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1 Dalton Cardiovascular Research Center, 2 Department of Veterinary Biomedical Sciences, and 3 Department of Physiology, and University of Missouri, Columbia, Missouri 65211
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
ACKNOWLEDGEMENTS
FOOTNOTES
REFERENCES
ABSTRACT 
Parker, Janet L., Mildred L. Mattox, and M. Harold Laughlin.
Contractile responsiveness of coronary arteries from exercise trained rats. J. Appl. Physiol. 83(2):
434-443, 1997.
The purpose of this study was to determine whether
exercise training alters vasomotor reactivity of rat coronary arteries.
In vitro isometric microvessel techniques were used to evaluate
vasomotor properties of proximal left anterior artery rings (1 ring per
animal) from exercise-trained rats (ET;
n = 10) subjected to a 12-wk treadmill training protocol (32 m/min, 15% incline, 1 h/day, 5 days/wk) and
control rats (C; n = 6) restricted to
cage activity. No differences in passive length-tension characteristics
or internal diameter (158 ± 9 and 166 ± 9 µm) were observed
between vessesls of C and ET rats. Concentration-response curves to
K+ (5-100 mM), prostaglandin
F2
(10
8-104
M), and norepinephrine
(108-104)
were unaltered (P > 0.05) in
coronary rings from ET rats compared with C rats; however, lower values
of the concentration producing 50% of the maximal contractile response
in rings from ET rats (P = 0.05)
suggest that contractile sensitivity to norepinephrine was
enhanced. Vasorelaxation responses to sodium nitroprusside (109-104
M) and adenosine
(109-104
M) were not different (P > 0.05)
between vessels of C and ET rats. However, relaxation responses to the
endothelium-dependent vasodilator acetylcholine (ACh;
1010-104
M) were significantly blunted (P < 0.001) in coronary rings from ET animals; maximal ACh relaxation
averaged 90 ± 5 and 46 ± 12%, respectively, in vessels of C
and ET groups. In additional experiments, two coronary rings (proximal
and distal) were isolated from each C
(n = 7) and ET
(n = 7) animal. Proximal coronary
artery rings from ET animals demonstrated decreased relaxation
responses to ACh; however, ACh-mediated relaxation of distal coronary
rings was not different between C and ET groups.
NG-monomethyl-L-arginine
(inhibitor of nitric oxide synthase) blocked ACh relaxation of all
rings. L-Arginine (substrate for
nitric oxide synthase) did not improve the blunted ACh relaxation in proximal coronary artery rings from ET rats. These studies suggest that
exercise-training selectively decreases endothelium-dependent (ACh) but
not endothelium-independent (sodium nitroprusside) relaxation responses
of rat proximal coronary arteries; endothelium-dependent relaxation of
distal coronary arteries is unaltered by training.
vascular smooth muscle; potassium; norepinephrine; prostaglandin
F2 INTRODUCTION EXERCISE TRAINING has been reported to induce increases
in coronary vasodilator capacity and capillary diffusion capacity (10,
13, 14). Laughlin and colleagues (10, 13, 14) and Parker
et al. (25) reported that the coronary circulation of exercise-trained
animals demonstrated enhanced adenosine (Ado)-induced vasodilation and
evidence of decreased The purpose of this study was to determine whether exercise training
induces alterations in vasoconstrictor responses and/or endothelium-dependent and -independent vasodilator responses of rat
coronary arteries. Our hypothesis was that exercise training alters the
contractile responses of rat coronary arteries to NE, SNP, and Ado in a
manner similar to that previously reported by our laboratory for
miniature swine (22). Vasoconstrictor responses were
studied by examining the concentration-contractile response relationship to exogenous K+ for
contractions mediated by opening voltage-gated
Ca2+ channels and to NE and
prostaglandin F2 METHODS 
; adenosine; nitroprusside; acetylcholine; L-arginine; NG-monomethyl-L-arginine; nitric oxide synthase; flow
-adrenergic responsiveness assessed in
vivo. Also, DiCarlo et al. (5) reported changes in
coronary vascular responses to vasoactive agents in conscious, exercise-trained dogs, and Bove and Dewey (3) reported that vasoconstrictor responses to phenylephrine were attenuated in left
anterior descending and circumflex coronary arteries of intact, exercise trained dogs. More recently, Oltman et al. (22) reported that
vasoconstrictor responses to norepinephrine (NE) and vasodilator responses to sodium nitroprusside (SNP) were blunted, whereas the
vasodilator response to Ado was enhanced in epicardial coronary arteries isolated from exercise trained pigs. These training-induced alterations appear to be relatively selective for vascular smooth muscle of epicardial coronary arteries, because peripheral arteries (femoral, renal, mesenteric) isolated from the same porcine training model did not demonstrate these changes (with the exception of NE
responses in renal arteries) (16). It is not known whether similar
adaptations are induced in coronary arterial smooth muscle of
exercise-trained rats.
