Vol. 274, Issue 4, H1075-H1081, April 1998
Monkey corpus cavernosum relaxation mediated by NO and other
relaxing factor derived from nerves
Tomio
Okamura,
Kazuhide
Ayajiki, and
Noboru
Toda
Department of Pharmacology, Shiga University of Medical Science,
Seta, Ohtsu 520-2192, Japan
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ABSTRACT |
Isolated monkey
corpus cavernosum muscle strips contracted with prostaglandin
F2
and treated with prazosin
responded to transmural electrical stimulation with frequency-related
relaxations that were abolished by tetrodotoxin. The nitric oxide (NO)
synthase inhibitor
NG-nitro-L-arginine
(L-NNA)
significantly attenuated but did not abolish the response;
L-arginine reversed the
inhibition. The neurogenic relaxation was not influenced in the strips
treated with atropine or calcitonin gene-related peptide
(CGRP)-(8
37), a CGRP-receptor antagonist, and those desensitized to
vasoactive intestinal polypeptide (VIP) or pituitary adenylate
cyclase-activating polypeptide (PACAP). Nerve fibers containing NADPH
diaphorase were histochemically demonstrated in cavernous tissues. The
relaxant response resistant to the NO synthase inhibitor was abolished by high K+ and tetrabutylammonium
but was unaffected by glibenclamide, charybdotoxin, apamin, ouabain,
SKF-525a, a cytochrome P-450
inhibitor, and oxyhemoglobin. It is concluded that neurogenic
relaxations of monkey corpus cavernosum muscle is associated partly
with NO released as a neurotransmitter and that other relaxing
factor(s) possibly responsible for
K+ channel opening also
participates; however, the type of
K+ channel involved is not
determined. Acetylcholine, VIP, CGRP, PACAP, and the
Na+ pump do not seem to be
involved in the neurogenic relaxation.
nitric oxide synthase inhibitor; potassium channel; penile
erection; neurotransmitter
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INTRODUCTION |
THE PENILE ERECTION is evoked by an elevation of
pressure of the corpus cavernosum that is associated with a relaxation
of cavernous smooth muscle. Originally relaxant substances, such as
acetylcholine and vasoactive intestinal polypeptide (VIP), liberated
from nerves were thought to be mediators of the penile erection (17,
25, 36). However, recent studies have provided evidence supporting the
hypothesis that nitric oxide (NO) derived from the nerve mainly
mediates the muscle relaxation in vitro and the increased
intracavernous pressure in vivo in a variety of mammals, including rats
(4, 7, 9), rabbits (27), dogs (12, 33, 34), and humans (26). There are
species variations in the mechanism of relaxation of the corpus
cavernosum. Involvement of vasodilators other than NO, acetylcholine,
and VIP is also reported (10).
Our preliminary study suggested that neurogenic relaxation of monkey
corpus cavernosum muscle was reduced but not abolished by treatment
with NO synthase inhibitors. Therefore, the present study was
undertaken to clarify the involvement of nerve-derived NO in the
relaxation and to analyze the mechanism of neurogenic relaxation
resistant to NO synthesis inhibition in the corpus cavernosum isolated
from Japanese monkeys.
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METHODS |
The studies review board at the Shiga University of Medical Sciences
approved the use of monkey corpus cavernosum in this study.
Preparation. Male Japanese monkeys
(Macaca fuscata), weighing 6-10
kg, were anesthetized with intramuscular injections of ketamine (40 mg/kg) and with intravenous injections of thiopental sodium (20 mg/kg)
and killed by bleeding from the carotid arteries. The penis was rapidly
removed, and corpora cavernosa were isolated. The tunica albuginea was
removed, and two strips (~3 × 5 × 10 mm) from each
individual were obtained. The specimens were vertically fixed between
hooks in a muscle bath (20-ml capacity) containing the nutrient
solution, which was aerated with a mixture of 95% O2-5%
CO2 and maintained at 37 ± 0.3°C. The hook anchoring the upper end of the strips was connected
to the lever of a force-displacement transducer. The resting tension
was adjusted to 0.25 g, which is optimal for inducing the maximal
contraction. Constituents of the solution were as follows (in mM): 120 NaCl, 5.4 KCl, 2.2 CaCl2, 1.0 MgCl2, 25.0 NaHCO3, and 5.6 dextrose. The pH
of the solution was 7.36-7.42. Before the start of experiments,
all of the strips were allowed to equilibrate in the bathing media for 60-90 min, during which time the medium was replaced every
10-15 min.
