Effects of central and peripheral urocortin on fed and fasted
gastroduodenal motor activity in conscious rats
Naoki
Kihara1,
Masaki
Fujimura1,
Ikuo
Yamamoto1,
Etsuro
Itoh3,
Akio
Inui4, and
Mineko
Fujimiya2
Departments of 1 Surgery and 2 Anatomy, Shiga
University of Medical Science, Otsu, Shiga 520-2192;
3 Pharmaceutical Research Department, Ube Research Laboratory,
Ube, Yamaguchi 755-8633; and 4 Second Department of Internal
Medicine, Kobe University School of Medicine, Kobe 650-0017, Japan
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ABSTRACT |
Since few
previous studies have examined the effects of urocortin on
physiological fed and fasted gastrointestinal motility, we administered
urocortin intracerebroventricularly (icv) or intravenously (iv) in
freely moving conscious rats and examined the changes in antral and
duodenal motility. Icv and iv injection of urocortin disrupted fasted
motor patterns of gastroduodenal motility, which were replaced by
fed-like motor patterns. When urocortin was given icv and iv in the fed
state, the motor activity remained like the fed patterns but % motor
index (%MI) was decreased in the antrum and increased in the duodenum.
Increase in the %MI in the duodenum induced by urocortin was shown as
a nonpropagated event, since the transit of nonnutrient contents in the
duodenum was decreased by icv and iv injection of urocortin. Changes in
the gastroduodenal motility induced by icv injection of urocortin were
abolished in animals with truncal vagotomy but not altered in animals
with mechanical sympathectomy, suggesting that the vagal pathway may
mediate the central action of urocortin. Neither urocortin antiserum
nor
-helical CRF-(9-41) affected fed and fasted
gastroduodenal motility, suggesting that endogenous urocortin is not
involved in regulation of basal gastroduodenal motility.
intracerebroventricular injection; intravenous injection; stomach; duodenum
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INTRODUCTION |
UROCORTIN, A
NEWLY CHARACTERIZED mammalian neuropeptide and a member of the
corticotropin-releasing factor (CRF) family, acts as an endogenous
ligand for the CRF receptor (58). This peptide is
implicated in various functions, such as feeding suppression (50,
51), anxiety (40), and cardiovascular
(44) and gastrointestinal (41) regulation.
Such a variety of actions may be accounted for by a wide distribution
of CRF receptor in the brain (10, 31, 32, 33, 46, 48) and
peripheral organs (26, 31). The effects of centrally
administered CRF on gastrointestinal functions have been widely
investigated in rats (7, 11, 29, 35, 50, 53) and mice
(49). Intracerebroventricular (icv) or intracisternal
injection of CRF caused the inhibition of gastric emptying of
nonnutrient liquid meals (7, 29, 49, 53) or nutrient solid
meals (11, 28, 35, 50), yet conversely caused the
stimulation of colonic transit (17, 30, 37, 59). These
effects of centrally administered CRF on gastric and colonic motility
were similar to abdominal surgery- (4, 5) or stress- (8, 17, 30, 37, 39, 54, 59) induced gastric ileus or
colonic dysfunction. Since central administration of CRF antagonists blocked the postoperative or stress-induced alterations of
gastrointestinal dysfunction, motor responses in the gut produced by
stress are thought to be mediated by the endogenous CRF in the brain
(17, 30, 37, 59). Peripheral administration of CRF or
urocortin also inhibited gastric emptying in rats (41),
mice (2, 49), and dogs (43). Recent studies
indicate that urocortin binds both CRF type 1 and 2 receptors but shows
a higher affinity for CRF type 2 receptor than for CRF type 1 receptor
(6, 58). To examine the receptor subtypes involved in the
regulation of gastrointestinal motility induced by centrally or
peripherally administered CRF and urocortin, the functional potency of
selective or nonselective CRF type 1 and 2 receptor antagonists in
blocking the effects of agonists has been examined (35,
41).
-Helical CRF (9-41) is known as a dual
antagonist to CRF type 1 and 2 receptors (14) but is more
selective for CRF type 2 receptor than for CRF type 1 receptor on the
basis of functional and binding studies (26, 41). In fact,
CRF type 2 receptor in both brain and peripheral organs is mainly
involved in the regulation of gastrointestinal motility (35,
41).
