Clinical Science (1992) 82, 55-62 (Printed in Great Britain)
55
A comparison of the pharmacological and mechanical
properties in vitro of large and small pulmonary arteries
of the rat
R. M. LEACH, C. H. C. WORT, 1. R. CAMERON and J.
P. T. WARD
Respiratory Research Laboratory, Division of Medicine, UMDS ( S t Thomas’ Campus), London, U.K.
(Received 14 Januaryl3 July 1991; accepted 31 July 1991)
INTRODUCTION
1. The mechanical and pharmacological properties of
small pulmonary arteries (100-300 p m normalized
lumen diameter) were directly compared with those of the
left main pulmonary artery (1-2 mm) from the rat. The
active and passive length-tension characteristics and
responses to a variety of agonists and antagonists were
dependent on arterial diameter.
2. Maximum contractile function was obtained in both
groups of vessels when stretched so as to give an equivalent transmural pressure of 30 mmHg. This is substantially lower than that found for systemic vessels, and
reflects the normal low pulmonary arterial pressure.
3. Noradrenaline was a powerful vasoconstrictor in large
but not small pulmonary arteries ( P < 0.001). In contrast,
bradykinin produced a significantly greater response in
the small arteries (P<O.OOl). In comparison with large
pulmonary arteries, small arteries were more sensitive to
noradrenaline ( P < 0.05) and 5-hydroxytryptamine
(P< 0.001), less sensitive to endothelin-1 ( P < 0.001)
and had the same sensitivity to prostaglandin Fza.
4. The mechanism that maintains the low arterial tone of
the pulmonary circulation is unknown, but it may involve
the release of relaxing factors from the endothelium. In
this preparation, basal resting tone could not be demonstrated in either large or small arteries.
5. Acetylcholine-induced relaxation of pre-contracted
pulmonary arteries was reduced or absent in the small
artery, despite histological evidence of an intact endothelium. In large arteries pre-contracted with
prostaglandin FZa, acetylcholine (100 pmol/l) caused
88.2% relaxation compared with 25.2% in the small
artery.
6. These results suggest that it would be unwise to
extrapolate from results obtained either in large pulmonary artery preparations or in perfused lungs to what
may occur in the functionally important resistance
arteries.
In the systemic circulation, the properties of arteries
alter with decreasing diameter. Small muscular resistance
arteries have different receptor properties, dependence
on extracellular Ca2+,resting basal tone and sensitivity to
non-adrenergic neurotransmitters when compared with
large elastic conduit arteries [l, 21. There has been
comparatively little investigation of the small pulmonary
arteries, but indirect evidence suggests that they may have
an important role in the control of the pulmonary circulation [3,4].Previous investigations in the pulmonary circulation have been conducted primarily in isolated lung
perfusion experiments o r using catheterization techniques
in the intact animal [5-71. Data from studies in vifro of
isolated conduit pulmonary arteries and veins are often at
variance with the findings documented in whole-lung perfusion experiments, and may reflect differences in the
vasoreactivity of the specific components of the vascular
bed. The powerful vasoconstriction in response to
catecholamines observed in isolated pulmonary artery
preparations is not observed in either the pulmonary
circulation of the intact animal or the perfused lung
preparation [5,6] and varies according to the degree of aand P-adrenoceptor stimulation and the initial resting
tone. Vascular occlusion techniques in perfused lung
experiments also provide evidence for differential activity
of the individual compartments of the pulmonary circulation. The potent HI-mediated vasoconstrictor action of
histamine on the pulmonary circulation is located
primarily in the venous circulation with only a minimal
effect in pulmonary arteries [8-lo]. When pulmonary
vascular tone is raised, a H,-mediated histamine vasodilatation may occur in arteries and small veins, but the
vasoconstrictor response is maintained in the large veins
[lo].
Using a Mulvany/Halpern small-vessel myograph [ 111,
the contractile and pharmacological responses of large
and small pulmonary arteries have been compared. We
Key words: pulmonary circulation, pulmonary vascular reactivity, small-vessel myography.