The strips were placed between stimulating electrodes. The gaps between
the strip and the electrodes were wide enough to allow undisturbed
contraction and relaxation and yet sufficiently narrow to effectively
stimulate intramural nerve terminals. A train of 0.2-ms square pulses
of supramaximal intensity were transmurally applied at frequencies of
2, 5, and 20 Hz for periods of 100, 40, and 10 s, respectively, or in
other series at 0.2, 0.5, 2, and 5 Hz for 40 s. The stimulus pulses
were delivered every 10 min by an electronic stimulator (Nihon-Kohden
Kogyo, Tokyo, Japan).
Functional study. Isometric
contractions and relaxations were displayed on an ink-writing
oscillograph (Nihon-Kohden Kogyo). The contractile response to 30 mM
K+ was obtained, and then the
strips were washed three times with fresh media and equilibrated for
30-40 min. The strips were partially contracted with prostaglandin
(PG) F2 (1-7 µM); the
contractions were in a range between 30 and 45% of the contraction
induced by K+ (30 mM). Transmural
electrical stimulation at 5 Hz was applied repeatedly at intervals of
10 min until steady responses were obtained, and then blocking agents
were applied. In some preparations, in which the stimulation-induced
relaxation was stabilized in the presence of 0.1 mM
NG-nitro-L-arginine
(L-NNA), the bathing fluid was
replaced by the solution in which LiCl was substituted for NaCl, and
L-NNA (0.1 mM) was included; the
muscle tone was then adjusted to about the same level as that before
the change of the solution. At the end of each series of experiment,
papaverine (0.1 mM) was added to attain the maximal relaxation;
relaxations induced by electrical stimulation and vasodilator agents
relative to those induced by papaverine were presented.
Histochemistry. The corpus cavernosum
was rapidly removed and fixed in ice-cold 0.1 M phosphate-buffered
saline (PBS, pH 7.4) containing 0.3% glutaraldehyde and 4%
paraformaldehyde for 10 min, then postfixed overnight in 0.1 M PBS with
4% paraformaldehyde followed by cryoprotection in 15% sucrose. The
fixed blocks were cut into thin sections (20 µm thick) in a cryostat
(
18°C) (Cryotom, Nakagawa Seisakusho, Tokyo, Japan) and
mounted onto gelatin-chrome-alm-coated glass slides. The sections were
then rinsed in 0.1 M PBS. NADPH diaphorase staining was performed by
incubating glass-mounted sections with 0.1 M PBS, pH 8.0, containing 1 mM NADPH (Kohjin, Tokyo, Japan), 2 mM nitroblue tetrazolium (Sigma
Chemical, St. Louis, MO), and 0.3% Triton X-100 at 37°C. The
period of incubation (range 15-30 min) was determined by staining
intensity. The reaction was terminated by washing the sections in 0.1 M
PBS containing 0.3% Triton X-100. Histochemical control experiments,
in which NADPH was excluded from the reaction mixture, gave no positive staining.
