Influence of Changes in pH on
the Mechanical Activity of Cardiac Muscle
By Hrvoje Lorkovic, Ph.D.
Downloaded from http://circres.ahajournals.org/ by guest on June 17, 2017
ABSTRACT
In a comprehensive experimental study, the influence of changes in extraand intracellular pH on the mechanical and electrical activity of frog heart
ventricle preparations has been investigated. The twitch tension and potassium
contracture tension were decreased in acid solutions, provided that the concentration of calcium or the frequency of stimulation was low. In solutions
with a high calcium concentration, prolonged exposure to acid leads to an
increase of the potassium contractures. The electrical activity, measured by
intracellular microelectrodes, is abolished in acid solutions if the change of
solution in the extracellular space is enforced by perpendicular jet superfusion;
with slow perfusion it is well maintained except when [Ca] is high. The
lowering of intracellular pH by applying a solution saturated with 80% CO2
while keeping the extracellular pH constant causes a rapid drop in twitch
tension followed by partial recovery. The relaxation phase of single twitches
is prolonged and potassium contractures are markedly increased after the
high CO2 has acted for several minutes. When both intra- and extracellular
pH are decreased, both twitches and potassium contractures are markedly
decreased. The electrical activity is maintained. Some of the effects of variations in pH are discussed in terms of changes in the processes of mechanical
activation and inactivation.
ADDITIONAL KEY WORDS
heart contraction
concentration
increased Pco 2
potassium contracture
• Contractions of cardiac muscle are decreased in acid solutions (1). Extracellular
electrograms (2) have shown that the electrical activity persists in moderately acid solutions. More recently, the excitability of
Purldnje fibers of sheep heart was found to
be lower in acid solutions; however, the action
potential could still be elicited at pH 5, and
its duration was longer than at normal extracellular pH (3). These results made it unlikely that the reduction of the mechanical
activity of cardiac muscle by acid solutions
resulted from an action of H ions on the
time course of the action potential and proFrom the Division of Physiology and Pharmacology,
National Institute for Medical Research, Mill Hill,
London, England.
This work was supported by a fellowship to the
author by the Wellcome Foundation and by a grant
(NB-02712) from the U. S. Public Health Service.
Accepted for publication August 7, 1966.
CircnUtum Ruttrcb, Vol. XIX, Oaohtt 1966
electrical activity of the heart
intracellular acidity
frog ventricle
vided grounds for inquiring how the process
of mechanical activation was affected by a
change in the extracellular pH. One hypothesis considered in this paper is that negatively charged membrane molecules, whose
existence has been postulated in connection
with the antagonistic action of sodium and
calcium ions on the heart (4), have a high
affinity for hydrogen ions, so that these enter
effectively into the competition even at low
concentrations. The possibility that some effects
of extracellular acidity might result from a
decrease in intracellular pH also could not
be discounted. The cellular pH of large striated
muscle fibers has been shown to be remarkably
independent of the pH of the external solution (5, 6). In heart muscle, however, the
high area-volume ratio and the lower buffer
capacity (7) could render the internal pH
more dependent on extracellular pH. Intracellular structures involved in the regulation
711
712
LORKOVId
of the concentration of calcium could then be
affected by pH changes and so could the receptor sites for calcium. The effects of
selectively decreasing the intracellular pH
with CO2 were therefore tested.
Downloaded from http://circres.ahajournals.org/ by guest on June 17, 2017
Methods
Ventricular strips, 0.5 to 1.0 mm thick, were
dissected from hearts of frogs (male and female
English Rana temporaria or Rana pipiens of
Wisconsin). The frogs, purchased mainly during
summer months, were kept in a cold room for
several weeks.
The strips were mounted in a small, channellike chamber ( 2 x 2 mm cross section) through
which solutions flowed at about 5 ml per min. The
chamber was connected with a six-way tap permitting rapid change of the bath solution (8).
In several experiments the jet-superfusion technique of frog ventricle halves as described by
Gray et al. (9) was applied. It permits an effective and more rapid change of the extracellular
fluid.