Abbreviations: EC, concentration required to produce half-maximal tension; EDRF, endothelium-derived relaxing factor; K+PSS, physiological salt solution in which KCI
replaced NaCI; PSS, physiological salt solution (see the text for composition).
Correspondence: D r R. M. Leach, Division of Medicine, S t Thomas’ Hospital, Lambeth Palace Road, London SEI 7EH.
56
R. M. Leach et al.
have previously demonstrated that arteries with an
internal diameter of 200-400 p m produce considerably
more force in response to K + depolarization than those
either smaller (100-200 p m ) or larger (400-2000 pm),
and may play a role in the generation of pulmonary
vascular resistance [3].
METHODS
Vessel mounting
Male Cummin Sprague Europe rats (250-350 g) were
anaesthetized with ether, and the heart and lungs were
removed. The left main pulmonary artery (1-2 mm
diameter) and a small pulmonary arterial vessel (100-300
p m diameter) were mounted as ring preparations on a
dual (single bathing chamber) myograph. Dissection was
performed under a binocular x 40 operating microscope.
The bronchial tree was dissected open along its path and
then gently lifted and dissected free from the artery
beneath [3]. The lung tissue surrounding the remainder of
the artery was dissected free without touching the artery.
Arteries were mounted as described previously [3, 111.
The vessels were bathed in physiological salt solution
(PSS) containing (mmol/l): NaCl 118, NaHCO, 24,
MgSO, 1, NaH2P0, 0.435, glucose 5.56, sodium pyruvate 5 , CaCI, 1.8 and KCl 4, and equilibrated with 95%
0 , / 5 % CO, (pH 7.35) at 37°C. After an equilibration
period of 1 h, the resting tension-internal circumference
characteristics of the vessel were determined and the
internal circumference of the vessel was adjusted to give
maximum developed tension (see normalization procedure below).
Normalization
Mean pulmonary transmural pressure is considerably
less than in the systemic circulation [4,5] and the normalization procedure was modified to adjust for this difference but was otherwise identical with that described
for systemic vessels [ 111. Length-tension experiments
demonstrated that maximum developed tension in
response to K + (75 mmol/l) depolarization occurred at a
resting internal circumference (L,,) at which the calculated equivalent transmural pressure was 30 mmHg in
both large and small pulmonary arteries (see Fig. 1).In the
normalization procedure, the internal circumference ( L )
was set to L,,, rather than to 0.9L,,,,, as described in
systemic arteries, to prevent excessive stretching of the
vessels. From the L,,, an estimate of the cylindrical lumen
diameter (d30) that the vessel would have had when
relaxed and under a transmural pressure of 30 mmHg can
be determined (d,, = L 3, /x) .
Experimental protocols
In order to assess the length-tension characteristics in
large and small pulmonary arteries, two groups of vessels
with similar internal diameters were compared. Ten left
main pulmonary arteries (1.5-2.0 mm diameter) were
compared with 10 intrapulmonary arterial resistance
vessels (100-300 p m diameter). The internal circumference ( L) of the arteries bathed in a relaxing solution was
increased in a stepwise manner and the passive tension
developed was recorded until the wall tension was about
2 mN/mm. After a relaxation period of 1 h, the developed
tension to maximal K + depolarization (75 mmol/l KCI)
was determined at identical internal circumference steps
to those used in the resting tension-internal circumference experiment. The 75 mmol/l KCI PSS (K+ PSS) solution was prepared by equimolar substitution of KCI for
NaCl in normal PSS. At each step the resting tension was
allowed to plateau for 2 min before the vessel was
exposed to K + PSS and the tension was recorded after 2
min. The artery was then washed with PSS for 5 min and
allowed to relax to the resting baseline tension before the
next stepwise increase in internal circumference.
After normalization the arteries were rested for 30 rnin.