(PGF2) for receptor-mediated
contractions. Vasodilator responses were studied by
examining the concentration-relaxation response relationships to NP and
Ado for direct vascular smooth muscle relaxation. Endothelium-dependent vasodilation was evaluated by using the receptor agonist acetylcholine (ACh).
D/2) or
L = 2f + D(2 + ), where
D is wire diameter and
f is the distance between the wires as
measured by the micrometer. Coronary rings were set at
L80, where
L80 = 0.8 L100 and
L100 is the
internal circumference the vessel would have under a transmural pressure of 100 mmHg (12, 25). On a subgroup of ET
(n = 5) and C
(n = 5) rats, complete length-active
tension curves were performed by using progressive vessel stretches and
repetitive contractile responses to
K+ (40 mM) at each level of
stretch. We determined that values of L80 were at or
near (within 5-10%) the optimal degree of stretch (L) where maximal active tension to
K+ is developed
(Lmax). After
equilibration, vessels were contracted three times with 100 mM
K+, rinsed with standard
bicarbonate buffer, and reequilibrated before the experiment was begun.
Contractile responsiveness.
Concentration-response curves to contractile agonists (NE,
PGF2) were obtained by adding
cumulatively increasing concentrations of the appropriate drug to the
reservoir superfusate of the microvessel myograph.
Concentration-contraction response curves to KCl were performed by
using perfusate solutions containing increasing concentrations of KCl
(substituted for NaCl to maintain constant osmolarity). Contractile
responses are presented as absolute contractile tension (mN/mm) and as
EC50 values calculated (when
appropriate) as the concentration producing 50% of the maximal
contractile response.
Relaxation responses.
To evaluate relaxation responses, coronary vessels were first
precontracted with equieffective concentrations of
PGF2 (approximately
half-maximal K+ contraction), and
the contraction was allowed to stabilize before the
concentration-relaxation response was performed to the vasodilator (Ado, ACh, SNP). Equivalent
PGF2 precontractions in vessels from C and ET groups were used for relaxation studies. Responses are
presented as a percentage of the precontraction (percent relaxation). Values for IC50 were calculated as
the drug concentration that produced 50% of the maximal relaxation
response.
Oxidative enzyme activity.
At the time the animals were killed, 50-mg samples were taken from the
middle of the medial and long heads of triceps brachii muscles. Immediately after dissection, the muscle samples
were frozen in liquid nitrogen and stored at 
70°C until
assay. The muscle samples were homogenized on ice in 0.4 M KCl and 20%
ethanol and centrifuged (23,000 g),
and the supernatant was dialyzed against 2 mM phosphate buffer (pH
7.4). Citrate synthase (CS) activity was determined
spectrophotometrically at 412 nm at 25°C as described by Srere
(33). Protein content was determined with the Bradford dye-binding
procedure with bovine albumin as the standard. These measurements were
used to validate the degree of adaptation of the skeletal muscle in
response to the exercise-training protocol, compared with respective
muscle assays of the sedentary C animals.
Drugs and solutions.
The Krebs bicarbonate buffer contained (in mM) 131.5 NaCl, 5.0 KCl, 1.2 NaH2PO4,
1.2 MgCl2, 2.5 CaCl2, 25.0 NaHCO3, and 11.2 glucose. This
solution was aerated with 95%
O2-5%
CO2 (pH 7.4) and was maintained at
37 ± 0.5°C. The KCl concentration was increased in designated
solutions. Unless otherwise specified, drugs were obtained from Sigma
Chemical (St. Louis, MO). All solutions contained 3 µM propranolol
(to block 
-adrenergic effects of NE) and 0.025 mM EDTA (to minimize
oxidation of NE). The drugs used were
PGF2 (Upjohn), Ado (Eastman
Kodak), NG-monomethyl-L-arginine
(L-NMMA; Calbiochem), NE, and
ACh (Sigma Chemical); concentrated stock solutions were prepared with
deionized water.
Data analysis and statistics.
Drug concentration-response curves were compared by using two-way
analysis of variance for repeated measures.
EC50 and
IC50 values were calculated by
using nonlinear regression analysis of the concentration-response data.