Isometric tension was measured with a strain
gauge. The electrical activity was recorded from
the part of the strip which was fixed to the bottom of the chamber. Intracellular glass microelectrodes filled with 3 M KC1 were used. Signals
of both the mechanical and electrical activity
were displayed on a Tektronix 502 oscilloscope
and on an Offner pen recorder. The strips were
driven by supramaximal shocks 1 to 4 msec in
duration from a stimulator at frequencies ranging from 0.5 to 24 impulses per min. Silver or
platinum wires mounted in the bottom of the
chamber served as stimulating electrodes.
The normal Ringer solution contained 110 mM
NaCl, 2.5 mM KC1 and 1 mM CaCl2 and was
buffered by tris-maleate at a concentration between 1 and 8 mM as stated. The pH, measured with a glass electrode, was set by adding
NaOH to the solution. The concentration of NaCl
was set to keep the sodium concentration constant
in solutions of different pH. In some experiments
the concentration of sodium and calcium was
lowered while keeping the quotient of [Na] 2 /
[Ca] constant. In these experiments NaCl was
replaced by sucrose. The composition of the solutions containing CO 2 buffers is given in the text.
Contractures of heart preparations were produced by Ringer solutions to which 25 to 120
mmoles solid KC1 had been added to each liter.
These solutions were allowed to act until the peak
of the contracture was_ reached.
Results
EFFECTS OF CHANGES IN EXTERNAL pH
In Figure 1 is shown the effect of lowering
pH9O pH5-O
25O("
O
2O
10
3O
]25O
O
sec
1
_ J m9
FIGURE 1
Frog ventricle strip (Rana temporaria). Twitch tension
(a) and action potentials (b). Strip in Ringer's solution
by 1 m/n tris maleate. At arrow, the pH was changed
from 9.0 to 5.0. (c) Twitches and action potentials
from the same experiment (indicated by numbers
1, 2, and 3) recorded at a faster time scale.
the pH in the external solution from 9.0 to
5.0 on the twitch tension and the electrical
activity of a frog heart ventricle strip stimulated at 6 impulses/min. In this preparation,
the mechanical activity decreased and almost
disappeared, whereas the membrane potential
and the amplitude of the action potential were
little changed. The duration of the action potential in this and other preparations was
slightly increased early after the application of
the acid solution. A small increase in twitch
tension sometimes occurred in the first minute after the decrease in pH, probably as a
result of the prolongation of action potentials.
On rare occasions, the twitch tension was
decidedly increased during the first 5 to 15
min in the acid solution. This latter effect
was attributable not only to a prolongation
of the action potential but also to a faster rise
of the twitch tension. Evidently, the effect of
low pH on the twitch response was compounded of several factors, some of which tended
to increase the twitch tension.
When the predominant effect was a decrease
in twitch tension, its time course was generally
complex. Thus in Figure 1 the decrease which
began on lowering the pH slowed down after
a few minutes but later the twitch tension
CircuUtkm Ruircb, Vol. XIX, Octobtr 1966
713
EFFECTS OF pH ON CARDIAC MUSCLE
Downloaded from http://circres.ahajournals.org/ by guest on June 17, 2017
again fell more rapidly. By varying the experimental conditions the different phases
could be made more distinct. When the concentration of calcium was low (less than 1
mM), or with a normal calcium concentration
when the frequency of stimulation was low
(less than 2 per min), a simple, approximately
exponential, decline to a lower level was observed (Figs. 2a and 3a). At high concentrations of calcium (3 to 6 mM; Fig. 2b) or
with normal calcium concentration when a
high frequency of stimulation (more than S
per min) was used, the initial fall in twitch
tension was interrupted for some time before
a second decline in tension occurred. The
appearance of both the mechanical and electrical output of the preparations during this
second decline often resembled that occurring
in fatigue: the resting membrane potential
was decreased, the action potentials were
shortened, and their amplitude diminished
until ultimately they failed. The rate of rise
of the twitch tension decreased, however, before any pronounced changes of the action
potentials occurred. The failure of the action
potentials could not, therefore, be considered
as a primary cause of the decrease in twitch
tension.
P H9O
pH&O
Ca ImM
Ilk)
Ca 5mM
0
4
8 mm
FIGURE 1
Frog ventricle strip (Rana temporaria). Twitch tension
and tension developed in response to (a) 100 IUM and
buffered by 5 m.w tris maleate. At arrow, pH was
changed from 9.0 to 5.0. The change to a high [K+]
is indicated by the thick line below the record. Both
records were obtained on the same preparation.