The vessels were then preconditioned by repeated 2 min
exposures to K + PSS before being returned to PSS. The
concentration-response characteristics of the arteries to
K+ depolarization before and after preincubation with
cocaine were determined. Cocaine blockade of the amine
pump prevents perivascular adrenergic nerve endings
modulating the effects of exogenous noradrenaline and
potassium on the isolated artery [ 121. Depolarizing solutions of progressively increasing K + concentration
(10-100 mmol/l) were added to the bathing chamber at 2
rnin intervals after emptying the bath of the preceding K+
depolarizing solution. Maximum developed tension
occurred at 75 mmol/l KCI (K+ PSS).
CaZ+dose-response curves were assessed in K + PSS
solution. The vessels were initially preincubated in a
Ca2+-free PSS solution containing 0.5 mmol/l EGTA.
The bathing solution was replaced with Ca2+-freeK + PSS
( + 0.5 mmol/l EGTA) and subsequently with solutions
containing increasing concentrations of Ca2+ (without
EGTA). The response was allowed to plateau between
each concentration. Higher concentrations were not
assessed because this resulted in Ca2+precipitation in the
K + PSS solution.
The concentration-response characteristics of the
vessels to noradrenaline, 5-hydroxytryptamine, prostaglandin FZa,endothelin-1, histamine and ADP were
determined by exposing the vessels to increasing concentrations of the activating solutions at 2 min intervals by
adding drugs directly to the bathing solution. The
responses to bradykinin (1x lo-’ mol/l) and angiotensin
I1 ( 5 x lo-‘ mol/l), vasoconstrictors in the rat pulmonary
circulation, were assessed at a single concentration as
these drugs did not produce plateau responses and
demonstrated tachphylaxis after the initial contraction.
The results are reported as developed tension in mN/mm
vessel length at each concentration, and the maximum
response as a percentage of the response to K + PSS. The
effect of adrenergic innervation on the responses to noradrenaline was assessed by preincubation of the arteries
with cocaine (3 x
mol/l, IZ = 4). The consequences of
a- and P,-adrenoceptor blockade on the contraction in
response to noradrenaline were also assessed by pre-
Pharmacological and mechanical properties of pulmonary arteries
57
incubation of the vessels with phentolamine (1x
mol/l, n = 4 ) and practolol (1x
mol/l, n=4),
respectively.
Acetylcholine-induced relaxation was assessed in
stable pre-contracted small and large pulmonary arteries.
The arteries were pre-contracted with noradrenaline
(3x
mol/l), prostaglandin F,, (1x
mol/l),
endothelin-1 (1x
mol/l) or K+ PSS solution. Acetylcholine was added directly to the myograph bath to
produce increasing concentrations ( 10-y-10-4 mol/l) in
the bathing solution. The degree of relaxation is reported
as a percentage reduction of the stable pre-contraction.
Materials
All chemicals were supplied by Sigma (Poole, Dorset,
U.K.), except phentolamine (Regitine; Ciba, Basle,
Switzerland), propranolol (Inderal; ICI, Alderley Park,
Macclesfield, Cheshire, U.K.), cocaine chloride (May and
Baker, Dagenham, Essex, U.K.) and porcine endothelin-1
(PeninsulaLaboratories, Belmont, CA, U.S.A.).
I .o
0.8
I .2
I .4
Internal circumference (l/L30)
Statistical analysis
All values are mean sf^^^. Data were compared by
using Student's t-test. The concentration required to
produce half-maximal tension (EC,,) was calculated using
a commercial software package (Enzfitter; Biosoft,
Cambridge, U.K.) and expressed as the pD,( - logEC5,)
for the purposes of statistical comparison.
5.0
-
4.0
-
E
RESULTS
iP
T
Z
(b)
3.0
-
T
E
v
PT
Active and resting tension-internal circumference
relationships
Fig. 1 demonstrates the passive tension-internal
circumference characteristics and the active developed
tension (to maximum K+ depo1arization)-internal
circumference characteristics in both large and small
pulmonary arterial vessels. In all subsequent experiments
the passive tension-internal circumference characteristics
were determined and the internal circumference was
adjusted to give a calculated resting transmural pressure
of 30 mmHg at which the maximum active tension is
developed.