Student's unpaired t-test was used to
compare intergroup differences in vessel dimensions, EC50 and
IC50 values, maximal responses,
and levels of preconstriction before exposure to vasodilator agents.
Statistically significant differences were defined as
P < 0.05. Data are presented as
means ± SE.
RESULTS
1 · g
wet wt1;
P < 0.05), a 45% greater CS
activity in the red portion of the long head of triceps brachii (51 ± 4 vs. 35 ± 3 µmol · min1 · g
wet wt1;
P < 0.05), and a 25% greater CS
activity in the soleus muscle of ET compared with C rats (66 ± 3 vs. 53 ± 4 µmol · min1 · g
wet wt1;
P < 0.05). Hindlimb skeletal muscle
of the ET rats had greater blood flow and capillary exchange capacity
than did that of the sedentary C rats (31, 32). These data indicate
that the ET rats included in the present study exhibited classic signs
of the exercise-trained state (2, 28).
Vessel characteristics.
Dimensional characteristics of proximal coronary artery segments
isolated from C untrained rats and ET rats (series
1) are summarized in Table
1. No significant differences
were noted in vessel luminal diameter, axial length, or passive
length-tension characteristics of the vessels (Table 1). However, wall
thickness of coronary segments of arteries of ET rats was significantly larger (P < 0.001) than of coronary
artery segments of C rats. Values for circumferential length at
L80 and
L100 were not
significantly different between the two groups; similarly, resting
tension at L80
was not different between coronary rings of ET and sedentary C rats.
Similarly, length-active tension relationships determined in a subgroup
of C (n = 5) and ET rats
(n = 5) indicated no differences in
circumferential length at
Lmax (optimum of
length-active tension relationship) or in resting tension at
Lmax (not shown).
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(Fig. 1).
Concentration-response curves to
PGF2
(108-104
M) were not different in vessels from C and ET rats. Contractions obtained in response to the maximal
PGF2 concentration used (104 M) averaged 2.7 ± 0.3 and 2.9 ± 0.2 mN/mm (P > 0.05), respectively, in C and ET groups; these values were comparable
to maximal K+-induced contractions
(EC50 for
PGF2 was not calculated because we were unsure whether 104
M values represented maximal responses). As also shown in Fig. 1,
coronary arteries exhibited concentration-dependent contractile responses to NE over the concentration range
108-104
M. Maximal contractions obtained to NE were, however, substantially smaller than those obtained to either
K+ or
PGF2 (Fig. 1). Maximal NE
contractions averaged 0.9 ± 0.3 and 0.9 ± 0.2 mN/mm
(P > 0.05), and no statistically
significant difference was obtained between the complete
concentration-response curves of the two groups. However, there was an
increased sensitivity to NE over the lower concentration range
(108-106
M) in vessels from ET rats; calculated
EC50 values for NE responses averaged 5.6 ± 0.9 and 3.3 ± 0.6 µM
(P = 0.05) in coronary arteries from C
and ET rats, respectively.
(108-104
M; B), and norepinephrine
(108-104
M, evaluated in presence of 3 µM propranolol;
C) in proximal coronary artery
segments isolated from control and exercise-trained rats. Values are
means ± SE from no. of animals in parentheses. NS, not
significantly different. Inset, values
of concentration producing 50% of maximal contractile response
(EC50) for control (C) and
exercise-trained (T) rats.
Relaxation responses. Relaxation responses to endothelium-dependent (ACh) and endothelium-independent (SNP, Ado) vasodilators were obtained in coronary arteries precontracted with PGF2
(concentration yielding approximate half-maximal K+
contractions). Importantly,
PGF2 precontractions were not different (P > 0.05) in
control and ET groups; these values are indicated for each group in the
legends of Figs. 2, 3, 4, 5. As illustrated in Fig.
2, coronary arteries from both groups
exhibited concentration-dependent relaxation responses to ACh
(1010-104
M). However, concentration-relaxation responses to ACh in coronary arteries from ET rats were significantly decreased
(P < 0.001) compared with coronary
arteries from C rats. Maximal relaxation to ACh averaged 46.1 ± 12.1 and 89.8 ± 5.4% (P < 0.01), respectively, in arteries from ET and C rats, indicating a
non-competitive-like inhibition of maximal relaxation.
IC50 values for ACh relaxation averaged 456.6 ± 197.5 and 64.4 ± 40.6 nM
(P < 0.05), respectively.
