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-induced fever in rats
Departments of 1 Physiology and 2 Psychosomatic Medicine, Faculty of Medicine, Kyushu University, Fukuoka 812-8582, Japan
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ABSTRACT |
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Top
Abstract
Introduction
Materials & Methods
Results
Discussion
References
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We have previously reported that central
injection of PGE2 induces
hyperthermia through its actions on
EP1 receptors in rats. Because the
increase in local synthesis of
PGE2 is assumed to be a necessary
process in a fever caused by central injection of interleukin-1
(IL-1), an EP1 receptor
antagonist (SC-19220) should inhibit the IL-1-induced fever. To test
this hypothesis, we observed the effect of SC-19220 on the fever
produced by injection of recombinant human IL-1 (rhIL-1) into the
lateral cerebroventricle (LCV) in conscious rats. Administration of
SC-19220 (100 µg) into the LCV 15 min before LCV injection of
rhIL-1 (4 ng) suppressed an initial rise in colonic temperature for
30 min, producing a fever with a longer latency to onset and a longer
time to peak elevation. SC-19220, given 60 min after the central
administration of rhIL-1, also suppressed the rhIL-1-induced
fever 15-60 min after its injection. These findings suggest that
the central IL-1-induced fever in rats is mediated, at least partly,
by activation of EP1 receptors by
PGE2.
prostaglandin E2; SC-19220
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INTRODUCTION |
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Top
Abstract
Introduction
Materials & Methods
Results
Discussion
References
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INTERLEUKIN (IL)-1 is a principal component of
endogenous pyrogens, which also include inflammatory cytokines such as
IL-6, tumor necrosis factor-
, and interferon- (see review in Ref. 9). Although it has been generally recognized that IL-1 released from immune cells peripherally produces fever by signaling the brain
via various routes (2), there is evidence supporting the view that
IL-1 in the brain is also responsible for a significant portion of
fever during peripheral inflammation. IL-1 and its mRNA increase in
brain tissues including the hypothalamus not only after various types
of brain insults such as cerebral infection and ischemia (7)
but also after peripheral inflammation (20) and peripheral
administration of endotoxin (19). Fever caused by peripheral injection
of endotoxin is diminished by central injection of antiserum to IL-1
(8) and IL-1 receptor antagonist (11). Peripheral injection of both
endogenous and exogenous pyrogens increases the concentration of PGE in
the brain (1, 5, 10), and the increase in
PGE2 is inhibited by a
cyclooxygenase inhibitor (10). Furthermore, fever caused by peripheral
injection of endogenous pyrogen is reduced by an intrahypothalamic
injection of sodium salicylate (5). These findings, taken together,
suggest that IL-1 and PGE2 in
the brain are responsible for at least part of the fever after
peripheral infection and inflammation.
We have reported that hyperthermia caused by intrahypothalamic or
intracerebroventricular injection of
PGE2 in rats is mediated through
EP1 receptors (15, 16). The
PGE2-induced hyperthermia is
mimicked by an EP1 receptor
agonist,
17-phenyl-
-trinor-PGE2, but
neither by EP2 nor
EP3 receptor agonist, and is
inhibited by the simultaneous injection of an
EP1 receptor antagonist, SC-19220 (15, 16). Furthermore, the brain sites where
PGE2 and
17-phenyl--trinor-PGE2 were
microinjected to produce hyperthermia in rats are located in the
preoptic hypothalamus and its neighboring basal forebrain including the
anterior wall of the third ventricle (15), which correspond to those
sites where IL-1 may produce fever in rabbits (13). These findings
indicate the possibility that IL-1 in the brain induces fever by
stimulating EP1 receptors. In the
present study, we investigated whether the fever following
intracerebroventricular injection of recombinant human IL-1
(rhIL-1) is inhibited by intracerebroventricular injection of
SC-19220 in rats.
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MATERIALS AND METHODS |
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Top
Abstract
Introduction
Materials & Methods
Results
Discussion
References
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Male Wistar rats (Kyudo, Tosu, Japan) weighing 270-300 g were housed two or three per cage at an ambient temperature of 23 ± 1°C on a 12:12-h light-dark cycle with lights on at 0800. Food and water were given ad libitum. Under anesthesia with pentobarbital sodium (50 mg/kg ip), rats were stereotaxically implanted with a 23-gauge stainless steel cannula containing 30-gauge stainless steel wire as a stylet in the lateral cerebroventricle (LCV). The coordinates were anterior, 0.8 mm posterior to the bregma; lateral, 1.5 mm from the midline; depth, 4.0 mm from the surface of the skull. The correct placement of the cannula in the LCV was confirmed by the rise of cerebrospinal fluid in the cannula. The cannula was then fixed to the skull with acrylic dental cement. After surgery, the rats were administered sulfamethoxide (100 mg/rat ip), returned to the colony, and housed individually. During a postsurgical recovery period, the animals were placed in a cylindrical wire cage for a few hours every day to habituate them to the experimental environment.