Statistics and drugs used. The results
shown in the text, Figs. 1-7, and Table 1 are expressed as means ± SE. Statistical analyses were made using the Student's paired
and unpaired t-test and Tukey's method after one-way analysis of variance. Drugs used were
L-NNA, NG-nitro-D-arginine
(D-NNA), VIP, CGRP-(837),
pituitary adenylate cyclase-activating polypeptide (PACAP)-27,
charybdotoxin (Peptide Institute, Minoh, Japan),
NG-nitro-L-arginine methyl ester
(L-NAME; RBI Research
Biochemicals, Natick, MA),
L- and
D-arginine, tetrabutylammonium
(TBA), methylene blue trihydrate (Nacalai Tesque, Kyoto, Japan),
tetrodotoxin (Sankyo, Tokyo, Japan), atropine sulfate (Tanabe, Osaka,
Japan), prazosin hydrochloride (Pfizer, Tokyo, Japan), apamin (RBI
Research Biochemicals), glibenclamide, 4-aminopyridine (Sigma
Chemical), levcromakalim (SmithKline Beecham, Surrey, UK),
ouabain (Merck, Darmstadt, Germany), PGF2 (Pharmacia-Upjohn, Tokyo,
Japan), and papaverine hydrochloride (Dainippon, Osaka, Japan).
Responses to NO were obtained by adding NaNO2 solution adjusted to pH 2 (8), and concentrations of the solution in the bathing media were
expressed as those of NO. Oxyhemoglobin was prepared by adding a
10-fold molar excess of the reducing agent sodium dithionite to a 1 mM
solution of commercial hemoglobin (Sigma Chemical) in distilled water.
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RESULTS |
Neurogenic response as affected by
L-NNA.
In a preliminary study on strips of the monkey corpus cavernosum
partially contracted with PGF2,
transmural electrical stimulation at 5 Hz produced a relaxation of 50.6 ± 12.5% (n = 5 from separate
monkeys) that was potentiated to 74.2 ± 4.0%
(n = 5, 33.2 ± 11.9% increase,
P < 0.05, paired
t-test) by treatment with 10 µM
prazosin. Therefore, mechanisms of neurogenic relaxation were analyzed
in the strips treated with prazosin. The electrical stimulation
elicited frequency-related relaxations that were abolished by 0.3 µM
tetrodotoxin. Mean values of the relaxation induced at 2, 5, and 20 Hz
for 100, 40, and 10 s, respectively, were 54.0 ± 3.3%
(n = 7), 75.3 ± 3.5%
(n = 7), and 67.4 ± 4.1%
(n = 5), respectively, relative to
those elicited by 0.1 mM papaverine. Reproducible responses were
obtained with 5-Hz stimulation; thus, unless otherwise mentioned, the
analysis of mechanisms underlying the response was carried out in the
strips stimulated at this frequency.
Relaxations induced by 5-Hz stimulation were not influenced by
treatment with 0.1 µM atropine (n = 4) but were attenuated by 10 µM
L-NNA as shown in Fig.
1. The inhibition was reversed by the
addition of L-arginine (1 mM).
Quantitative data with 5-Hz stimulation are summarized in Fig.
2.
D-NNA (10 µM) did not alter the neurogenic relaxation, and
D-arginine (1 mM) failed to
restore the response depressed by
L-NNA. Prolonged electrical
stimulation at 1 Hz elicited a sustained relaxation that was not
affected by atropine. During the nerve stimulation,
L-NNA (10 µM) produced a
contraction, and L-arginine (1 mM) abolished the response (Fig. 3).
Tetrodotoxin abolished the stimulation-induced relaxation. Similar
results were also obtained in two additional strips from separate
monkeys. Exogenously applied NO (10 µM) produced relaxations of 32.0 ± 6.7% (n = 5), which were not
influenced by L-NNA.
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Fig. 1.
Typical tracing of relaxant response to transmural electrical
stimulation (TES; 2, 5, and 20 Hz) of a monkey corpus cavernosum strip
before and after treatment with
NG-nitro-L-arginine
(L-NNA; 10 µM),
L-arginine
(L-Arg; 1 mM), and tetrodotoxin
(TTX, 0.3 µM). The strip was partially contracted with 1 µM
prostaglandin (PG) F2 and
treated with 10 µM prazosin. Papaverine (PA; 0.1 mM) produced the
maximal relaxation.
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Fig. 2.