CkcmUliom Resurcb,
Vol. XIX, Ociobtr 1966
pH5-O
pH9O
0-5/min,2mMCo
2O
O
40
so
4 IO
12
4 9
100 120
8/min,2mMCa
24
34
26
28
38
FIGUJtE 3
Frog ventricle strip (Rana temporaria). Twitch tension
and tension developed in response to 100 mM KCl.
The Ca** concentration was 2 mM in a and b and
0.5 ran in c. The frequency of stimulation was 0.5
per min in a; 8 per min in b and 24 per min in c.
The symbols have the same meaning as in Figure 2.
Note change in time scale in record b. The solutions
were buffered by 1 mM tris maleate.
THE Co3*—H+ COMPETITION
Several experiments were designed to test
the hypothesis that the intensity of the mechanical activity depends on a competition
between hydrogen and calcium ions for sites
involved in the mechanical activation of the
muscle.
A possible test at a constant twitch tension
was to keep a given ratio of sodium-tocalcium concentration while varying the absolute concentrations of these ions (4). According to the law of mass action, hydrogen ions
should compete more effectively for the same
anionic sites when the concentration of sodium and calcium ions is low. The solutions
used contained 115 mM Na and 1 mM Ca,
or 20 mM Na and 0.03 mM Ca. Contrary to
expectations, no difference was observed between the decrease in the twitch tension produced by lowering the pH from 9.0 to 5.0
in the two solutions.
714
Downloaded from http://circres.ahajournals.org/ by guest on June 17, 2017
Another approach to the same question was
to test whether the curve relating the twitch
tension to the calcium concentration was
shifted to higher calcium concentration in
low pH. The basic assumptions were that the
quantity of available activator is a function
of the concentrations of hydrogen and calcium, and that the peak twitch tension is
proportional to the maximum activation obtainable in solutions of a given composition.
It is unlikely, however, that the second assumption applies to more than a limited degree. Considerable differences were observed
in the time course of the twitch tension development at various calcium concentrations.
With low calcium (0.2 to 0.5 HIM), tension
rose in a roughly linear fashion throughout
the action potential. In high calcium (more
than 3 IDM), tension rose rapidly to a maximum and then slowly declined before the
action potential was over. This was probably
due to a less pronounced plateau of the action
potential in high calcium. Nearly identical observations have been reported by Niedergerke (10).
Since the tension-[Ca2+] relation is a sigmoid curve, a shift of the curve in the direction of higher [Ca2+] by lowering the pH
would be expected to reflect itself in changes
of twitch tension most clearly at intermediate,
and less at low or high, concentrations of
calcium. The results showed instead that the
decrease in twitch tension due to a lowering
of pH from 9.0 to 5.0 was about the same at
1, 2, and 5 mM Ca. Unfortunately, the experiments were hampered by the partial irreversibility of the low pH effects and even more
by the secondary decline in the twitch tension
when the concentration of calcium was high.
It was therefore often impossible to distinguish
between the effects of the Ca2+-IT competition
and the secondary decrease in twitch tension
mentioned previously.
JET SUPERFUSION
When the perpendicular jet-superfusion
technique was used, the twitch tension was
completely abolished by a solution at pH 5.0
after 5 to 10 sec (Fig. 4). It required 20 to
60 sec to obtain the same effect with pH 5.5.
LORKOVI6
I
B
2min
a
b
c
100
mv
I sec
FIGURE 4
Frog ventricle (halved; Rana pipiens). (A) twitch tension in a preparation temporarily submitted to quick
perpendicular superfusion (indicated by a thick line
below the record). At arrow, pH was changed from
82 to 5.0. (B) action potentials from a similar preparation. A solution with pH 5.0 was applied throughout.
(a) Slow perfusion; between a and b quick superfusion was applied for 5 sec. (b) Superimposed tracings show inhibition of electrical responses after quick
superfusion. (c) Superimposed tracings starting 40
sec after the superfusion was slowed down show
reappearance of action potentials. Frequency of stimulation; 15 per min. The solutions were buffered by
5 min tris maleate.
