradycardia and AV nodal block had already developed but the atria1 action potential amplitude was still relatively well maintained.
I
1 con
/
1
I
5
4
3
- Log
iodoocetote
(M 1
FIGURE 5. Relationship between intraatrial conduction time and IAA concentration under aerobic and hypoxic conditions. Atria1 stimulation rate 180/min. The ordinate is separated in two parts indicating intraatrial conduction time or incidence of intraatrial conduction block. Values are means h S.D. from six experiments. (0) PO, = 40 mmHg; (0) PO, = 460 mmHg.
Effect of IAA
under aerobic and anaerobic conditions on the AV nodal conduction
The alterations of the AV nodal electrical activity were not systematically studied using intracellular recording and various driving rates but some limited results were obtained determining the metabolic effects on the A-H interval in the His bundle electrogram at a constant atria1 rate (901 min). The results obtained from preparations in which the SA node was excised are summarized in Figure 6. Under aerobic conditions, 5 x 1O-4 M IAA caused complete AV nodal block in all experiments; 10~~ and 7.5 x 1O-5 M IAA had no significant effect. Hypoxia alone in the absence of IAA inhibited the AV nodal conduction but no block was induced during the 30 min observation period. During combination of 7.5 x 10e5 M IAA with hypoxia, AV nodal conduction was invariably blocked before all atria1 electrical activity was abolished. Electraphysiologic In order inhibition effects of conditions, 133 * 15
effects of deoxyglucose under aerobic and anaerobic conduction
to substantiate that the electrophysiologic effects of IAA were due to of glycolysis rather than to unspecific blockade of the SH groups, the deoxyglucose were also determined. In six experiments under aerobic deoxyglucose caused no significant change of the sinus rate (control to 125 f 19 beats/min; P > 0.05) and of the SA nodal action potential
Glycolysis
and
Nodal
Potentials
- Log iodocetote
CM i
FIGURE 6. Relationship between AV nodal conduction and IAA concentration hypoxic conditions. Atria1 stimulation rate SO/min. The ordinate is separated AV nodal conduction time or incidence of AV nodal block. Values are means ments. (0) PO, = 40 mmHg; (0) PO, = 460 mmHg.
817
under aerobic and in two parts indicating i SD. from six experi-
amplitude (78 + 11 to 76 & 12 mV; P > 0.05). Typical recordings obtained from the same respective SA nodal and atria1 fibers before and after deoxyglucose are shown in Figure 7. The only significant alteration was a small reduction of the atria1 action potential duration (126 & 19 to 108 f 20 ms; P < 0.01) associated with no significant change of the atria1 action potential amplitude (102 f 5.2 to 100 i 8.1 mV; P > 0.05). The AV nodal conduction time remained unchanged (65 5 15 to 68 & 2 1 ms; P > 0.05). The effects of deoxyglucose on atria1 and lower AV nodal action potential and on the A-H interval in the His bundle electrogram are demonstrated in Figure 8. Combination of deoxyglucose with hypoxia in six additional experiments resulted in complete inexcitability of the preparation within 30 min [Figure 8(c)]. The sequence of events resembled those obtained with the combined effects of 7.5 x 1O-5 M IAA plus hypoxia: during an initial period, complete AV nodal block rapidly
IS FIGURE 7. Effect of 50 rnM Z-deoxyglucose under aerobic and hypoxic conditions on SA nodal and atria1 action potentials and AV nodal conduction. In each panel, the upper trace shows action potentials obtained from the same SA nodal fiber, the middle trace shows action potentials obtained from the same atria1 fiber and the lower trace is a His bundle electrogram. (a) 45-min control conditions; (b) 30-min deoxyglucose; (c) 18-min combination of deoxyglucose and hypoxia. Note complete AV nodal block and almost abolished SA nodal potentials but still preserved atria1 action potentials. M.C.C.
21
818
J. Senges
et
al.
developed associated with marked atria1 bradycardia and depression of the SA nodal action potential amplitude. Simultaneously, the atria1 action potential duration was shortened but the atria1 action potential amplitude remained almost unchanged (Figure 7). Subsequently, the atria1 action potential amplitude was also reduced resulting in abolition of all electrical activity. (0 1
(b)
FIGURE 8. Effect of 50 rn~ Z-deoxyglucose under aerobic and hypoxic conditions on atria1 and lower (NH) AV nodal action potentials and on AV nodal conduction at constant atria1 stimulation (90/min). In each panel, the upper and middle trace shows action potentials from the same respective atria1 and lower AV nodal fiber; the lower trace is a His bundle electrogram. (a) 45-min control conditions; (b) 30-min deoxyglucose; (c) (slower time base), 30-min combination of deoxyglucose plus hypoxia. Note that under aerobic conditions deoxyglucose induces a small reduction of the plateau phase of both the atria1 and the nodal action potential and that complete inexcitability develops following combination with hypoxia (marked stimulus artefacts due to increased stimulation strength).