Although a number of previous studies have shown the effects of central
or peripheral administration of CRF/urocortin on gastrointestinal motility, a mere transit of meals from the stomach has been used as a
marker of gastric motility. It is well known that gastrointestinal motility under physiological conditions consists of fasted and fed
motor patterns and that each of them is regulated by different mechanisms (23). In fact, previous studies have shown that
centrally administered neuropeptides regulate the fed and fasted
patterns of motility. Neuropeptide Y (NPY), a feeding-stimulatory
peptide in the brain, induced the fasted pattern in the duodenum when given to the fed rats (15), whereas bombesin, a
feeding-inhibitory peptide, induced the fed pattern in the small
intestine when given to fasted dogs (20). Therefore, it
seems necessary to investigate the action of CRF/urocortin, which is
known as a feeding-inhibitory peptide in the brain, on the
physiological fed and fasted motor activities of the gastrointestinal tract.
In the present study, urocortin was used as a ligand for CRF receptors
because urocortin is known to be two- to threefold more potent than CRF
in inhibiting food intake and gastric emptying (2, 41).
Urocortin was given either centrally or peripherally, and the effects
on the fed and fasted patterns of gastroduodenal motility were
examined. Changes in the motor activity in the fed state of the
duodenum induced by urocortin were assessed compared with the transit
of nonnutrient duodenal contents.
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MATERIALS AND METHODS |
Male Wistar rats (Clea Japan, Tokyo, Japan), weighing
200-250 g at the start of the experiments, were used. Rats were
housed under controlled temperature (21-24°C) and light (lights
on 8:00-20:00) conditions with free access to laboratory chow
pellets (CE-2; Clea Japan) and water. Care of animals was conducted in
accordance with the Guide for Use of Experimental Animals (Shiga
University of Medical Science).
Animal preparation.
The rats were anesthetized with intraperitoneal injection of
pentobarbital sodium (50 mg/kg Nembutal; Abbott Laboratories), placed
in a stereotaxic apparatus, and implanted with guide cannula (25-gauge;
Eicom, Kyoto, Japan), which reached the right lateral ventricle. Stereotaxic coordinates were 0.8 mm posterior to bregma, 2.0 mm right lateral to the midline, and 4.5 mm below the outer surface of
the skull using a Kopf stereotaxic frame with the incisor bar set at
the horizontal plane passing through bregma and lambda. The guide
cannula was secured with dental cement anchored by two stainless steel
screws fixed on the dorsal surface of the skull. After surgery, a dummy
cannula (Eicom) was inserted into each guide cannula, and a screw cap
(Eicom) was put on the guide cannula to prevent blockade. The animals
were allowed to recover for at least 4 days after this operation. On
the day of the experiment, a dummy cannula was replaced by a
microinjection cannula (AMI-5; Eicom) connected to a polyethylene tube
(PE-50, Clay Adams). The placement of the cannula was verified at the
end of the experiment by injection of 10 µl dye (0.05% cresyl
violet) and examination of brain slices.
At 4-6 days after the brain operation, rats were deprived of food
and given free access to water for 18 h before the abdominal operation. They were anesthetized with pentobarbital sodium (50 mg/kg),
and the motility recording device was implanted as follows. Two
manometric catheters (3-Fr, 1 mm diameter; ATOM, Tokyo, Japan) with
side holes were inserted through the gastric fistula, and tips were
placed at the gastric antrum and 3 cm distal to the pylorus. Catheters
were fixed at the gastric wall by purse-string suture and run
subcutaneously to emerge at the crown of the neck and were secured at
the animal's skin. In some animals, a catheter (3-Fr, 1 mm diameter)
was placed in the right jugular vein instead of icv cannulation, run
subcutaneously to emerge at the crown with the manometric catheter, and
used for intravenous (iv) administration of peptides. The catheter was
filled with heparinized saline to prevent obstruction. During the first
postoperative day, the animals were allowed water but no food, and the
experiment was performed 1 wk after the operation. Animals were fasted
18 h before the experiment.
Icv and iv injection of peptide.
Urocortin (Peptide Institute, Osaka, Japan) was dissolved with 1%
acetate solution and frozen until use. Urocortin doses of 0.01 µg,
0.1 µg, 1 µg, and 5 µg were dissolved in saline in a 10-µl
volume for icv injection or a 0.5-ml volume for iv injection. Each dose
of urocortin was administered icv through the microinjection cannula
anchored in the skull or iv through the jugular vein cannula. Vehicle
control was made by icv or iv injection of saline.