When the rapid flow of solution (about 2 ml
per sec) was slowed down to less than 0.2
ml per sec, twitches reappeared. The abrupt
decrease in twitch tension in pH 5.5 suggested that a failure of the action potential
might underlie the abolition of twitches. This
was confirmed with intracellular microelectrodes (Fig. 4B, a, b). After the flow of solution was slowed down again, a short, spike-like
potential change appeared; it soon increased
in size and eventually was followed by a plateau (Fig. 4B, c). The spike and the plateau
sometimes occurred separately.
POTASSIUM CONTAACTURES
In the analysis of the antagonism between
calcium and sodium, extensive use has been
made of potassium chloride as a depolarizing
agent (4, 11). In view of the effects of pH
variations on the excitability of heart muscle
and on the duration of the action potential, it
seemed desirable to apply this technique to
the present problem. The rate of tension deCircaUiiom Rtsunb,
Vol. XIX, Octobtr
1966
EFFECTS OF pH ON CARDIAC MUSCLE
715
Downloaded from http://circres.ahajournals.org/ by guest on June 17, 2017
velopment during contracture of heart muscle
strips is limited by diffusion of potassium in
the extracellular space. Faster responses are
obtainable by jet superfusion with solutions
with a high potassium concentration (9). To
compare them with the results obtained with
twitches, the effects of changes in pH on the
potassium contractures elicited both by normal and jet superfusion were investigated.
The tension developed in electrically driven
strip preparations from Rana temporaria in
response to slowly added potassium was decreased in acid solutions but only so long as
the calcium concentration or rate of stimulation was low (Figs. 2a and 3a). The decrease
in contracture tension was best seen when
concentrations of potassium that provoked
submaximum conrractures were applied; it
therefore probably reflects an increase in the
threshold depolarization for tension production (Fig. 5). When a high calcium concentration or a high frequency of stimulation was
used and the twitch tension had entered into
the second decline in acid solution, the contracture tension was increased above the control level (Figs. 2b and 3b). This increase
was not due to a longer but rather to a
100
steeper rise of the contracture tension. The
threshold potassium concentration at which
contractures were obtained was reduced in
these circumstances. Twitch tension and tension developed in response to potassium thus
moved in opposite direction during the second
phase of the action of acid.
The results obtained with ventricle strips
from Rana pipiens were essentially the same
as those described, except for a less pronounced second phase of the decrease in
twitch tension and the concomitant increase
in potassium contractures.
To investigate contracture responses of jetsuperfused preparations, 200 mM KCI was
first applied. The resulting response provided
an estimate of the maximum tension for the
preparation. Submaximum responses provoked
by 70 mmoles KCI added to a liter of Ringer's
solution were used to test the effect of pH
changes. Before the application of the solution with a high potassium concentration the
preparation was flushed by Ringer's solutions at pH 8.2, 6.0, 5.5 or 5.6 for 15 to 30 sec.
The results are presented in Figure 6 and
Table 1. The response to 200 mM KCI was not
affected by changes in pH. The contracture
tension in response to 70 mM KCI decreased
in a roughly linear fashion as pH was lowered,
but the effect was not larger than that obtained with strip preparations. Prolonged exposure to low pH did not affect the results.
EFFECTS OF CHANGES IN INTERNAL pH ONLY
High CO2 tension and HCO~3 buffers were
used to lower the intracellular pH while keep•z
50
25
80
40
KCI
120
[mM)
FIGURE 5
Frog ventricle strip (Rana temporaria). Relative peak
tension at pH 9 (solid line and symbols) and pH 5.8
(dashed line and open symbols) as a function of the
potassium concentration. The high potassium solutions
at pH 5& were applied 5 min after the pH had been
changed.
Ci'C.Ulicm Risnrcb, Vol. XIX, Ociobtr 1966
Imta
FIGURE 6
Frog ventricle (halved; Rana pipiens). Twitches and
potassium contractures during quick superfusion (2
ml per sec). The pH was 8.2 in a and c; it was 5.0 in
b (between the arrows) and d (throughout). The application of high KCI is indicated by the thick line below the record. The concentration of KCI was 70 IUM
in a and b; it was 200 m,w (without NaCl) in c and d.
All solutions contained 2 w Ca.