Modifications by
NG-nitro-D-arginine
(D-NNA; 10 µM),
L-NNA (10 µM),
L-NNA + D-arginine
(+D-Arg; 1 mM),
L-NNA + L-Arg
(+L-Arg; 1 mM), and TTX (0.3 µM) of relaxation induced by TES (5 Hz) of monkey corpus cavernosum
strips contracted with PGF2 and
treated with prazosin (10 µM). Abscissa represents
stimulation-induced relaxations relative to those caused by 0.1 mM PA.
a P < 0.01: significantly different from control (C);
b P < 0.01: significantly different from value with
D-NNA;
c P < 0.01: significantly different from value with
L-NNA + L-Arg;
d P < 0.01: significantly different from
L-NNA;
e P < 0.01: significantly different from value with
L-NNA + D-Arg (Tukey's method). Nos. in
parentheses indicate no. of strips from separate monkeys. Vertical
bars, SEs.
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Fig. 3.
Typical tracing of response to TES (1 Hz) for 80 s at beginning and
~120 min from on to off, as affected by atropine (0.1 µM),
L-NNA (10 µM),
L-Arg (10 mM), and TTX (0.3 µM) in a monkey corpus cavernosum strip contracted with 1.3 µM
PGF2 and 10 µM prazosin. PA
(0.1 mM) produced the maximal relaxation.
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L-NNA in a concentration of 10 µM did not abolish but partially inhibited the response to electrical
nerve stimulation (from 70.7 ± 5.4 to 23.4 ± 2.7%,
n = 9, 63.3 ± 5.2%
inhibition; Fig. 2). Additional inhibition was not obtained by
increasing the concentration to 0.1 mM (from 75.6 ± 3.7 to 26.3 ± 5.7%, n = 7, 65.4 ± 3.5% inhibition). The addition of
L-NAME (0.1 mM) to
L-NNA (0.1 mM)-treated strips
(n = 4) did not further inhibit the
response. The incomplete inhibition by 0.1 mM
L-NNA was also seen in the
responses to nerve stimulation at frequencies <5 Hz for 40 s (Fig.
4). Paired comparisons in the results from
five strips from separate monkeys indicated that the inhibitions by
L-NNA in the responses at 0.2, 0.5, 2, and 5 Hz averaged 77.1 ± 6.3, 80.4 ± 3.0, 52.5 ± 6.2, and 51.8 ± 5.7%, respectively; the values at 0.2 and 0.5 Hz
were significantly different from those at 2 and 5 Hz
(P < 0.05). Despite a marked inhibition of the response to the low frequencies of stimulation, significant relaxations were seen in the presence of
L-NNA. However, tetrodotoxin
applied at the end of each series abolished the stimulation-induced relaxation. Finally, the mechanism underlying the
L-NNA-resistant neurogenic
relaxation was analyzed at 5 Hz under treatment with 0.1 mM
L-NNA.
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Fig. 4.
Modifications by L-NNA (0.1 mM)
and TTX (0.3 µM) of response to TES at low frequency range
(0.2-5 Hz) of monkey corpus cavernosum strips contracted with
PGF2 and treated with prazosin
(10 µM). Abscissa represents stimulation-induced relaxations relative
to those induced by 0.1 mM PA.
a P < 0.01: significantly different from control;
b P < 0.01, c P < 0.05: significantly different from value with TTX (Tukey's
method). n, No. of strips from
separate monkeys. Vertical bars, SEs.
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Neurogenic relaxation resistant to NO synthase
inhibition. The response seen in
L-NNA-treated strips was not
affected by treatment with CGRP-(837) (0.1 µM) (24.0 ± 4.3 vs.
26.3 ± 3.5%, n = 4) or in the
strips desensitized to VIP by repeated applications (10 nM VIP,
4-5 times) of the peptide (29.2 ± 4.6 and 28.3 ± 4.3%, n = 4). The concentration
of CGRP-(837) is sufficient to significantly inhibit the response to
CGRP (35). PACAP (0.1 µM) produced a relaxation averaging 15.2 ± 1.7% (n = 5). Repeated applications (2-3 times) abolished the responsiveness to this peptide of the strips, in which the stimulation-induced relaxation (36.4 ± 5.4%, n = 5) was not different from that
before the addition of PACAP (37.8 ± 6.4%). Treatment with
oxyhemoglobin (16 µM, n = 5) and methylene blue (10 µM, n = 3) did
not inhibit the neurogenic response.