Measurement of gastroduodenal motility.
Gastroduodenal motility was measured in the conscious, freely moving
rats by the manometric method. On the day of the experiment, the
manometric catheter was connected to a pressure transducer (TP-400T;
Nihon Koden Kogyo, Tokyo, Japan), and the catheter was protected from
biting by a flexible metal sheath and connected to the infusion swivel
(dual type, 20-gauge; Instech Laboratories, Plymouth Meeting, PA) to
allow free movement. The catheter was continuously infused with
bubble-free 0.9% saline at a rate of 2 ml/h by a low-compliance
capillary infusion system using a heavy-duty pump (CVF-3100; Nihon
Koden). The data were recorded on a polygraph (RM-6100; Nihon Koden)
and stored in a MacLab system (MacLab/8e, AD Instruments Ply, Power
Book 2400c/240; Apple Computer).
After the sequence of typical fasted pattern of motility was observed,
rats were given two pellets of laboratory chow (~8 g). After the
gastroduodenal motility was ascertained to have changed into the fed
pattern, icv injection of each dose of peptide was performed. In some
experiments, icv injection was performed in animals that showed the
fasted pattern of motility. Effects of iv injection of peptides through
the jugular vein cannula were also examined. In other experiments, 50 µg of
-helical CRF-(9-41) (Sigma, St. Louis, MO)
were injected icv or iv before icv injection of 5 µg urocortin or
injected iv or icv before iv injection of 5 µg urocortin.
The transit of nonnutrient duodenal contents induced by icv and iv
injection of urocortin was examined. One-half milliliter of optimum
cutting temperature compound (Tissue-TEK; Miles, Elkhart, IN),
containing 5% toluidine blue (13), was injected through the manometric catheter inserted into the duodenum 10 min after icv or
iv injection of urocortin (5 µg). The rats were killed after 20 min,
and duodenal transit was measured as the distance (cm) of the most
distal point of the stain from the pylorus.
Immunoneutralization of intrinsic urocortin.
After the sequence of fasted or fed pattern was observed, 5 µl
anti-urocortin antiserum (Yanaihara Institute, Shizuoka, Japan) or
normal rabbit serum plus 5 µl saline were injected icv or iv, and the
change in gastroduodenal motility was observed. In other experiments, 5 µl of anti-urocortin antiserum were injected icv or iv before icv or
iv injection of urocortin in both the fasted and fed states.
Truncal vagotomy and mechanical sympathectomy.
Four days before the measurement of gastroduodenal motility, either
truncal vagotomy or mechanical sympathectomy was performed. Under
median laparotomy on anesthetized rats, the lower part of the esophagus
was exposed and the anterior and posterior branches of the vagal nerve
were incised. For the mechanical sympathectomy, the roots of celiac and
superior mesenteric arteries were exposed and prevertebral ganglia
between these arteries were completely removed. Sham-operated controls
for vagotomy and sympathectomy were made by laparotomy on anesthetized
rats. Vagotomized rats were fed by liquid diet after the operation to
avoid excess distention of the stomach.
Analysis of motor activity and statistical analysis.
To examine the motor activity, 10 rats were used for the measurement of
normal fed and fasted patterns and 3 rats were used for each
experiment. The frequency of the fasted pattern was obtained from the
average of the onset of phase III-like activities per hour, and that of
the fed pattern was obtained from the average of the onset of spike
waves per minute (15). The drug effects on the fed motor
activity were evaluated by changes in the %motor index (MI) before and
after drug administration. MI was defined as the summation of the
amplitude of contractions per minute, and %MI was calculated as
100 × (mean MI for 30 min after drug infusion)/(mean MI for 30 min before drug infusion). Results were expressed as means ± SD;
however, in graphs (Figs. 3B, 5B, 7C, and 8C) results were expressed as means ± SE.
Statistical analysis of the data in Figs. 3B, 5B,
7C, and 8C was performed by one-way ANOVA
followed by Fisher's protected least significant differences test.
Values of P < 0.05 were considered statistically significant.
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RESULTS |
Gastroduodenal fed and fasted motor activity in conscious rats.
In the fasted state, the cyclic changes of pressure waves were detected
in both the antrum and duodenum by a manometric method, including the
quiescence period, during which relatively low amplitude of
contractions occurred (phase I-like contraction), followed by a
grouping of strong contractions (phase III-like contraction) (Fig.