10-104
M) in proximal coronary artery rings isolated from control and exercise-trained rats. Values are means ± SE from no. of animals in
parentheses. Vessels were precontracted with equipotent concentrations of prostaglandin F2
(PGF2) and allowed to
restabilize before exposure to acetylcholine. Levels of preconstriction
averaged 1.73 ± 0.23 and 2.01 ± 0.16 nN/mm in sedentary control
and trained groups, respectively (NS).
Inset, drug concentration that
produced 50% of maximal relaxation response
(IC50) values for control and exercise-trained rats.
9-104
M; A) and adenosine
(109-104
M; B) in proximal coronary artery
rings isolated from control and exercise-trained rats. Values are means ± SE from no. of animals in parentheses. Rings were precontracted
with equipotent concentrations of
PGF2 and allowed to restabilize
before exposure to nitroprusside. Levels of preconstriction were not
different (P > 0.05) between sedentary control and trained groups (nitroprusside: 1.76 ± 0.28 vs. 1.82 ± 0.21 nN/mm; adenosine: 1.98 ± 0.12 vs. 2.09 ± 0.18 nN/mm in sedentary control and trained groups respectively).
Inset, IC50 values for control and
exercise-trained rats.
and allowed to restabilize
before exposure to acetylcholine. Levels of preconstriction were not
different (P > 0.05) between
sedentary control and trained groups (proximal: 2.81 ± 0.45 vs.
3.23 ± 0.56 nN/mm; distal: 2.84 ± 0.43 vs. 2.97 ± 0.55 nN/mm in sedentary control and trained groups, respectively).
) and exercise-trained
(B; 
) rats. Values are means ± SE from no. of animals in parentheses.
Relaxation responses to the endothelium-independent vasodilators SNP and Ado are illustrated in Fig. 3. In vessels precontracted with PGF2
(approximate half-maximal contraction;
PGF2 concentrations indicated
in Fig. 1 legend), SNP (109
to 104 M) produced
concentration-dependent relaxation responses. However, in
contrast to ACh-mediated relaxation, SNP relaxation responses were not
different (P > 0.05) in coronary
arteries from C and ET rats, and maximal (100%) relaxation of the
PGF2 contraction was achieved
in both groups. Similarly, SNP
IC50 values were not significantly
different and averaged 16.83 ± 9.63 and 6.81 ± 1.14 nM,
respectively, in arteries from C and ET rats. Ado relaxed PGF2 contractions of both
groups of vessels; however, the potency of Ado as a vasodilator in
coronaries from either C or ET rats was far less than that of either
SNP or ACh (Fig. 3). Essentially, no Ado-mediated relaxation was
observed in either group over the concentration range
109-106
M. At 104 M, Ado relaxation
averaged 56.20 ± 10.70 and 54.90 ± 6.11%
(P > 0.05) in coronary arteries from
C and ET rats, respectively. IC50
values for Ado relaxation were not calculated, because maximal relaxation had not been obtained even at
104 M Ado.
Series 2: Proximal and distal coronary arteries.
As described in METHODS, two coronary
rings (proximal and distal segments) were isolated from each heart
removed from the animals in series 2. These rings were used to further evaluate the impaired relaxation
response to ACh. Vessel dimensions of the proximal and distal coronary
ring segments isolated from this group of animals are illustrated in
Table 2. No significant
differences were observed between any proximal and distal coronary ring
characteristic. As shown in Fig.
4A, relaxation
responses to ACh were significantly decreased
(P < 0.01) in the proximal coronary
rings from hearts isolated from ET rats compared with rings isolated
from C rats, confirming the results obtained in series
1 (Fig. 2). Maximal percent relaxation to ACh averaged
30.8 ± 6.2 and 74.0 ± 15.2% (P < 0.001) in rings from trained
and control rats respectively; IC50 values averaged 89.3 ± 30.6 and 88.7 ± 20.3 nM (P > 0.05), respectively. In contrast, ACh relaxation
responses obtained in the distal coronary ring (Fig.
4B) were not blunted and were not significantly different (P > 0.05) in rings from C and ET rats. Maximal relaxation in both
groups averaged 65-70% (Fig.
4B).
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5 M) was not different
in proximal and distal coronary arteries of either C or ET groups (Fig.
6), confirming results of series 1 (Fig. 3).