The neurogenic response in the strips treated with 0.1 mM
L-NNA was not significantly
reduced by 1 mM TBA but was abolished at 3 mM (Fig.
5). In addition, applications of 20 mM
K+ abolished the relaxation (Fig.
6), which was restored by replacement of
one-half of the bathing media with normal fluid containing 0.1 mM
L-NNA. Complete recovery of the
response was observed by repeating the same procedure to reduce the
applied K+ concentration to
one-fourth. The results are quantitatively compared in
Fig. 7. Treatment with other compounds that
are recognized to interfere with the transmembrane passage of
K+ through channels did not
significantly inhibit the response to nerve stimulation. The data with
charybdotoxin, apamin, 4-aminopyridine, glibenclamide, and
Ba2+ are shown in Table
1. Levcromakalim, a
K+ channel opener, at 0.1 and 1 µM produced relaxations of cavernous strips of 20.2 ± 4.5 and
76.6 ± 6.6% (n = 5),
respectively.
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Fig. 5.
Modifications by L-NNA (0.1 mM)
and tetrabutylammonium (TBA; 1 and 3 mM) of relaxant response to TES (5 Hz) of monkey corpus cavernosum strips contracted with
PGF2 and treated with prazosin
(10 µM). Abscissa represents stimulation-induced relaxations relative
to those induced by 0.1 mM PA.
a P < 0.05: significantly different from control (C);
b P < 0.01: significantly different from value with
L-NNA;
c P < 0.05: significantly different from value with 1 mM TBA (Tukey's
method). Nos. in parentheses indicate no. of strips from separate
monkeys. Vertical bars, SEs.
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Fig. 6.
Typical tracing of relaxation induced by TES (5 Hz) of a monkey corpus
cavernosum strip before and after
L-NNA (0.1 mM), TBA (1 mM),
excess K+, depletion of
K+ with control media containing
0.1 mM L-NNA (same procedure was
done twice), and tetrodotoxin (TTX, 0.3 µM). The strip was partially
contracted with PGF2 (1.2 µM)
and prazosin (10 µM); a at top right
and bottom left are same response. PA (0.1 mM) produced maximal
relaxation.
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Fig. 7.
Modifications by L-NNA (0.1 mM),
excess K+ (K, 20 mM), and
K+ depletion [by
L-NNA-containing solution;
depleted to one-half (K 1/2) and one-fourth
K+ strength (K 1/4)] of
response to TES (5 Hz) of monkey corpus cavernosum strips contracted
with PGF2. Abscissa denotes
stimulation-induced relaxations relative to those caused by 0.1 mM PA.
a P < 0.01: significantly different from control (C);
b P < 0.01: significantly different from value with
L-NNA;
c P < 0.01: significantly different from value with K;
d P < 0.05: significantly different from value with K 1/2 (Tukey's
method). Nos. in parentheses indicate no. of strips from separate
monkeys. Vertical bars, SEs.
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Table 1.
Effects of charybdotoxin, apamin, 4-aminopyridine, glibenclamide,
Ba2+, and SKF-525a on relaxant response to transmural
electrical stimulation at 5 Hz
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Under treatment with 0.1 mM
L-NNA, relaxations induced by
electrical stimulation were not attenuated in the cavernous strips exposed for 30 min to 1 µM ouabain (38.9 ± 5.4 vs. 41.6 ± 5.9%, n = 7). Substitution of
isotonic LiCl for NaCl did not significantly alter the
stimulation-induced response during the observation period of 60 min;
mean values were 31.8 ± 5.5%
(n = 6) in control media and
36.2 ± 5.2% 30 min after exposure to the experimental solution.
According to Hecker et al. (13), the response to endothelium-derived
relaxing factor resistant to NO synthase inhibitors is significantly
attenuated by a cytochrome P-450
inhibitor, SKF-525a, suggesting the involvement of epoxide. Treatment
with this inhibitor (0.1 mM) did not reduce the response to nerve
stimulation (Table 1).