Basal tone and Kt and Ca2+sensitivity
Resting tone was not demonstrated in either large or
small pulmonary arteries. Preincubation in Ca*+-free
solutions or Ca2+-freesolutions with EGTA (0.5 mmol/l)
did not alter the steady-state baseline resting tension.
Exposure to acetylcholine ( lo-, mol/l), propranolol
(3x
mol/l), phentolamine
mol/l), nitroprusside (5 x l o V hmol/l), verapamil (1X
mol/l), or
caffeine (10 mmol/l) also had no effect on baseline
tension. After pre-contraction of the arteries with K+
PSS, Ca2+-free solution, nitroprusside (5 x
mol/l)
I
0.8
I
I .o
I
P
1
I
I .2
I
I
I .4
Internal circumference (1/l.3G)
Fig. I. Passive resting tension ( 0 ) and active tension in
response to maximum K+ depolarization ( 0 ) at increasing
internal circumference (expressed as relative internal circumference, L/L,,) for small (100-300 pm, n = 10, 0 ) and large (1-2
mm, n = 10, b) pulmonary arteries. The broken line represents the
30 mmHg isobar.
and verapamil (1 X
mol/l) all resulted in complete
relaxation.
Fig. 2 demonstrates the concentration-response to K+
depolarization as measured tension in mN/mm. In both
large (diameter 1821 f 4 8 pm, n = 8 ) and small arteries
(diameter 2 7 7 f 2 6 pm, n = 8 ) maximum tension was
developed at 70 mmol/l [K+]with an ECsoof 27.1 mmol/l
R. M. Leach et at,
58
KCI (pD, 2.42 f0.03) in the large and 34.6 mmol/l KCI
(pD, 2.54f0.02; P < O . O l ) in the small vessel. Fig. 3
illustrates the Ca2+ concentration-response in K+ PSS in
terms of measured wall tension in mN/mm. The EC,,
values in large (diameter 1 8 9 0 f 4 2 , n = 6 ) and small
(diameter 254 f22, n = 6) arteries were 0.27 mol/l CaZ+
(pD2 0.62f0.11) and 0.49 mmol/l CaZ+ (pDz 0.32
f0.03), respectively ( P < 0.05).
Response to agonist stimulation
Fig. 4 compares the concentration-response curves to
noradrenaline ( IZ = 15), 5-hydroxytryptamine ( n= 1l),
prostaglandin FZa ( n= 17) and endothelin-1 ( n= 7) as
developed tension (mN/mm) in large (1-2 mm) and small
(100-300 p m ) pulmonary arteries. The maximum
responses in large and small arteries, as a percentage of
the contraction with 75 mmol/l KCI, were for noradrenaline 93.9 f7.1% and 6.35 k 2.2%, respectively, for
5hydroxytryptamine 41.6 f 4.6% and 11.1f 1.7%,
respectively, for prostaglandin F2a 79.8 f 4.8% and
84.3 f5.4%, respectively, and for endothelin-1
174 f 10% and 121 k 11%, respectively.
In the small arteries there was a small response to
noradrenaline in seven out of 15 experiments and the
EC,, and mean dose responses were determined from
these experiments. In eight small arteries there was no
response to noradrenaline when stimulated at baseline
resting tone. After partial pre-contraction of the small
arteries with either KCL (35 mmol/l) or prostaglandin F,,
(1 X
mol/l), we were unable to demonstrate either
potentiation of the response to noradrenaline or relaxation in those arteries that did not respond. In the large
arteries ( n= 15), low concentrations of noradrenaline
produced contraction, but when the concentration in the
bathing solution was increased above 3 x
mol/l,
partial relaxation of the previous plateau contraction was
observed. Preliminary investigations ( n = 4) suggest that
fi-adrenoceptor blockade with practolol (1x 10-6mol/l)
will reverse the relaxation observed with high concentrations of noradrenaline or prevent it after preincubation. In
large arteries, a-adrenoceptor blockade with phentolamine (1x los6 mol/l) completely abolished the response
to noradrenaline in both pre-contracted arteries and after
preincubation with phentolamine before the addition of
noradrenaline ( n= 4). Blockade of the perivascular
adrenergic nerve endings by preincubation with cocaine
(5 x
mol/l) did not alter the response to noradrenaline at any concentration.