716
LORKOVIt
TABLE 1
Relative Peak Tension of Contractures Protioked by 70 nut KCl in Five Jet-Superfused Frog
Hearts at pH 5.0 to 8.2*
PH
Relative
tension (%)
8.2
6.0
100
74 ±5.7
(SEM)
5.5
5.0
65 ±6.7
53 ± 8.5
•Tension developed at pH 8.2 was taken as 100%.
Downloaded from http://circres.ahajournals.org/ by guest on June 17, 2017
ing the external pH constant. The pH of a
solution which contained 110 mM NaHCO,
instead of NaCl and was saturated with 80%
CO2 was 6.7. In frog sartorius fibers a decrease
of internal pH by about one unit would be
produced by this solution (12). Since the buffer capacity of the heart muscle is about onehalf of that of a skeletal muscle (13), CO2
should be more effective in depressing the
intracellular pH in the heart. The control
solution, also at pH 6.7, contained 7.9 mM
NaHCOs and was saturated with 5.5% CO2.
The effect of an application of high CO2 at
constant pH is shown in Figure 7B. Immediately upon admission of 80% CO2, there
was a rapid decline in twitch tension. About
1 min later the twitch tension reached a
a
CO2 5-5%
HCOj7-9mM
A
b
CO, BO°/o
HCO7 M5mM
°
6 m m
<
^
\
bllrrar*
b(5mln)
120
H
5
FIGURE 7
Frog ventricle strip (Rana temporaria). (A) action
potentials (top) and twitch tension (bottom) recorded
from a preparation which was first in solution a containing 7.9 mm NaHCOs and equilibrated with 5.5%
COS (first record). The second and third records were
taken 1 and 5 min after the application of solution b
containing 115 mM NaHCO} and equilibrated with
80% CO,. Note the altered calibration for the mechanical tension in the third record. The voltage step following the action potential in the first record is equal
to 100 mv. (B) twitch tension and tension developed
in response to 100 mM KCl (indicated as in Figure 3)
in solutions a and b. Tetanization (8 shocks per sec) is
indicated by the squares above the record. The solutions contained 1 mM Ca.
minimum, which was followed by a transient
increase. In most preparations the twitch tension settled at a steady level less than 10% of
the original tension; in some it dropped to
only about 20% and tended to increase later.
The amplitude of the action potential was
unchanged or even increased. In Figure 7A
action potentials and twitches recorded at a
fast time scale are shown. The action potentials grew longer during the initial fall of tension, sometimes by more than 30%, later they
became shorter, decreasing to about 60% of
the initial length. A gradual prolongation of the
relaxation phase of the twitches and a slight
contracture were always observed to accompany the small secondary increase in twitch
tension, in rhythmically stimulated preparations. After 5 to 10 min the contracture and the
rate of relaxation reached a steady state. When
stimulation was stopped simultaneously with
the application of high CO2, the relaxation
phase of the first twitch after a rest of 5 min
was not prolonged, but the prolongation was
fully established after 3 or 4 twitches. A rest of
5 min had no influence on the rate of relaxation once the slowing had been established.
The extent of the slowing in the relaxation
phase was not much reduced by omitting Ca2*
from the high CO2 solution at any time.
To distinguish between the effects of CO2
and HCO~a experiments were done in which
a solution containing 115 mM HCO~8 without
CO2 (pH about 9) was applied 5 min ahead
of the same solution saturated with 80% COa.
The application of high bicarbonate usually
decreased the twitch tension to about 80%.
Upon application of high CO2, the same
changes in twitch tension occurred as described previously.
Potassium contractures were greatly increased by the high CO2 treatment at constant
CircuUtion Ruurcb, Vol. XIX, Ocioitr 1966
EFFECTS OF pH ON CARDIAC MUSCLE
Downloaded from http://circres.ahajournals.org/ by guest on June 17, 2017
external pH, and the threshold for contractiires was reduced. The maximum sensitivity to potassium was reached 5 to 10 min
after the application of high CO2 (Fig. 7B),
but some effect was already noticeable after
30 sec. The effect of potassium was not further increased by raising the stimulation frequency before the application of potassium.