Histochemical study. In the trabecular
meshwork of a corpus cavernosum strip, abundant nerve fibers and
bundles containing NADPH diaphorase were demonstrated histochemically
(Fig. 8). Similar results were also
obtained in the strips from two additional monkeys.
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Fig. 8.
NADPH diaphorase histochemistry of a monkey corpus cavernosum strip.
There are abundant positively stained nerve fibers in the trabecular
meshworks. Bar, 50 µm.
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DISCUSSION |
The present study revealed that corpus cavernosum muscle strips
obtained from the Japanese monkey responded to electrical nerve
stimulation with a frequency-related relaxation that was significantly
inhibited by treatment with high concentrations (10 and 100 µM) of an
NO synthase inhibitor, L-NNA,
but not by D-NNA. The inhibition
was reversed by L- but not
D-arginine. Exogenously applied
NO relaxed the cavernous muscle. NO synthesized from
L-arginine in nerve terminals
appears to partially mediate the relaxation of monkey corpus cavernosum
participating in the penile erection. Histochemical demonstration of
nerve fibers containing NADPH diaphorase in monkey cavernous trabecular
meshworks (present study) and NO synthase in dog corpora cavernosa (12)
supports the hypothesis of nitroxidergic innervation (31). Atropine did
not alter the neurogenic response, suggesting that the release of
endothelium-derived NO by a mediation of muscarinic receptor
stimulation by acetylcholine (2) derived from nerves is not involved in
the response. VIP, the other candidate of neurotransmitter (17), and
CGRP, a vasodilator neurotransmitter (1, 18), are unlikely to mediate
the relaxation because the strips made tachyphylactic to VIP and those
treated with a sufficient concentration of the CGRP receptor antagonist CGRP-(837) (35) did not inhibit the response to nerve stimulation. PACAP is demonstrated to be colocalized with VIP in nerve structures in
human cavernous tissues and to be effective in relaxing isolated cavernous strips (14), as seen in the monkey strips in the present study. However, the relaxation in the monkey tissue was not so evident,
and the response to nerve stimulation was not reduced in the strips
made unresponsive to the peptide, suggesting that PACAP used here is
not involved in the neurogenic response. Therefore, it is unlikely that
VIP, CGRP, and PACAP are neurotransmitters responsible for cavernous
muscle relaxation.
Continuous nerve stimulation at a low frequency (1 Hz, Fig. 3) produced
a sustained, intense relaxation that might reflect a prolonged penile
erection under effective and nondeteriorating neural influences in
vivo. The relaxation was interrupted by an inhibition of NO synthesis
and a blockade of nerve action potentials.
The relaxation induced by transmural electrical stimulation was
abolished by tetrodotoxin but inhibited only partially by treatment
with L-NNA. Raising the
concentration of the inhibitor from 10 to 100 µM did not produce
additional inhibition. Cohen et al. (5) have demonstrated that the
endothelium-dependent relaxation and hyperpolarization and the NO
release induced by acetylcholine in the rabbit carotid artery are not
abolished even by combined treatment with high concentrations of
L-NNA and
L-NAME, suggesting the NO
release is resistant to NO synthase inhibitors. However, this does not
seem to be the case in our preparation, because the response in
L-NNA (100 µM)-treated strips
was not influenced by oxyhemoglobin, a NO scavenger, or methylene blue in concentrations sufficient to abolish the response to neurogenic and
endothelial NO in monkey (30, 32) and canine blood vessels (29).