DISCUSSION
Exercise training has been shown to induce increases in coronary blood
flow capacity (10, 13-15) and alterations in the responsiveness of
the coronary circulation to vasoactive agents in experimental animals
(3, 5, 7, 11, 12, 19-22, 25, 35). Studies of smooth muscle
vasomotor reactivity of coronary arteries of exercise-trained animals
have produced conflicting results (3, 22, 26). Bove and Dewey (3)
reported that constrictor responses to phenylephrine were attenuated in
trained dogs, whereas Rogers et al. (26) reported that vasoconstrictor
responses to phenylephrine and NE were not altered by training. Oltman
et al. (22) reported that vasoconstrictor responses to NE were
attenuated in exercise-trained miniature swine, whereas vasodilator
responses to Ado were enhanced. Haskell et al. (8) reported that
coronary arteries of human ultradistance runners have an increased
dilating capacity compared with those of normal human subjects. The
present study represents, to our knowledge, the first report involving
the in vitro evaluation of coronary artery contraction and relaxation
function using coronary arteries isolated from exercise-trained rats.
Significant findings include: 1)
vasoconstrictor dose-response relationships to depolarization (K+) and receptor-mediated
(PGF2 and NE) contractile
mechanisms were not altered by exercise training, although sensitivity
to low concentrations of NE was increased (reduced
EC50 values); 2) vasodilator responses to SNP and
Ado were not altered by training; and
3) vasodilator responses to ACh in
proximal (but not distal) coronary arteries from ET rats were
attenuated compared with responses of coronary arteries from C rats.
There were no significant differences between the lumen diameter, axial length, or passive tension characteristics of the coronary arteries of ET and C rats. However, the mean wall thickness of arteries from ET rats (series 1) was significantly greater than arteries from C rats. This observation is consistent with the results of Segal et al. (29), who found that treadmill exercise training in rats produced 12-18% increases in the medial wall thickness in the abdominal aorta and the femoral, axillary, superior mesenteric, and coeliac arteries and increased total wall area in the aorta and the femoral and axillary arteries. There were no differences in lumen diameter in any of these arteries of ET rats compared with sedentary C rats (29). Our experiments do not reveal the mechanisms involved in this increased thickness of the arterial wall. It is possible that this represents an adaption that decreases tangential wall stress, as proposed by Segal et al.
Coronary contractile responses. Concentration-response relationships to K+ and the receptor agonist PGF2 were not altered in
coronary arteries from ET rats. Thus it appears that neither cell
membrane receptor sites for these agents nor the intracellular
signal-transduction mechanisms responsible for smooth muscle
contraction were affected in this model of exercise training.
Similarly, Oltman et al. (22) reported that contractile responses to
K+,
PGF2, and endothelin were not
altered in coronary artery rings isolated from exercise-trained
miniature swine. However, in contrast to
PGF2 and endothelin, Oltman et
al. also reported that proximal coronary artery rings isolated from
trained pigs developed less contractile tension in response to NE. Our
results indicate that this exercise training-induced reduction in
NE-induced contractions does not occur in rat coronary arteries.
Although concentration-response curves to NE were unaltered, calculated EC50 values indicated that
training unexpectedly enhanced sensitivity to lower concentrations of
NE in rat coronary arteries. Mechanisms responsible for this finding
remain unclear. Potentially, decreased 2-adrenergic-stimulated release
of NO from endothelium would enhance concomitant
1-adrenergic receptor-mediated
vasoconstrictor responses. Although Rogers et al. (26) reported that
endothelium-dependent relaxation of canine coronary arteries to
2-adrenergic agonists was
unaltered by exercise training, this model did not also exhibit enhanced contractile sensitivity to NE. In our rat model, decreased agonist-stimulated synthesis/release of endothelium-derived relaxing factor (EDRF)/NO (discussed in Coronary relaxation
responses: ACh) could theoretically contribute to
increased sensitivity to NE, at least in proximal rat coronary
arteries.
Coronary relaxation responses: SNP and Ado.
Our finding that vasorelaxation responses to Ado and SNP are not
altered in coronary arteries from ET rats surprised us, because Oltman
et al. (22) reported increased sensitivity to Ado and blunted
relaxation to SNP in isolated epicardial arteries from trained pigs.
The results of three previous studies lead to our hypothesis that
coronary arteries from ET rats would exhibit increased sensitivity to
Ado and SNP (5, 7, 13). Laughlin et al. (13) reported that coronary
blood flow was greater in the hearts of exercise-trained pigs during
maximal vasodilation with Ado. DiCarlo et al. (5) reported that
sensitivity of coronary resistance vessels to Ado, as reflected in
measurements of changes in coronary blood flow velocity in response to
intra-coronary infusion of pharmacological agents, was enhanced in
exercise-trained dogs. Also, the dilating capacity of proximal coronary
arteries was found to be greater in ultradistance runners than in
normal human subjects (7). The difference in our results and what was
expected based on the reports of Laughlin et al. (13), DiCarlo et al. (5), and Haskell et al. (8) may be due to the fact that our experiments
were performed in isolated arteries in which neural, mechanical, and
humoral influences could be eliminated, and their experiments were
conducted in conscious subjects. However, the reason training had
different effects in rats cannot be established at this time.