There is now considerable evidence that
voltage and time-dependent processes determine the tension response of skeletal muscle
to depolarization (8), and it is probable that
in heart muscle a similar, but slower system
exists. The question therefore arises whether
the striking differences between the behavior
of twitches and contractures when intracellular pH is reduced by CO2 can be ascribed
to the difference in the time course of the
depolarization in the two cases. If it is assumed that a process of activation and a process of inactivation are both slowed by high
CO2, the transient depolarization by an action
potential might be unable to activate the contractile mechanism, whereas a long-lasting depolarization by potassium may be more effective than normally. The tension response
to a series of closely spaced action potentials
might then build up to nearer the response
produced by potassium. In some preparations,
the frequency of stimulation was therefore increased to 8 to 32 per sec (only a fraction of
the stimuli being effective) for short periods
at different times after the admission of CO^After 1 min, before the slight secondary increase in tension had developed, the tension
response was very weak. However, a strong
superposition of the mechanical response was
found 5 min after CO2 had been applied.
The tetanic tension obtained in this way was
5 to 30 times larger than the twitch tension
(Fig. 7B) and comparable to the contracture
tension provoked by 100 ITIM potassium.
The twitch tension increased above the
control values on return to the low COi> solution and remained elevated for more than half
an hour. The rate of relaxation decreased and
approached normal values during this time.
The changes in the mechanical activity induced by high COj were essentially the same
CircuUtio* Rtiurch, Vol. XIX, Oaobtr 1966
717
when ventricle halves from Rana pipiens were
jet superfused. A just superthreshold contracture was produced by about 40 ITIM KCI
(Fig. 8); 5 min after the high CO2 solution
had been applied, contractures of more than
50% of maximum tension were produced by
the same concentration of potassium. The rate
of rise of tension was slow in spite of the
jet superfusion, which is in keeping with a
slowed activation as suggested earlier. One
minute after the original solution with low CO2
had been reapplied, maximum tension was developed by the preparation in response to the
same potassium concentration, and the tension
development was quick. The rate of activation thus appears to become normal early after
COj is withdrawn, whereas rate of inactivation remains slow for a much longer time.
EFFECTS OF A SIMULTANEOUS DECREASE IN
EXTERNAL AND INTERNAL pH
A Ringer's solution containing 5 mM bicarbonate was used in these experiments. In
equilibrium with the room atmosphere the
pH of this solution was 8.4 to 8.8. When 80$
CO2 was bubbled through it, the pH dropped
to 5.3. The effect of varying the CO2 content of such a solution is shown in Figure 9.
A rapid fall in twitch tension occurred immediately on admission of CO2. The membrane
potential decreased progressively in some
preparations; in others there was only an initial slight depolarization and the membrane
potential was well maintained afterwards. The
action potential was prolonged at the beginning of the high COu application but returned
to control values later. The twitch tension,
II min
Frog ventricle (halved; Rana pipiens). The twitches
and contractures provoked by 40 min KCl (indicated
as before ) in solution a and b (see legend for Figure
7). Jet superfusion was started about 20 sec before the
application of 40 n\M KCl and stopped toward the
end of the contracture.
718
LORKOVie
pH8-5
pH8-5
Downloaded from http://circres.ahajournals.org/ by guest on June 17, 2017
IOO
mV
sec I
FIGURE 9
Frog ventricle strip (Rana temporaria). Twitch tension
(a) and action potentials (b) in a Ringer solution containing 5 mM NaHCOy At arrow, a similar solution
equilibrated with 80% COt was applied, (c) Three
stages of the experiment recorded at a faster time
scale.
depressed by CO2) recovered only slightly
when the concentration of calcium was increased from 1 to 5 to 10 mM. A slowing of
relaxation was sometimes observed when the
calcium concentration was high, but the potassium contracture was always less than in
an alkaline solution without CO2, which contained calcium in the same concentration;
tetanization was ineffective. The twitch tension recovered almost to control value upon
reapplication of the alkaline solution.
Discussion
The complexity of the present results becomes more comprehensible if the probability
is borne in mind that the relation between
electrical and mechanical activity in muscular
tissues is reciprocal in nature: the time course
of the action potential may influence the production of tension, and reactions connected
with contraction and its activation may influence the action potentials, as has recently
been demonstrated for skeletal muscle (14).
All the systems involved can probably be influenced by a change in pH. Thus the in-
crease in twitch tension sometimes seen soon
after the extracellular pH was lowered was
probably due, at least in part, to a prolongation in the action potential such as has been
seen in Purkinje fibers (3). The decrease in
action potential height and duration which
developed after some time in acid solution,
when the calcium concentration was high,
could possibly be a consequence of changes
in the processes connected with contraction.