Prostaglandin F,, produced stable plateau responses in
both the large and small arteries, which returned rapidly
to the baseline on washing with PSS. In comparison, the
contraction in response to endothelin-1 in the small artery
produced a plateau response lasting several hours which
could not be reversed by repeated washing. In contrast,
the response in the large artery slowly diminished to the
baseline over a period of 45 min, even in the continued
presence of the drug. 5-Hydroxytryptamine-inducedtone
gradually diminished in both large and small vessels.
Bradykinin produced a powerful but transient contraction in the small but not the large artery. The maximum
response to bradykinin (1x
mol/l) was 1.15f0.13
mN/mm (62f4.4% of maximum K+ depolarization) in
small arteries (diameter 2 4 8 f 15 ,urn, n = 17) and
0.07f0.01 mN/mm (5.2+0.8% of maximum K+
2.0
2.5
2.0
I .5
E
E
t
E
I .5
v
c
:.
1.0
al
c
u
r”
0.5
I
I
0
I
I
I
I
2
3
I
4
[Cali] (rnmol/l)
[KCI] (mmolll)
Fig. 2. Response to Kt depolarization in large (1-2 m m
diameter, n=8, 0) and small (100-300 pm diameter, n=8, 0 )
pulmonary arteries expressed in terms of wall tension
Fig. 3. Response to increasing Ca” concentration in Kt PSS for
large (1-2 mm diameter, n=6, 0 ) and small (100-300 pm
diameter, n=6, 0 ) pulmonary arteries expressed in terms of wall
tension
59
Pharmacological and mechanical properties of pulmonary arteries
T
J
I
r
T
Table I. Maximum developed tension to agonlst stimulation in
large and small pulmonary arteries. Abbreviation: NS, not significant.
!I
Agonist
i
Prostaglandin F,,
1
I
I
1’/ ;
I
I
7
Noradrenaline
I
6
/
i/
5-Hydroxytry ptamine
Endothelin-I
r T
7
6
5
4
3
(b)
1
T
2.0
7,
?
t
E
v
0
c
.-
2
Y
I
0
9
I , I9 fO.07
0.58f0.08
( n = I I)
2.01 f0.13
I .99 f0.33
17)
0.08 f 0 . 0 2
(n=7)
0.23 f 0 . 0 3
( n = I I)
2.18k0.41
(n=7)
< 0.05
(0.00 I
<O.OOI
NS
8
7
6
5
- log([Agonist] (mol/l)}
histamine or ADP. In comparison with large arteries,
small pulmonary arteries showed a significantly smaller
(P<0.001) response to maximum noradrenaline stimulation (1x
mol/l), but a significantly larger ( P < O . O O l )
response to maximally effective bradykinin (1x
mol/l) (Table 1). The relative potencies of noradrenaline,
5-hydroxytryptamine,prostaglandin F2aand endothelin-1
in large and small pulmonary arteries are shown in Table
2. In comparison with large pulmonary arteries, small
arteries were more sensitive to noradrenaline (P<0.05)
and 5-hydroxytryptamine (P<0.001), less sensitive to
endothelin-1 ( P < O . O O l ) and had the same sensitivity to
prostaglandin Fza.
Fig. 5 shows acetylcholine-induced relaxation of large
and small pulmonary arteries, after pre-contraction with
K + PSS, noradrenaline, prostaglandin Fzaand endothelin1. In small arteries, acetylcholine only caused relaxation
after pre-contraction with prostaglandin Fza;there was no
relaxation in response to acetylcholine after pre-contraction with K + PSS or endothelin-1, but small transient
contractions were observed. In the presence of noradrenaline, acetylcholine resulted in similar transient
contractions to those described above in eight out of 15
experiments. Histological examination of eight large and
eight small arteries after experimentation confirmed the
presence of an intact endothelium.