Coronary relaxation responses: ACh.
In contrast to the results obtained with direct smooth muscle
vasodilators (SNP, Ado), endothelium-dependent relaxation to ACh was
unaltered in distal coronaries and blunted in proximal coronary
arteries from ET rats. Furthermore, the pronounced inhibition of
maximal relaxation (Figs. 2 and 3A)
suggests a noncompetitive-like inhibitory action of training in the
proximal coronary artery of the rat that was not observed in the distal
coronary segment.
These observed differences in effects of training on ACh-mediated
responses in proximal and distal arteries were surprising and are
difficult to explain. Training-induced inhibited relaxation to ACh
cannot be attributed to the inability of the underlying coronary smooth
muscle to respond to NO or guanosine 3
,5-cyclic monophosphate (cGMP), because relaxation responses to the
cGMP-dependent NO-donor SNP were unaffected in these vessels.
Similarly, SNP relaxation responses also indicate that vessel wall
restructuring (increased vessel wall thickness) is unlikely to
compromise the ability of the smooth muscle to maximally relax to NO,
because unaltered and complete (100%) relaxation was attained to SNP
in these rings. Furthermore, in series
2, no significant differences were observed in vessel
ring dimensions betweeen proximal and distal ring segments (Table
2). However, in these experiments we noted that the
interventricular coronary artery traversed the ventricle with few
branches until near the apex. The distal segment was anatomically more
intramyocardial than was the proximal segment. This suggests the
possibility that the two segments were potentially exposed to differing
levels of vasoactive metabolites or other unknown factors released from
contracting myocardium during bouts of exercise. In addition to these
potential in vivo differences, underlying cellular mechanisms for this
unexpected finding remain to be determined. Cellular mechanistic
possibilities include altered endothelial
Ca2+ mobilization
[producing decreased
Ca2+-dependent activation of
endothelial cell NOS (ecNOS)]; differential ecNOS downregulation
in proximal vs. distal coronary arteries, and/or altered
receptor-second messenger coupling mechanisms in rat proximal coronary
arteries. Impaired maximal responses, however, suggest
that competitive inhibitory actions at endothelial muscarinic receptor
sites are not involved (because increased levels of agonist did not
overcome the training effect).
A recent study of Wang et al. (35) reported that endothelium-dependent
vasodilation was enhanced by 7 days of exercise training. However, two
other studies report that exercise training did not alter
endothelium-dependent responses of conduit coronary arteries (23, 26).
Rogers et al. (26) reported that responsiveness of isolated dog conduit
coronary arteries to substance P and
2-adrenergic agonists was
unchanged by training, and Oltman et al. (23) reported that endothelium
mediated vasodilation of epicardial porcine coronary arteries was not
altered by 12-16 wk of exercise training. In these studies,
vasodilator responses to bradykinin, substance P, clonidine, serotonin,
and the calcium ionophore A-23187 were all similar in coronary arteries
of trained and control pigs (23). Similarly, McAllister et al. (16)
recently reported that the same model of exercise training did not
alter endothelium-dependent vasodilation of porcine peripheral and
mesenteric arteries. The difference between the results of Wang et al.
(35) and those of the present study, Rogers et al. (26), and Oltman et
al. (23) could be due to one or more of the following:
1) Wang et al. examined ACh-induced
vasodilation of coronary arteries in conscious dogs, whereas we
examined responses in vitro; 2)
there were species differences; and
3) duration of the exercise training programs was different [1 wk for Wang et al. (35); 12 wk in the
present study and that of Oltman et al. (23)]. We will discuss each of these possible explanations below.