1). The frequency of the onset of the
phase III-like activity in the antrum was 5.17 ± 0.51/h
(n = 6), and that in the duodenum was 5.63 ± 1.30/h (n = 6); those were in accordance with the
migrating myoelectrical complex (MMC) (53). After food
intake, the fasted motor pattern was disrupted and replaced by the fed
motor pattern, which consisted of irregular contractions of high
frequency: 3.8 ± 0.6/min (n = 10) in the stomach
and 1.8 ± 0.6 /min (n = 10) in the duodenum. Such
fed motor patterns continued for 109.0 ± 7.8 min
(n = 5) in the stomach and 119.0 ± 30.9 min
(n = 5) in the duodenum, followed by the onset of the
phase III-like activities (Fig. 1).

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Fig. 1.
Simultaneous recording of antral and duodenal motility in
a conscious, freely moving rat. In both the antrum and duodenum, the
fasted pattern, which consists of the cyclic change of phase I- and
phase III-like contractions, is changed by feeding into the fed
pattern, which consists of irregular contractions with high
frequency.
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Effects of icv and iv injection of urocortin on fed and fasted
motor activities.
When icv injection of urocortin (1 and 5 µg) was given in the fasted
state, the fasted pattern of motility, consisting of consecutive phase
I and phase III-like contractions found in the antrum and duodenum,
changed into the fed-like motor pattern of irregular contractions with
high frequency (Fig. 2). The fed-like motor pattern induced by icv injection of urocortin (5 µg) continued for 171.0 ± 26.0 min (n = 3) both in the stomach
and duodenum. When icv injection of urocortin (1 and 5 µg) was given
in the fed state, the motility pattern remained a fed pattern in both the antrum and duodenum (Fig.
3A); however, the %MI of the
pressure waves was significantly reduced in the stomach and was
significantly increased in the duodenum (Fig. 3B). Changes
in the %MI of the pressure waves in the stomach and duodenum induced
by different doses of icv urocortin are shown in Fig. 3B.

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Fig. 2.
Effect of intracerebroventricular (icv) injection of urocortin (5 µg/rat) on fasted motor activity of the antrum and duodenum. The
fasted pattern is changed into the fed-like motor pattern immediately
after icv injection of urocortin in both the antrum and duodenum.
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Fig. 3.
A: effect of icv injection of urocortin (5 µg/rat) on fed motor activity of the antrum and duodenum. The
motility pattern remains a fed pattern; however, the amplitude of the
pressure wave is decreased in the antrum but increased in the duodenum
after icv injection of urocortin. B: comparison of %motor
index (%MI) in the antrum and duodenum after icv injection of saline
and various concentrations of urocortin in the fed state. Values are
means ± SE from 3 animals. *P < 0.05 and
**P < 0.01.
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The effects of iv injection of urocortin on antral and duodenal
motility were also examined. With the iv injection of urocortin (1 and
5 µg) given in the fasted state, the fasted motor pattern observed in
the antrum and duodenum was replaced by the fed motor pattern, which
continued for 146.0 ± 18.0 min (n = 3) in both the stomach and duodenum (Fig. 4). When
iv injection of urocortin (5 µg) was given in the fed state, the fed
motor patterns remained but the %MI was significantly reduced in the
stomach and significantly increased in the duodenum (Fig.
5). Intravenous injection of
0.1 and 0.01 µg of urocortin had no effect on the gastroduodenal
motility.

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Fig. 4.
Effect of intravenous (iv) injection of urocortin (5 µg/rat) on fasted motor activity of the antrum and duodenum. The
fasted pattern is changed into the fed-like motor pattern after iv
injection of urocortin in both the antrum and duodenum.
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Fig. 5.
A: effect of iv injection of urocortin (5 µg/rat) on fed motor activity of the antrum and duodenum. The
motility pattern remains a fed pattern; however, the amplitude of the
pressure wave is decreased in the antrum but increased in the duodenum
after iv injection of urocortin. B: comparison of %MI in
the antrum and duodenum after iv injection of saline and different
concentrations of urocortin in the fed state of animals. Values are
means ± SE from 3 animals. **P < 0.01.
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The transit of nonnutrient duodenal contents was measured by the
toluidine blue migration induced by icv and iv injection of urocortin.
