Creatine and Creatinine Metabolism - Physiological Reviews

July 2000 CREATINE AND CREATININE METABOLISM 1127
D. CK
Physiological and biochemical aspects of the CK
isoenzyme system have been briefly introduced at the
beginning of this article and were the subject of recent
reviews (837, 838, 1084, 1124). Instead of repeating the
relevant arguments, some major achievements made in
the last few years are summarized here.
Transgenic animals lacking single isoenzymes of CK
or combinations thereof have recently been analyzed
carefully (for reviews, see Refs. 502, 838; see also Refs. 73,
527, 818, 848, 942, 1026, 1047, 1091). Mutant mice lacking
the muscle form of cytosolic CK, M-CK, are viable and
fertile and display neither overt abnormalities nor alterations
in absolute muscle force (1043). However, the gastrocnemius-plantaris-soleus
muscle complex of these M-
CK knockout mice lacks the ability to perform burst
activity, i.e., at 1- and 5-Hz stimulation, the isometric
twitch force is considerably lower for the first 70 and 25 s,
respectively, than in controls. After these periods, the
ratios invert and mutants develop even greater force than
controls.
Graded mutants displaying a two-, three-, or sixfold
depression of MM-CK activity relative to controls showed
intermediary properties in as far as force generation is
concerned (1045). Interestingly, no flux through the CK
reaction could be detected by the 31 P-NMR inversion
transfer technique in muscles having 0, 16, and 34% of
wild-type MM-CK activity, whereas significant fluxes of
similar size were measured in muscles having 50 and 100%
of wild-type activity. These findings clearly indicate that a
considerable portion of the CK flux in muscle must be
NMR invisible, and therefore question the usefulness of
NMR experiments for measuring total CK flux.
Single mutants lacking the sarcomeric mitochondrial
CK (Mi b-CK) gene showed no abnormalities in skeletal
muscle morphology and short-term performance (939).
Oxidative phosphorylation capacity also was normal in
both skeletal muscle and heart, except for an impaired
stimulation of mitochondrial respiration by Cr (73, 939).
These findings might raise questions about the physiological
functions of Mi-CK. It has to be kept in mind, however,
that the proportion of Mi-CK to total CK activity in
skeletal muscle is generally rather low. Because this proportion
is considerably higher in rodent heart, it will be
more interesting to study the effects of the Mi b-CK knockout
mutation on heart function.
Double mutants deficient in both M-CK and Mi b-CK,
despite being apparently normal, displayed even more
pronounced disturbances in gastrocnemius and diaphragm
muscle performance than M-CK-deficient single
mutants (527, 938, 1091). Tetanic force, power, work, and
the rates of tension development and relaxation were
considerably decreased. The enhanced endurance performance
seen in M-CK-deficient gastrocnemius muscle was
not observed in double mutants. Cultured myotubes of
CK-deficient double mutants showed abnormal Ca 2� handling
in response to depolarization by ACh or KCl, as
evidenced by a 30% decrease in the [Ca 2� ] amplitude, a
60% increase in the plateau level of Ca 2� after stimulation,
and decreased rates of Ca 2� release and sequestration.
Furthermore, double mutants displayed ultrastructural
abnormalities, most notably conspicuous tubular aggregates
of SR membranes in fast skeletal muscle fibers.
Such tubular aggregates are also seen in various myopathies
with electrolyte disturbances and may represent a
metabolic adaptation to abnormal Ca 2� handling.
Isolated perfused hearts from wild-type, M-CK-deficient,
and both M- and sarcomeric Mi-CK-deficient mice
displayed comparable isovolumic left ventricular contractile
performance at baseline and responded to increased
heart rate and perfusate [Ca 2� ] with similar increases in
rate-pressure product (848). During baseline perfusion,
[ADP] was significantly higher in M- plus Mi-CK-deficient
hearts (214 �M) than in either wild-type or M-CK-deficient
hearts (150 �M). In contrast, the �G for ATP hydrolysis
was not significantly different between the three groups.
Increasing heart rate and perfusate [Ca 2� ] increased
[ADP] in M- plus Mi-CK-deficient hearts to 407 �M and
decreased |�G ATP| by 3.6 kJ/mol. On the other hand, in
either wild-type or M-CK-deficient hearts, it had no significant
effect on [ADP] and decreased |�G ATP| by only 1.7
kJ/mol. Consequently, increases in cardiac work in Mplus
sarcomeric Mi-CK-deficient mice become more “energetically
costly” in terms of high-energy phosphate use,
accumulation of ADP, and decreases in the free energy of
ATP hydrolysis (848).
Because the importance of the CK/PCr/Cr system for
high-energy phosphate transport had been demonstrated
most elegantly in sea urchin spermatozoa (1011), it was
straightforward to test the effects of a disruption of the
proposed PCr shuttle in mouse spermatozoa which contain
ubiquitous mitochondrial CK (Mi a-CK) and the brain
isoenzyme of cytosolic CK (B-CK) (942). Surprisingly,
transgenic mice deficient in ubiquitous Mi-CK displayed
normal viability and fertility as well as normal motility
patterns of isolated spermatozoa. In summary, the mild
phenotype of some of the CK knockout mutants was
rather unexpected. Upon closer examination, however,
the CK-knockout mice stress the importance of the CK
system for high-energy phosphate metabolism and transport,
proper Ca 2� handling and, thus, muscle performance.
In contrast to these CK-knockout mice, gain of function
effects were studied in transgenic mice expressing
B-CK or ubiquitous Mi-CK in liver, a tissue that is normally
devoid of CK (32, 96, 347, 454, 502, 642, 643, 717, 846).
These livers, in the presence of Cr and PCr, revealed a
protection of [ATP] and decreased adenine nucleotide
degradation during a fructose load (96), delayed ATP