First, we chose to use isolated coronary arteries in our experiments
because this in vitro approach eliminates metabolic, neural,
mechanical, and humoral influences on coronary tone that may be present
to a variable extent in vivo. In the experiments of Wang et al. (35),
infusion of ACh may have increased intracoronary flow, producing
flow-induced vasodilation. To test for this possibility, Wang et al.
demonstrated that ACh-induced vasodilation was greater in coronary
arteries of trained dogs even when flow was held constant. This
suggests that training-induced changes in flow-induced vasodilation do
not contribute to these changes. However, metabolic, neural, and
humoral influences present in the in vivo experiments of Wang et al.
may be at least partially responsible for the findings different from
those of the present study. Second, it is also possible that exercise
training produces enhanced endothelium-dependent vasodilation in
coronary arteries of dogs but not in rat coronary arteries. The fact
that exercise produces four- to fivefold increases in coronary blood
flow in dogs but only increases coronary blood flow by 20-30% in
rats is consistent with this notion (6). Rats have relatively high
resting coronary blood flows (6). Thus, if flow and/or shear
stress are signals for adaptation, the signal produced by exercise is
less for rats. Also, training-induced bradycardia has been shown to be
more modest in rats than in dogs. For example, we have shown that a
treadmill training program similar to that used in the present study
produced only a 6% decrease in resting heart rate of rats (28).
Because of lower resting heart rates in large mammals (such as dogs,
pigs, and humans), training produces a 20-30% decrease in resting
heart rate (28). Thus exercise training may not produce enhanced
endothelium-mediated vasodilation in the coronary arteries of rats, at
least partly because training appears to have less impact on the heart
of rats than on the hearts of larger mammals. However, Rogers et al.
(26) reported that exercise training of dogs for 11 wk did not alter endothelium-dependent vasodilator responses of
2-adrenergic agonists or
substance P. This observation argues against the notion that these
conflicting results are due to species differences, or alternatively, that training-induced enhanced endothelium-dependent relaxation responses are agonist specific for muscarinic receptors in dogs.
Third, the present study and those of Rogers et al. (26) and Oltman et
al. (23) examined coronary vasodilator responses in animals that appear
to have attained steady state of adaptation to exercise training as
reflected in increased skeletal muscle oxidative capacity. The training
programs were of sufficient duration to ensure that adaptations would
be expected to have been completed, whereas Wang et al. (35) purposely
conducted their experiments after only 1 wk of training, before these
training-induced adaptations would have occurred. Exercise
training-induced adaptations are time-dependent processes that are
initiated by acute bouts of exercise and reach completion after a
period of time (2, 27). For example, training-induced proliferation of
capillaries and mitochondria in skeletal muscle appears to commence
shortly after the initial bouts of exercise and to be complete after
4-8 wk of training (2, 27). It is possible that coronary vascular adaptation occurs in a similar, progressive, time-dependent manner. If
this is true, then early in the adaptive process, repetitive bouts of
exercise (two 1-h bouts/day in the study of Wang et al.) may produce a
relatively rapid upregulation of ecNOS, resulting in enhanced
endothelium-dependent vasodilation as reported by Wang et
al. Thus endothelium-mediated control of coronary artery diameter may be enhanced early in the exercise-adaptive process but
returns to normal later in the training period, when structural adaptations (11, 36) return coronary shear stress during exercise to
lower levels.
Effects of inhibition of NOS and cyclooxygenase.
ACh-induced dilation has been shown to be a receptor-mediated,
endothelium-dependent response in arteries of both rats and dogs (8, 9,
34). Although debate continues concerning whether EDRF-NO accounts for
all or part of ACh-induced vasodilation (9), Tschudi et al. (34)
reported that ACh does not stimulate the cyclooxygenase system in rat
coronary arteries. Indomethacin did not alter ACh-induced vasodilation
in rat coronary arteries (34), whereas treatment with
L-NMMA markedly reduced the
maximal response to ACh and
N
-nitro-L-arginine methyl ester
blocked ACh-induced vasodilation in rat coronary artery. Therefore, in
the present study we chose to investigate the possibility that
ACh-induced vasodilation was attenuated after exercise training (in
proximal arteries) because of a decrease in the release of NO from
coronary arteries of trained rats.
L-NMMA, an inhibitor of NOS
(24), produced inhibition of ACh-induced relaxation in vessels from ET
and C rats (Fig. 5), confirming previous results of Tschudi et al.
(34). Also, treatment with
L-arginine did not improve
ACh-induced relaxation in proximal coronary arteries from ET rats,
suggesting that the depressed response is not the result of
insufficient substrate for NOS. However, it is possible that the
increased wall thickness would result in increased diffusion distance
for EDRF-NO so that the measured sensitivity of vasodilator responses
of the vessels to ACh would decrease (although maximal responsiveness
would likely remain unaffected). This explanation, however, appears
unlikely because reduced sensitivity to ACh was not observed in
series 2, although maximal relaxation
was significantly reduced.