Within 20 min after icv and iv injection of 5 µg urocortin, the
distances migrated were 29.2 ± 3.0 cm (n = 3) and 30.2 ± 2.8 cm (n = 3), respectively. These values
were significantly lower (P < 0.05) compared with icv
(40.0 ± 3.4 cm, n = 3) and iv (41.2 ± 4.0, n = 3) injections of saline.
Effects of CRF receptor antagonist and urocortin antibody on
urocortin-induced change in the gastroduodenal motility.
To examine whether the effects of icv and iv injection of urocortin on
the motor activity were mediated by CRF receptors in the brain and
periphery, respectively, CRF receptor antagonist
-helical
CRF-(9-41) was injected before urocortin. Changes in the pattern of gastroduodenal motility induced by icv and iv injection of urocortin during fasted states seen in Fig. 2 and Fig. 4 were antagonized by
-helical CRF-(9-41) given by the
same route (Fig. 6). Similarly, changes
in the pattern of gastroduodenal motility induced by icv and iv
injection of urocortin during fed states seen in Fig. 3A and
Fig. 5A were antagonized by
-helical
CRF-(9-41) given by the same route (Fig.
7, A and C, and
Fig. 8, A and C).

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Fig. 6.
A: effect of icv injection of -helical ( h-) corticotropin-releasing factor (CRF)-(9-41) (50 µg/rat) before the icv injection of urocortin (5 µg/rat) on the
fasted motor activity of the antrum and duodenum. B: effect
of iv injection of -helical CRF-(9-41) (50 µg/rat) before the iv injection of urocortin (5 µg/rat) on the
fasted motor activity of the antrum and duodenum. Changes in the motor
pattern induced by icv and iv injection of urocortin seen in Figs. 2
and 4 were completely antagonized by -helical
CRF-(9-41).
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Fig. 7.
A: effect of icv injection of -helical
CRF-(9-41) (50 µg/rat) before the icv injection of
urocortin (5 µg/rat) on the fed motor activity of the antrum and
duodenum. B: effect of iv injection of -helical
CRF-(9-41) (50 µg/rat) before the icv injection of
urocortin (5 µg/rat) on the fed motor activity of the antrum and
duodenum. Pretreatment of icv injection of -helical
CRF-(9-41) completely antagonizes the change in the
motor activity induced by icv injection of urocortin seen in Fig.
3A, whereas pretreatment with iv injection of -helical
CRF-(9-41) does not alter the change in the motor
activity induced by icv injection of urocortin. C:
comparison of %MI in the antrum and duodenum induced by injection of
saline, urocortin (icv), urocortin (icv) + -helical
CRF-(9-41) (icv), and urocortin (icv) + -helical CRF-(9-41) (iv). Values are means ± SE from 3 animals. **P < 0.01. No significant change
was found between %MI induced by saline and urocortin (icv) + -helical CRF-(9-41) (icv) or between %MI induced
by urocortin (icv) and urocortin (icv) + -helical
CRF-(9-41) (iv) in both antrum and duodenum.
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Fig. 8.
A: effect of iv injection of -helical
CRF-(9-41) (50 µg/rat) before the iv (5 µg/rat)
injection of urocortin on the fed motor activity in the antrum and
duodenum. B: effect of icv injection of -helical
CRF-(9-41) (50 µg/rat) before the iv (5 µg/rat)
injection of urocortin on the fed motor activity of the antrum and
duodenum. Pretreatment with iv injection of -helical
CRF-(9-41) completely antagonizes the change in the
motor activity induced by iv injection of urocortin seen in Fig.
5A, whereas pretreatment with icv injection of -helical
CRF-(9-41) does not alter the change in the motor
activity induced by iv injection of urocortin. C: comparison
of %MI in the antrum and duodenum induced by injection of saline,
urocortin (iv), urocortin (iv) + -helical
CRF-(9-41) (iv), and urocortin (iv) + -helical CRF-(9-41) (icv). Values are means ± SE from 3 animals. **P < 0.01. No significant
change was found between %MI induced by saline and urocortin (iv) + -helical CRF-(9-41) (iv) or between %MI induced
by urocortin (iv) and urocortin (iv) + -helical
CRF-(9-41) (icv) in both antrum and duodenum.
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To rule out the possible leakage of icv-injected peptides into the
systemic circulation, CRF receptor antagonist
-helical CRF-(9-41) was injected iv before icv injection of
urocortin. Changes in the pattern of gastroduodenal motility induced by
icv injection of urocortin during fed states seen in Fig. 3A
were not altered by the systemic administration of
-helical
CRF-(9-41) (Fig. 7, B and C).