Such changes may well not occur in noncontractile Purkinje fibers. In jet-perfused preparations yet another type of electrical failure
occurred: the action potentials disappeared
abruptly and completely.
A roughly exponential decrease of mechanical responses was observed after lowering the
extracellular pH when the frequency of stimulation and the calcium concentration were
low. Under these conditions the twitches and
potassium-induced contractures fell to about
the same extent and the action potentials
were well maintained. The dominant effect
was then probably a decrease in the availability of calcium ions. But the situation did
not lend itself to quantitative treatment in
terms of the model applicable to Ca*+-Na+
antagonism (11), though some evidence for a
weak H+-Ca2+ antagonism was obtained in
potassium contractures. In the most sensitive
region between pH 5 and pH 7 a 100-fold
increase in [H+] was always less effective
than a twofold increase in [Na+]. Preliminary
results suggest that a different relationship
exists in skeletal muscle, where the threshold
depolarization for mechanical activation is
more sensitive to changes in [H+] than in
[Na+]. It is improbable that the failure of the
low sodium-low calcium solutions to potentiate
the effects of low pH was due to the lowering
of the mechanical threshold that has been
observed in these solutions (11). A lack of
positive staircase when the frequency of stimulation in the low sodium solutions is increased (15) was also observed. Although both
of the effects mentioned make it difficult to
compare the effects of pH at different sodium
levels, they do not prevent the conclusion
that the effect of low pH is not exerted via
Circulation Rts—rcb, Vol. XIX, Oaobtr
1966
719
EFFECTS OF pH ON CARDIAC MUSCLE
Downloaded from http://circres.ahajournals.org/ by guest on June 17, 2017
the mechanism underlying the Ca2+-Na+ competition.
The increase of the potassium contractures
after lowering the extracellular pH when the
external calcium concentration of it is elevated or when, as with frequent stimulation,
the calcium turnover is increased (13), may
be due to an increase of the calcium concentration in the sarcoplasm. This is suggested
by the small contracture which generally developed in solutions with a high calcium
concentration. It seems possible therefore that
low pH slows the removal of calcium from
the interior of the cells.
It has been found that calcium release from
skeletal muscles increases when they are
treated with high CO2 (16). If a similar effect
occurs in cardiac muscle and if the calcium
involved is released from some store in the
fibers, a rise in the intracellular concentration of ionic calcium to near that which is
needed for mechanical activation could occur.
An intracellular release of Ca2+ by high CO?
cannot, however, account for the fact that the
relaxation in a twitch is not slowed initially
when stimulation is first applied 5 or more
min after application of CO2. The gradual
appearance of the slow relaxation seems to
indicate that the removal of calcium from
some specific localities is inhibited. The common feature of the two conditions leading
to increased contractures thus appears to be an
increase in the intracellular ionized calcium.
The decrease in twitch tension produced
by simultaneous reduction of the extracellular
and intracellular pH was more pronounced
than when the intracellular pH only was lowered. The amplitude of the contracture response was never increased, even in high Ca
solution. Since contractures were increased in
high calcium both when the extracellular and
when the intracellular pH were lowered separately, it is difficult to explain this result.
One conclusion which can be drawn on
the basis of the present experiments is that
a slowing of inactivation of the contractile
element, as reflected in a slow relaxation, is
not necessarily linked to a slowing in activaOrcuUltcm Rnmrch, Vol. XIX, Ociobtr 1966
tion as reflected in small twitches. This was
shown by the fact that relaxation was slow
long after the twitch tension had fully recovered after intracellular acidification with
CO2.
Acknowledgments
I would like to thank Sir Peter Medawar for hospitality and facilities at the National Institute for
Medical Research and Dr. O. F. Hutter for suggesting the subject of investigation and for his continued
interest in this work. I also gratefully acknowledge
my debt to Dr. C. Edwards who allowed me to finish
some of the experiments in his laboratory at the Department of Physiology, University of Minnesota. A
preliminary report of part of the work has appeared in
J. Physiol. (London) 177, 36-37P.
References
1. DALY, J. DE BURGH, AND CLARK, A. J.: Action of
ions upon the frog's heart. J. Physiol. (London) 54: 367-383, 1921.