Integration of findings: Role of chronic flow and shear stress
changes in coronary adaptation to exercise training.
Delp et al. (4) reported enhanced ACh-induced
vasorelaxation of aorta isolated from rats trained with an exercise
training program similar to the one used in the present study. These
results indicated that exercise can produce enhanced
endothelium-dependent vasorelaxation in arteries of the rat. Miller and
colleagues (17, 18) demonstrated enhanced, endothelium-dependent
relaxation to ACh, Ado diphosphate, and
2-adrenergic stimulation in
canine femoral arteries as a result of chronic increases in blood flow. Chronic increases in blood flow were produced with arteriovenous fistulas, which resulted in increased femoral arterial blood flow proximal to the fistula (17, 18). Delp et al. (4) argued that blood
flow was the primary signal for adaptive increases in
endothelium-dependent vasodilation in their exercise-trained rats. This
argument is supported by recent studies documenting that chronic high
blood flow in vivo (via arteriovenous fistula) significantly enhances
NOS mRNA, immunoreactive NOS (Western blots), and
Ca2+-dependent NOS activity in rat
aorta (21). Similarly, Sessa et al. (30) documented increased NOS mRNA
expression in the aorta of dogs after chronic exercise training. It is
possible that the primary reason that exercise training did not produce an increase in ACh-induced vasorelaxation in the proximal coronary arteries of our trained rats is that this exercise model does not
produce a large enough (or sufficiently prolonged) increase in coronary
blood flow and/or shear rate in the coronary artery of rats to
stimulate these adaptive responses. Treadmill exercise only produces a
modest increase above baseline values in rat coronary blood flow (6).
In contrast, exercise produces a 200-300% increase in blood flow
through the rat abdominal aorta (4). This is in the range of increased
flow reported by Miller et al. (17, 18) in dogs. Thus available
information indicates that this intensity of exercise does not produce
a large enough increase in coronary blood flow in rats to signal the
type of adaptations reported to occur in endothelium-mediated vascular
control in the aorta of exercise-trained rats.
The present results indicate that in the proximal portion of the
coronary artery, ACh-induced vasodilation is attenuated after exercise
training in rats. If this change in endothelium-dependent vascular
control is also related to changes in flow (or shear stress), it
suggests that flow (shear stress) may be decreased by training. Because
coronary blood flow in rats is known to be increased by 20-30%
(6) during treadmill exercise, these observations suggest that coronary
blood flow is decreased in trained rats during the nonexercising time
(23 h/day) when the rats are not training. We have no data that allow
critical evaluation of this possibility. Furthermore, the physiological
role of NO during bouts of exercise is unclear, in view of a recent
report indicating that blockade of NO synthesis does not alter coronary
blood flow during acute exercise in conscious dogs (1). Also, despite evidence for altered NOS regulation in response to chronic exercise or
flow/shear stress stimuli (1, 4, 17, 18, 21, 30), the functional
significance of these findings has not been clarified. Although shear-stress-induced release of NO (via increased flow) may
indeed contribute to exercise-induced vasodilation, other critical
roles of NO have been proposed, including modulation of myocardial
metabolism (1) and structural remodeling of the arterial wall during
chronic adaptations to flow-induced changes in shear stress (21).
Conclusion.
The results of this study indicate that exercise training does not
alter the passive compliance characteristics of rat proximal coronary
arteries, although the wall thickness of coronary arteries from trained
rats was increased. In addition, contractile responses induced by KCl,
PGF2, and NE and vasorelaxation
responses induced by SNP and Ado were similar in coronary arteries from ET and C rats (although sensitivity to low concentrations of NE was
increased by training). ACh-induced vasorelaxation was attenuated in
the proximal coronary artery of trained rats, whereas ACh-induced vasorelaxation was similar in the distal portion of the coronary arteries of trained and control rats. The blunted ACh response of
proximal coronary arteries was not altered by treatment with L-arginine or indomethacin.
These results suggest that endothelium-mediated vasorelaxation in
response to ACh is blunted in proximal (but not distal) coronary
arteries isolated from rats exercise trained at the intensity used in
this study.
ACKNOWLEDGEMENTS
The authors thank Donna Baumgartner for assistance in exercise training the animals and Lori Hoyt and Beth Farrow for excellent assistance in preparation of the manuscript for submission.
FOOTNOTES
Address for reprint requests: J. L. Parker, Dalton Cardiovascular Research Center, Univ. of Missouri, Columbia, MO 65211.
Received 7 May 1996; accepted in final form 4 April 1997.
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