To rule out the possible influx of systemically injected peptide into
the brain,
-helical CRF-(9-41) was injected icv
before iv injection of urocortin. Changes in the gastroduodenal motility induced by iv injection of urocortin during the fed state seen
in Fig. 5A were not altered by the icv injection of
-helical CRF-(9-41) (Fig. 8, B and
C). A single injection of
-helical CRF-(9-41) icv or iv did not change the fed and
fasted patterns of gastroduodenal motility (Figs. 6, A and
B; 7, A and B; and 8, A and
B).
The effects of passive immunoneutralization with urocortin antiserum on
gastroduodenal motility during fasted and fed states were examined.
Neither icv- nor iv-administered urocortin antiserum (5 µl) changed
the fasted (Fig. 9B) and fed
(Fig. 9D) patterns of gastroduodenal
motility. The same volume of urocortin antiserum blocked the changes in
the gastroduodenal motility induced by icv or iv injection of urocortin
during fasted (Fig. 9A) and fed (Fig. 9C) states.


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Fig. 9.
A and C: effects of icv and iv
injection of urocortin antibody (5 µl/rat) before the icv and iv
injection of urocortin (5 µg/rat) on the fasted (A) and
fed (C) motor activity in the antrum and duodenum.
Pretreatment with urocortin antibody blocked the changes in the
gastroduodenal motility induced by icv or iv injection of urocortin
during fasted and fed states. B and D: effects of
icv and iv injection of urocortin antibody (5 µl/rat) on the fasted
(B) and fed (D) motor activity. Neither icv nor
iv injection of urocortin antibody changed the fasted and fed patterns
of gastroduodenal motility.
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Involvement of the autonomic nervous system.
We examined the involvement of the autonomic nervous system using rats
that had received truncal vagotomy or sympathectomy. Normal fed and
fasted patterns of gastroduodenal motility were not affected by the
sympathectomy. The normal fed pattern was disrupted in both antrum and
duodenum in animals with truncal vagotomy, and the normal fasted
pattern was also somewhat disrupted in the duodenum but was intact in
the antrum in vagotomized animals (Fig.
10). When urocortin was injected icv in
vagotomized animals, changes in the fasted pattern seen in Fig. 2 were
completely abolished, especially in the antrum (Fig. 10A).
However, changes in the fasted pattern induced by iv injection of
urocortin seen in Fig. 4 were not basically altered by the vagotomy
(Fig. 10B). In contrast, sympathectomy failed to alter the
changes in fed and fasted motor activity induced by icv and iv
injection of urocortin (data not shown).

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Fig. 10.
Effects of icv (5 µg/rat; A) and iv (5 µg/rat; B) injection of urocortin on the fasted state of
animals that had received truncal vagotomy. Fasted pattern of motility
is not clearly seen in the duodenum. Vagotomy abolishes changes in the
fasted pattern induced by icv injection of urocortin seen in Fig. 2
(A). However, it does not alter changes in the fasted
pattern induced by iv injection of urocortin seen in Fig. 4
(B).
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DISCUSSION |
In the present study, we investigated the effects of urocortin
administered icv or iv on the fed and fasted motor activity of the
antrum and duodenum in freely moving conscious rats. When urocortin was
administered icv in the fasted state, the fasted patterns of antral and
duodenal motility were disrupted and the fed-like motor patterns
appeared. Similar results were obtained when urocortin was injected iv
in fasted rats, in which the fasted patterns of antral and duodenal
motility were replaced by the fed motor patterns. On the other hand,
when urocortin was administered icv in the fed state, the motor
activities in the antrum and duodenum were affected differently.