I
I .5
(n=
Acetylcholine-induced relaxation
I
E
E
I .23 f 0 . 0 7
(n = 17)
(n=7)
- log([Agonist] (mol/l)}
2.5
Small artery
(n= 15)
1
8
9
Large artery
P value
A
9
10
Maximum tension
(nM/mm)
4
3
Fig. 4. Response to increasing concentrations of endothelin-l
(n=7, O), noradrenaline ( n = IS, 0), 5-hydroxytryptamine
(n=ll, A ) and prostaglandin F,, (n=l7, A) In small (100-300
gm diameter, a) and large (1-2 mm diameter, b ) pulmonary
arteries expressed as wall tension
depolarization) in large arteries (diameter 1702 k 50 pm,
n = 19). Angiotensin I1 produced small transient contractions ( < 10% of maximum K + depolarization) in both
large and small vessels, but there was no response to
DISCUSSION
Small arteries from a variety of vascular beds have properties that reflect the specialization of those systems [l,
131. Until recently, most studies of pulmonary blood
vessels in vitro were performed on larger arteries and
veins and the results, although important, may not reflect
the mechanical and pharmacological properties of the
smaller vessels. Experiments in the isolated whole-lung
perfusion preparation and in the catheterized pulmonary
circulation of the intact animal have attempted to define
the role of the individual components of the pulmonary
circulation, including the muscular arteries, arterioles,
R. M. Leach et al.
60
Table 2. pD, (and EC,,) values for agonist concentration-response curves in large and small pulmonary arteries.
Abbreviation: NS, not significant.
Agonist
Prostaglandin F,,
Noradrenaline
5-Hydroxytryptamine
Endothelin-I
PD, (ECSO)
Large artery
Small artery
4.84k 0.08
(14.5pmolll)
4.99to.05
(10.3pmolll)
(n = 17)
(n= 17)
6.45 k0.I6
(0.35pmolll)
7.04 k0.22
(0.09pmolll)
(n= 15)
(n=7)
4.48t0.10
(33.1pmolll)
5.49k0.06
(3.24pmolll)
(n= I I)
(n=ll)
8.05k 0.06
7.45k0.02
(8.9I nmolll)
(n=7)
(33.I nmolll)
100 -
P value
80 -
NS
h
ae
(0.05
0
c
.Y
c1
Y
<o.oo I
8
a
8
3
Y
c
c
<0.001
(n=7)
venules and veins [8, 131. The small-vessel myograph has
not previously been used to study the pulmonary
circulation.
T h e length-tension experiments performed on large
and small pulmonary arteries demonstrate that maximal
vessel response to K + depolarization is achieved at a
transmural pressure equivalent to 3 0 mmHg, considerably less than that described in vessels from the
systemic circulation [ 111. Although 30 mmHg is greater
than the normal mean pulmonary artery pressure of
15-20 mmHg previously described in the rat [5], we have
elected to perform our experiments at this degree of
stretch to ensure maximal contractile function and experimental repeatability.
We have examined the physiological and pharmacological responses of isolated large and small pulmonary
arteries to determine whether arterial size may influence
its properties and its role in the pulmonary circulation.
Our results demonstrate that the responses to various
agonists and antagonists differed between large and small
arteries. In particular, noradrenaline produced powerful
contractions in the large, but little o r no response in the
small, pulmonary artery. Conversely, bradykinin had the
opposite effect. Our unpublished observations suggest
that there is a progressive change in responsiveness to
noradrenaline and bradykinin in arteries of intermediate
size. In order to understand the physiological significance
of such differences, the results of these experiments must
be discussed in conjunction with previous studies on
isolated lungs and itz vivo.