Discussion The present results show that under aerobic conditions the glycolytic inhibitors iodoacetate and deoxyglucose at respective concentrations of 7.5 x 10e5 and 5 x 10~~ M have no significant effect on cardiac automaticity and conduction. Under hypoxic conditions, iodoacetate and deoxyglucose at the respective concentrations induce (1) early electrophysiological changes including complete depression of SA or AV nodal function and marked shortening of the atria1 action potential duration and (2) more delayed effects including reduction of atria1 action potential amplitude and block of intraatrial conduction. The effects of higher IAA concentrations (5 x lo-* M) under aerobic conditions resemble those obtained with lower concentrations (7.5 x 10-s M IAA) combined with hypoxia.
Glycolysis
and
Nodal
Potentials
81’1
Iodoacetate is well known to inhibit the formation of p)-ruvate from gly-cogen by non-competitive inhibition of triophosphate but this drug is not a specific inhibitor of glycolysis in mammalian cardiac muscle and even in low concentrations it is likely that other enzyme systems are affcctcd [20]. Although in the present expcrimerits rather low concentrations of IAA were used, some unspecific inhibition of various metabolic pathway particularly in the presence of 5 x lo-” M IAA cannot be excluded. In contrast, deoxyglucose appears to be a specific and competitive inhibitor interacting with glycolysis at the phosphoglucoisomerase level 12’2 1. ‘T‘llc possibility that the depressant action of 50 nlM deoxyglucose was due to incrrascd osmolarity rather than to inhibition of glycolysis can be excluded since rrcc‘nt studies have shown that in the presence of hypoxia 50 mM glucose markedly improved the nodal electrical activity and 50 IYIM sucrose had no significant effect [17]. l’hat the upstroke of the action potential in SA and A\’ nodal fibers depends on a transmembrane slow inward current has been suggested by several authors [!/, 2.7, 251. Recent experiments have demonstrated a close relationship betbvccn hyposic nodal function and the extracellular glucose concentration supporting the hypoth& that the SA and AV nodal cells can utilize energy for production and maintenance of slow inward current-dependent electrical activity from both aerobic or anaerobic glycolytic metabolism [17]. The relevance of these experimental results to the human situation was strongly supported by clinical observations in patients with SA and/or AV nodal dysfunction who developed marked improvement of nodal aulomaticity and conduction after increase in extracellular glucose concentration [Ih‘]. The present data are in general agreement with this hypothesis suggesting that in the normoxic heart the nodal electrical activity is predominantly related to aerobic metabolism and is little or not affected by moderate inhibition of glycolysis. However, during selective administration of hypoxia some more or less depressed nodal function was still preserved and complete deterioration of all nodal electrical activity occurred exclusively following inhibition of both aerobic plus glycolytic pathways. The results obtained under hypoxic conditions support the suggestion that glycolytic energy production may effectively contribute to the maintenance of nodal electrical activity. The fast Na system-dependent atria1 action potential amplitude and the atria1 conduction was relatively long maintained even during IAA at high concentrations (5 :< 10m4 M) or during the combined effects of hypoxia plus 7.5 x 1OP” M lA.4. For comparison, the atria1 action potential plateau was much more sensitive to such metabolic interventions. In papillary muscle fibers, the hypoxic alterations of membrane properties have been proposed to result from an increased potassium conductance leading to an increased rate of repolarization [4, 18, 211. Howevcsr, more recent results have suggested that anaerobic glycolysis is involved in the rrgulation of the slow inward current rather than of the potassium conductance [3, 10, II, 131. In the present experiments, metabolically-induced inhibition of the slow inward current could well explain not only the depression of the nodal electrical activity but also the reduction of the atria1 action potential plateau. The contrast between maintained atria1 action potential amplitude but depressed nodal electrical activity can be explained either by differences in metabolic energy production of the various cell types or by differences in the metabolic regulation of the various ionic mechanisms. Since the margin of safety for generation and conduction of the fast Na-dependent electrical activity is very high as compared with slog inward current-dependent electrical activity [?I, this discrepancy in intrinsic generator properties of the local membrane could result in a different sensitivity of
820
J. Senges et al.
the two inward current systems to inhibition of aerobic or glycolytic metabolism. However, the delayed abolition of atria1 excitability during the combined effects of hypoxia plus glycolytic inhibitors suggests that the fast Na system was also related to the metabolic state of atria1 fibers. Sinoatrial block resembling type Mobitz II was exclusively obtained with 5 x 1O-4 M IAA but never with hypoxia and merits particular comment. This discrepancy could be explained by the fact that 5 x 1O-4 M IAA resulted in depression of both nodal and atria1 electrical activity whereas anoxia inhibited rather selectively the SA nodal function and failed to significantly affect the atria1 conduction. Recently it has been speculated that “all or none” behaviour of Mobitz type II SA block suggests that atria1 excitation may be occurring via a limited narrow path [6]. It is interesting to note that IAA-induced SA block type Mobitz was closely related to the simultaneous occurrence of conduction block within the atrium. Acknowledgement
We are grateful
to Mrs Katharina
Marquard
for technical
assistance.
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