Although the motor activities remained as fed patterns in both antrum
and duodenum, %MI was decreased in the antrum but increased in the
duodenum. Similar results were obtained when urocortin was administered
iv in the fed state; the motor activity remained like fed patterns and
%MI was decreased in the antrum but increased in the duodenum. The
effects of icv or iv injection of urocortin on the motor activity of
the gastrointestinal tract were mediated by CRF receptors in the brain
or peripheral organs, since CRF receptor antagonist
-helical
CRF-(9-41) injected via the same route as urocortin
blocked these effects. To rule out the possible leakage of icv-injected
peptide into the systemic circulation, iv injection of
-helical
CRF-(9-41) was combined with icv injection of
urocortin. Conversely, to rule out the possible influx of systemically
injected peptide into the brain, icv injection of
-helical
CRF-(9-41) was combined with the iv injection of urocortin. Results showed that iv injection of
-helical
CRF-(9-41) did not alter the effects of icv-injected
urocortin, and conversely icv injection of
-helical
CRF-(9-41) did not alter the effects of iv-injected
urocortin. These data suggest that central and peripheral urocortin may
exert actions independently through CRF receptors in the brain and
peripheral organs. The possible brain nuclei responsive to icv
injection of urocortin have been widely examined by the dual labeling
of c-fos induction by icv urocortin and mRNA expression of
CRF type 1 and 2 receptors (6). The c-fos
induction site after icv injection of urocortin in the brain was widely
distributed but not always parallel to the site expressing CRF type 2 receptor mRNA; however, it more or less overlapped with the site
expressing CRF type 1 receptor mRNA (6). Furthermore,
-helical CRF-(9-41) itself is known to be a dual
antagonist for CRF type 1 and 2 receptors (14). Since,
however, evidence has been shown that
-helical
CRF-(9-41) is more selective for the CRF type 2 receptor on the basis of functional and binding studies (26,
41), the effects of icv and iv injection of urocortin on the
motor activity of the gastrointestinal tract obtained in the present
study seem to be mediated mostly by CRF type 2 receptors in the brain
and peripheral organs.
It is widely accepted that CRF is involved in the postoperative or
stress-induced gastric ileus. From this point of view, in most of the
previous studies that investigated the effects of centrally or
peripherally administered CRF/urocortin on motor activities, emptying
of gastric contents has been extensively investigated (7, 11, 28,
29, 35, 49, 50, 53). A few previous studies have examined the
effects of centrally administered CRF on the gastric and duodenal
contractility measured by a strain-gauge force transducer implanted in
the stomach and duodenal wall in anesthetized rats (16,
21) and conscious dogs (28). In those experiments,
central administration of CRF inhibited the gastric contractility in
rats (16, 21) and stimulated the frequency of duodenal
contractions but inhibited the percentage of propagated duodenal
contractions in dogs (28). These opposite effects of CRF
on the contractile activity between stomach and duodenum seen in the
previous studies are consistent with the present results, in which %MI
was decreased in the stomach but increased in the duodenum by icv and
iv infusion of urocortin in the fed state of conscious rats. Both
present and previous studies show that the increase in the contractile
activity of the duodenum induced by CRF/urocortin was a nonpropagated
event. In most of the previous studies that examined the effects of
CRF/urocortin on the gastrointestinal motility, attention has been paid
only to the postprandial movement of the gastrointestinal tracts, but motor activities in the fasting state have not been considered.
The present results demonstrated that the fasted pattern of
gastroduodenal motility was disrupted by exogenously applied urocortin via icv or iv infusion. Several brain-gut peptides such as CCK, bombesin, and NPY are known as central or peripheral mediators that
regulate the fed and fasted motor activity of the gastrointestinal tract (15, 20, 47). CCK has been suggested as a mediator that causes the disruption of MMCs, which corresponds with the fasting
motility of the small intestine after a meal. Intraduodenal nutrient
stimulates the release of CCK, which disrupts the MMC through
peripheral CCKB receptors, and the released CCK acts on the
vagal afferent fibers and induces a release of central CCK, which
participates in MMC disruption through central CCKA
receptors (1, 47). Both peripheral and central
administration of bombesin cause the disruption of MMC (9,
45), and peripheral bombesin exerts the central actions through
vagal afferent pathways (57). NPY exerts the opposite
effects from CCK or bombesin; when NPY was administered icv in the fed
state of rats, the fed motor pattern of the duodenum was replaced by
the fasted motor pattern (15). The physiological
significance of fasted motor activity in the gastrointestinal tract has
been considered to be a mechanical and chemical cleansing of the empty
stomach and intestine (23) and therefore possibly linked
to the hunger sensation. In fact, icv administration of NPY stimulates
food intake and at the same time elicits the fasted pattern of the
small intestine (15). CCK and bombesin in the brain are
known as feeding-inhibitory peptides, and icv injection of CCK elicits
the disruption of fasted motor activities (20, 47). CRF
and urocortin are also feeding-inhibitory peptides and cause the
disruption of fasted motor activities, as shown in the present study.