Pulmonary vascular resistance is probably primarily
dependent on the smaller arterial vessels [3, 141, although
there will be a contribution from larger arteries and the
venous circulation. Catecholamine &nulation of perfused lungs, both as isolated preparations and in the intact
animal, results in a small (2O-4Oo/0) increase in pulmonary
vascular resistance [7,15]. This is highly dependent on the
underlying basal tone, autonomic innervation and a- and
@-adrenoceptor blockade [S, 7, 151. Our results suggest
.-0
w
-23
d
- log([Acetylcholine](molll))
100-
80 -
'.\
I\
7.
-
Ec
0
.e
c
8
Y
60 -
Y
-al
D
m
Y
L
.E
40 -
Y
23
d
20 i
0'
9
I
8
I
7
I
6
I
5
I
4
- log{[Acetylcholine](molll))
Fig. 5. Percentage relaxation of the stable contraction produced
by K' (75 mmolll, n = 6, A), endothelin-l (10.' molll, n = 5, V),
noradrenaline (3 pmolll, n = 13, A) and prostaglandin F,, (lo-'
molll, n =9, 0) in small (100-300 pm diameter, a ) and large (1-2
mm diameter, b) pulmonary arteries for increasing concentrations of acetylcholine
Pharmacologicaland mechanical properties of pulmonary arteries
that these small increases in resistance are related to constriction in the larger arteries, as the small vessels d o not
respond to noradrenaline (Fig. 4). The mechanism underlying this variation in response in vessels of differing size
may relate to either receptor density o r affinity [ l , 2, 151,
In this study, even at noradrenaline concentrations of
mol/l there was little o r no response in small
arteries, suggesting that the response is receptor-limited.
Further investigation of receptor occupancy and density
are required. In the large artery, a-adrenoceptor blockade
with phentolamine completely abolished the constrictor
response to noradrenaline. At high concentrations, noradrenaline caused relaxation of the pre-contracted large
artery. p,-Adrenoceptor blockade with practolol
prevented or reversed this relaxation. T h e lack of
constrictor response in the small arteries was not due to
an overlying P,-adrenoceptor-mediated vasorelaxation,
as practolol did not result in constriction. Both noradrenaline and 5-hydroxytryptamine may, however, have
additional effects that result in release of endotheliumderived relaxing factor (EDRF ) from the endothelium, as
has been found in systemic arteries [16].
The pulmonary vascular bed is in general well supplied
with adrenergic nerves, and sympathetic stimulation can
increase pulmonary vascular resistance [ 15, 171. Specific
staining methods for adrenaline and acetylcholinesterases
in the rat pulmonary circulation suggest that adrenergic
sympathetic innervation is confined to the extrapulmonary arteries [ 181. Even at noradrenaline concentrations approaching those reported in the synaptic cleft
(
mol/l) [2, 151we have been unable to demonstrate a
response to noradrenaline in the small pulmonary artery.
The absence of adrenergic nerves in the rat pulmonary
circulation may explain the differences in the response of
the large and small pulmonary arteries to noradrenaline.
It would also imply that sympathetic innervation of the rat
pulmonary circulation is dependent on alternative neurotransmitters, for example neuropeptide Y, which is
released from many sympathetic nerves [ l , 19,201.
Endothelin-1, a recently described vasoconstrictor
peptide, produced a potent and irreversible dose-dependent contraction in the small artery. The large artery was
significantly more sensitive to endothelin-1, but the
response was not sustained, returning to the baseline even
in the continued presence of the drug over a period of
45-60 minutes [21]. In comparison, application of endothelin isopeptides to the intact perfused lung may induce
either moderate and reversible increases in pulmonary
vascular resistance [22], o r where pulmonary vasomotor
tone is already increased, a reduction in vascular resistance [23]. The difference between the effect in isolated
vessels and that in lungs may be related to the fact that
endothelin-1 can release other vasodilators from isolated
lungs, and is itself substantially removed from the circulation by the lungs both in vivo and in vitro [24]. Endothelin-1 is produced by the endothelium and would be
expected to act as a local mediator with a role in regional
perfusion, rather than as a circulating agent. The results of
exogenous application into the circulation may not therefore reflect is physiological function.
61
The response of these isolated arteries to 5-hydroxytryptamine was small compared with the powerful vasoconstriction reported for whole-lung preparations [25].