2.
MINES, G. R.: On functional analysis by the ac-
tion of electrolytes. J. Physiol. (London) 46i
188-235, 1913.
3.
HECHT, H. H., AND HUTTER, O. F.: Action of
pH on Cardiac Purkinje Fibres in Electrophysiology of the Heart, ed. by B. Taccardi.
London, Pergamon Press, pp. 105-123, 1964.
4.
LUTTGAU,
H. C ,
AND NiEDEHGEHKE, R.:
An-
tagonism between Ca and Na ions on the
frog's heart. J. Physiol. (London) 143: 486505, 1958.
5.
CALDWELL, P. C : Studies on the internal pH of
large muscle and nerve fibres. J. Physiol. 142i
22, 1958.
6. KOSTYUK, P. C , AND SOROKINA, S. A.: On the
mechanism of hydrogen ion distribution between cell protoplasm and the medium. In
Symposium on Membrane Transport and Metabolism, ed. by F. Kleinzeller. Prague, Publishing House of Czechoslovak Academy of
Sciences, pp. 193-203, 1961.
7. BRODY, H.: Carbon dioxide dissocation curve of
frog heart muscle. Am. J. Physiol. 93: 190196, 1930.
8. HODCKIN, A. L., AND HOROWICZ, P.: Potassium
contractures in simple muscle fibres. J. Physiol.
(London) 153: 386-403, 1960.
9. GRAY, H. W., LAMB, J. F., AND MCGUIGAN,
J. A. S.: Potassium exchanges in quiescent and
beating frog's ventricle. J. Physiol. (London)
172: 25P, 1964.
10.
NIEDERCERKE, R.: Staircase phenomenon and the
action of calcium on the heart. J. Physiol.
(London) 134: 569-583, 1956.
11. NIEDERCERKE, R.: Potassium chloride contracture
720
of the heart and its modification by calcium.
J. Physiol. (London) 134: 584-599, 1956.
12. HILL, A. V.: Influence of the external medium
on the internal pH of muscle. Proc. Roy. Soc.
London, Ser. B 144: 1-21, 1955.
LORKOVId
muscle fibres. J. Physiol. (London) 178: 4567, 1965.
15. REITER, M.: Einfluss der Erregungsfrequenz auf
die Inotrope Wirkung von Calcium am Meerschweinchen-Papillarmuskel. Arch. Exper. Pathol. Pharmakol. 247: 330-338, 1964.
13. BRADY, A. J.: Excitation and excitation-contrac-
tion coupling in cardiac muscle. Ann. Rev.
Physiol. 26: 341-356, 1964.
14. LUTTCAU, H. C : Effect of metabolic inhibitors
on the fatigue of the action potential in single
16.
SWANT, R. C : Influence of pH on calcium ex-
change in skeletal muscle. Communications,
21 International Congress of Physiology. Science, Buenos Aires, p. 267, 1959.
Downloaded from http://circres.ahajournals.org/ by guest on June 17, 2017
Circulation Rtiurcb, Vol. XIX, Oclobir 1966
Influence of Changes in pH on the Mechanical Activity of Cardiac Muscle
HRVOJE LORKOVIC
Downloaded from http://circres.ahajournals.org/ by guest on June 17, 2017
Circ Res. 1966;19:711-720
doi: 10.1161/01.RES.19.4.711
Circulation Research is published by the American Heart Association, 7272 Greenville Avenue, Dallas, TX 75231
Copyright © 1966 American Heart Association, Inc. All rights reserved.
Print ISSN: 0009-7330. Online ISSN: 1524-4571
The online version of this article, along with updated information and services, is located on the
World Wide Web at:
http://circres.ahajournals.org/content/19/4/711
Permissions: Requests for permissions to reproduce figures, tables, or portions of articles originally published in
Circulation Research can be obtained via RightsLink, a service of the Copyright Clearance Center, not the
Editorial Office. Once the online version of the published article for which permission is being requested is
located, click Request Permissions in the middle column of the Web page under Services. Further information
about this process is available in the Permissions and Rights Question and Answer document.
Reprints: Information about reprints can be found online at:
http://www.lww.com/reprints
Subscriptions: Information about subscribing to Circulation Research is online at:
http://circres.ahajournals.org//subscriptions/