Therefore, the inhibitory nerve is postulated to liberate relaxant
substance(s) other than NO. In isolated rabbit corpus cavernosum
strips, the electrical stimulation-induced relaxation at high
frequencies is not abolished by
L-NNA (30 µM), oxyhemoglobin (10 µM), and methylene blue (10 µM) (16). In isolated canine corpus
cavernosum, the neurogenic relaxation is totally abolished by
L-NNA, suggesting that the
response is mediated exclusively by NO produced from
L-arginine (12). Possibility of
discriminating the release of different neurotransmitters from nerves
by changing stimulation frequencies has been described (20). This may
be the case in the material used here because the responses to low frequencies of stimulation (0.2 and 0.5 Hz) were more susceptible to
L-NNA than those to high
frequencies (2 and 5 Hz) (77-80 vs. 52-53% inhibition
obtained from the same 5 strips, Fig. 4), although the relaxation
resistant to the NO synthase inhibitor was still observed even when the
nerve was stimulated at low frequencies.
The stimulation-induced relaxation resistant to a high concentration of
L-NNA was abolished by raising
the external concentration of K+,
which did not abolish the relaxation induced by NO or
PGI2 (data not shown). Lowering
the external K+ concentration
reversed the response. According to Saito et al. (27), relaxations
induced by electrical field stimulation of rabbit corpus cavernosum
strips are potentiated by increasing the external
K+ concentrations. High
concentrations of TBA also abolished the neurogenic response.
Therefore, substance(s) opening K+
channels in smooth muscle cell membranes may be involved in the response of the monkey strips. Levcromakalim, an ATP-activated K+ channel opener, relaxed the
strips. However, glibenclamide, an ATP-activated
K+ channel inhibitor, and
charybdotoxin and apamin,
Ca2+-activated
K+ channel inhibitors, did not
significantly alter the response resistant to
L-NNA.
Ba2+ and 4-aminopyridine in
concentrations used were also ineffective. Because SKF-525a, a
cytochrome P-450 inhibitor, depressed
the endothelium-derived hyperpolarizing factor (EDHF)-mediated
relaxation in canine, bovine, and porcine coronary arteries that was
resistant to NO synthase inhibitors (13, 23), the authors suggested the
involvement of arachidonic acid metabolite synthesized by cytochrome
P-450, presumably epoxide. However,
this inhibitor in the same concentration failed to inhibit the response
to nerve stimulation. There are numerous papers demonstrating that the hyperpolarization and relaxation via EDHF are ascribed by opening of
certain types of K+ channels in
subprimate mammals (3, 15, 19, 21, 22, 37), on the basis of findings
with so-called selective K+
channel inhibitors. However, in the present study, the type of K+ channels responsible for non-NO
relaxing factor(s) could not be specified.
A possible role of
Na+-K+-adenosinetriphosphatase
in relaxations of human corpus cavernosum muscle is suggested (11).
Relaxations and membrane hyperpolarization in vascular smooth muscle
are reportedly mediated by the
Na+-K+
pump (6, 24), the activity being reduced by ouabain and replacement of
extracellular Na+ with
Li+. In our preparations treated
with high concentrations of
L-NNA, the relaxant response to
nerve stimulation was not influenced by ouabain or with substitution of
LiCl for NaCl. Involvement of a neuronal substance that activates this
enzyme would therefore be excluded. The present study revealed for the
first time a neurogenic relaxation that was mediated not only by NO but
also by other substances in monkey corpus cavernosum smooth muscle,
although a non-NO mechanism of neurogenic relaxation has been reported in nonvascular smooth muscle (28). Relaxations by this substance may be
due to opening of K+ channels, the
types of which were unidentified. Whether this type of channel is
present specifically in primate corpus cavernosum or is seen also in
other vascular smooth muscle from a variety of mammals remains to be
ascertained. Involvement of acetylcholine, VIP, CGRP, PACAP, and the
Na+ pump in the response is
excluded.
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ACKNOWLEDGEMENTS |
The authors thank Dr. K. Yoshida for technical assistance with the
histochemical study.
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FOOTNOTES |
This work was supported in part by the Grant-in-Aid for Scientific
Research (B) from the Ministry of Education, Science, Culture, and
Sports, Japan.
Address for reprint requests: N. Toda, Dept. of Pharmacology, Shiga
Univ. of Med. Sci., Seta, Ohtsu 520-2192, Japan.
Received 12 September 1997; accepted in final form 17 November
1997.
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Abstract
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