The large artery developed greater tension, but was less
sensitive to 5-hydroxytryptamine than the small artery.
Preliminary experiments have demonstrated that the
magnitude of the response to 5hydroxytryptamine is
significantly increased in the absence of a functioning or
intact endothelium [26]. This is probably related to the
reported actions of 5-hydroxytryptamine on EDRF
release [ 161. Prostaglandin Fzaranother locally produced
mediator, acted as a potent vasoconstrictor on both large
and small pulmonary arteries, although the effect was
significantly greater in the small artery. It is known to
cause persistent vasoconstriction in whole lungs [25].
Bradykinin is normally considered to be a vasodilator
of the pulmonary circulation, but when perfused through
rat lung preparations it acts as a vasoconstrictor [25]. In
small pulmonary arteries bradykinin caused a large but
transient contraction, with a much smaller response in the
large vessel. Similar small transient responses were
observed with angiotensin 11, which is a vasoconstrictor of
the perfused lung preparation [27]. The mechanism of the
tachyphylaxis in isolated arteries is unknown.
Myogenic tone is an intrinsic mechanism of vascular
smooth muscle, and is thought to account for 60% of the
resistance to flow in most vascular beds of the systemic
circulation in vivo, with the exception of the kidney [28].
Small resistance arteries appear to be responsible for a
major proportion of this resistance. Basal tone can be
demonstrated in vitro in isolated rat systemic resistance
arteries from several vascular beds [4]. It is dependent on
extracellular Ca2+,and is related to the degree of applied
stretch and endothelial function [ l , 29, 301. In contrast, it
is generally accepted that the resting tone of the pulmonary circulation is low [4, 51. Only small reductions in
baseline resting tone can be demonstrated in the isolated
perfused lung after application of a variety of vasodilatory
substances [7, 311. We were unable to show any intrinsic
tone in either large or small isolated pulmonary arteries,
despite the vessels being stretched to the point of maximal
contractile performance.
The endothelium may play an important role in the
maintenance of vascular tone in both the systemic and
pulmonary circulations [32, 331, and a large basal release
of EDRF could account for the low resting tone found in
pulmonary arteries. The vasorelaxant action of acetylcholine is thought to involve the release of EDRF [34],yet
we found that in the small pulmonary artery it was ineffective as a vasodilator after pre-contraction with high concentrations of KCl or endothelin-1, although it was
effective in large arteries, and in small arteries constricted
with prostaglandin FZa.Bradykinin also produces EDRFmediated relaxation in most preparations [35], but in the
small pulmonary artery it caused a powerful contraction,
with little or no response in the large artery. These results
imply that EDRF production in the endothelium of small
arteries is insensitive to acetylcholine and bradykinin, or
that production is already maximally stimulated, or alternatively that it does not occur. As acetylcholine does
62
R. M. Leach et al.
induce some relaxation against prostaglandin F2a there
must be some E D W activity, and conversely EDRF production cannot be at its maximum, although it may be
close to it. This does assume, however, that acetylcholine
is not causing relaxation by some other means under these
circumstances. It is also possible that in the small arteries
the technique itself damages the endothelium, but this has
not been found in small systemic arteries and histological
examination of the small pulmonary arteries at the end of
the experiment confirmed that the endothelium was
intact. Although the endothelium may play some role in
maintenance of the low basal resting tone of the pulmonary circulation, its relative importance is unclear.
Further work needs to be performed in order to define the
function of the endothelium in these vessels.
In conclusion, we have demonstrated a number of
important differences between the responses of large ( 1-2
mm diameter) and small (100-300 p m diameter) pulmonary arteries to various pharmacological agents. The
responses of these isolated vessels also differ significantly
from those previously reported for perfused lung preparations. These results suggest that it would be unwise to
extrapolate from results obtained either from large pulmonary artery preparations or from perfused lungs to
what may occur in the small resistance arteries. Clearly,
the comparison of a variety of preparations is required if
the action of mediators on the pulmonary circulation is to
be understood.
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