However, unlike CCK or bombesin, CRF and urocortin are not known to be
involved in the regulation of gastrointestinal motility in normal
condition [although they may act in stress-induced alteration of
gastric and colonic motility (17, 30, 37, 39, 54, 59)],
with the exception of the data in isolated rat colon, in which the
basal colonic motility was regulated by endogenous CRF
(34). This was confirmed by the present results; neither
CRF receptor antagonist
-helical CRF-(9-41) nor
urocortin antiserum administered icv and iv altered the fed and fasted
motor patterns of gastroduodenal motility. These results suggest that
urocortin originating from the brain and peripheral organs may be not
involved in the regulation of basal gastrointestinal motility. It has
been reported that urocortin-immunoreactive neurons are widely
distributed in the brain nuclei such as the supraoptic nucleus,
paraventricular nucleus, substantia nigra, ventral tegmental area, and
Edinger-Westphal nucleus (27, 56). Urocortin or urocortin
mRNA is also detected in the peripheral sites, including
enterochromaffin cells in the intestinal epithelium of the
human (24, 25), enteric neurons of the rat
(19), and human lymphocytes (3). This central
and peripheral urocortin activated by stress or operation may alter
gastrointestinal motility.
The present results showed that when urocortin was administered icv or
iv in the fed state, the fed motor pattern remained but %MI was
decreased in the antrum and increased in the duodenum. The results
showed that increase in %MI in the duodenum induced by icv or iv
injection of urocortin was a nonpropagated event, since the transit of
nonnutrient duodenal contents was decreased by icv or iv injection of
urocortin. The inhibitory effect of urocortin on the fed motor activity
in the antrum seems consistent with previous reports in which icv and
iv injection of CRF or urocortin inhibited gastric emptying
(35-37, 41, 43, 53). The duodenal effects were also
consistent with the previous reports in which icv infusion of CRF
increased the frequency of proximal duodenal contractions and decreased
the distally propagating contractions in the whole duodenum of dogs
(28) and iv injection of CRF increased the postprandial MI
but did not alter the propagated contractions in human duodenum
(38). Gastric and duodenal movements seem to be
interrelated because gastric emptying causes subsequent duodenal
filling and, conversely, duodenal distention elicits reduction in
gastric emptying (12, 22). Furthermore, nonpropagated contractions in the duodenum act in the resistance to gastric emptying
(18). A vago-vagal reflex may play an important role in
mediating such feedback control of gastric and duodenal motility (12, 22, 28). Because, however, opposite motor responses of antrum and duodenum occur after vagotomy, the intrinsic myenteric neurons such as nitrergic or nicotinic cholinergic pathways might be
involved in this mechanism (42, 55).
To examine the involvement of autonomic nerves in the action of
centrally administered urocortin on gastrointestinal motility, the
experiments were performed in vagotomized and sympathectomized animals.
Results showed that vagotomy abolished the icv urocortin-induced change
in the gastroduodenal motility but did not alter the iv urocortin-induced change in the motor activities. Sympathectomy failed
to alter the icv and iv urocortin-induced changes in the fed and fasted
patterns in both the antrum and duodenum. These data suggest that the
effects of icv urocortin on gastrointestinal motility are mediated by
vagal nerves.
In summary, the present study examined the effects of icv- and
iv-injected urocortin on the physiological fed and fasted motor activity of the antrum and duodenum in freely moving conscious rats.
Results showed that both peripherally and centrally administered urocortin disrupted the fasted motor patterns of gastroduodenal motility when administered to animals in the fasted state, whereas urocortin given icv or iv to animals in the fed state caused a decrease
in %MI of the antrum and an increase in %MI of the duodenum. The
transit was decreased in spite of the increase in the fed motor
activity in the duodenum.
 |
ACKNOWLEDGEMENTS |
This work was supported by grants from the Ministry of Education,
Science, Sports, and Culture of Japan (M. Fujimiya, M. Fujimura, and A. Inui)
 |
FOOTNOTES |
Address for reprint requests and other correspondence: M. Fujimiya, Dept. of Anatomy, Shiga Univ. of Medical Science, Seta, Otsu,
Shiga 520-2192 Japan (E-mail:
fujimiya{at}belle.shiga-med.ac.jp).
The costs of publication of this
article were defrayed in part by the
payment of page charges. The article
must therefore be hereby marked
"advertisement"
in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
Received 3 February 2000; accepted in final form 19 September 2000.
 |
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