Drugs.
rhIL-1 and SC-19220 were generous gifts: rhIL-1 (Lot No. 9K77)
from Drs. Y. Masui and Y. Hirai (Institute of Cellular Technology, Otsuka Pharmaceutical, Tokushima, Japan) and SC-19220 from Dr. R. A. Marks (Searle). rhIL-1 was dissolved in physiological saline and
stored at 
80°C and diluted with saline before use. SC-19220 was dissolved in DMSO before use, making 100 mg/3 ml of solution. All
solutions were passed through a 0.22-mm Millipore filter (Millipore, Tokyo, Japan) before injection. All glassware, syringes, and injection needles were autoclaved before use.
Experimental procedures. Experiments were performed at least 7 days after surgery. On the experimental day, each rat was loosely restrained in the cylindrical wire cage at 0900. The colonic temperature (Tco) of each animal was monitored automatically at 1-min intervals by a copper-constantan thermocouple inserted into the colon 4 cm beyond the anus. The temperature was allowed to stabilize for at least 2 h. To avoid the possibility that stress elicited by these procedures might affect the properties of Tco responses to injected drugs, all the experiments started after Tco had stabilized at normothermia and had shown no further changes for more than 20 min.
Then, the stylet was removed from the guide cannula and a 30-gauge injector cannula, connected to a 10-µl microsyringe, was inserted into the guide so that it protruded 0.5 mm beyond its tip. Drugs or the same volume of vehicles (3 or 5 µl) was delivered into the LCV over 60 s/µl. At the end of the injection, the injector cannula was kept in position for ~1 min and then quickly replaced by the stylet. There was no back flow of the fluid in this procedure. In the experiments, SC-19220 (100 µg/3 µl) or the vehicle, DMSO (3 µl), was injected into the LCV 15 min before or 60 min after the intracerebroventricular injection of either rhIL-1 (4 ng/5 µl) or
physiological saline (5 µl). The time when injection of rhIL-1 or
saline was started was designated as time
0, and changes in
Tco were observed during the
subsequent 4 h. All experiments were performed at a room temperature of
23 ± 1°C. Each rat was used for two experiments on different days
at least 7 days apart.
Data analysis. The values are presented as means ± SE. Significant differences were assessed by one-way ANOVA followed by Scheffé's test. Differences were considered to be significant at P < 0.05.
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RESULTS |
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Top
Abstract
Introduction
Materials & Methods
Results
Discussion
References
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In the first series of experiment, we injected SC-19220 (100 µg/3
µl) or the same volume of DMSO into the LCV 15 min before intracerebroventricular injection of rhIL-1 (4 ng/5 µl) or saline. The mean Tco at
time
zero of each group ranged from 37.87 ± 0.03 to 38.24 ± 0.09°C and did not differ significantly from
each other. The rats treated with DMSO followed by rhIL-1 injection
showed a rise in Tco that became
apparent 30 min after injection, reached a peak (1.06 ± 0.05°C)
at 120 min, and then remained at this level until the end of the
observation period (240 min) (Fig. 1). An administration of SC-19220 15 min before rhIL-1 injection suppressed an initial rise in Tco for 30 min,
producing a fever of similar magnitude (1.06 ± 0.15°C), but with
slower onset (60 min) and longer time to reach a peak (165 min).
Furthermore, pretreatment with SC-19220 significantly attenuated the
rhIL-1-induced rise in Tco
75-90 min after rhIL-1 injection. No changes in
Tco occurred in the SC-19220 + saline-treated rats and the DMSO + saline-treated rats.
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In the second series of experiment, we injected SC-19220 (100 µg/3
µl) or DMSO into the LCV 60 min after intracerebroventricular injection of either rhIL-1 (4 ng/5 µl) or saline. The mean
Tco at
time
zero of each group ranged from 37.91 ± 0.17 to 38.29 ± 0.08°C and did not differ significantly from
each other. The rats that received rhIL-1 followed by injection of
DMSO 60 min later produced fever with similar time courses and
magnitude (Fig. 2) as that of rats that
received DMSO followed by rhIL-1 (Fig. 1). The similar rise in
Tco in rhIL-1 + SC-19220-treated rats was significantly inhibited by SC-19220, when
given 60 min after rhIL-1 injection, from 75 to 120 min (Fig. 2).
The antipyretic activity of SC-19220 was maximal 30 min after its
injection. Thirty minutes thereafter,
Tco returned to the level at the
time of SC-19220 injection and then continued to rise maximally to a
level that was similar to that of rhIL-1 + DMSO-treated rats. There
was no change in Tco in the saline + SC-19220-treated rats and the saline + DMSO-treated rats during the
observation periods.
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DISCUSSION |
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Top
Abstract
Introduction
Materials & Methods
Results
Discussion
References
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The present study demonstrates that SC-19220, a specific
EP1 receptor antagonist (18), may
inhibit fever induced by an intracerebroventricular injection of
rhIL-1 in the rat. IL-1 is known to specifically increase the
release of PGE2 from the
hypothalamic explants (14). We have reported that the hyperthermia
induced by the central injections of
PGE2 is blocked by SC-19220 (15,
16). Furthermore, the central injection of an
EP1 agonist,
17-phenyl--trinor-PGE2, produced a rise in Tco (15, 16).
These findings, taken together, suggest that the central
IL-1-induced fever is mediated, at least partly, through activation
of EP1-receptors by
PGE2 in the rat.
The present finding does not conform well with that of a previous study
(3), which demonstrated the failure of SC-19220 (intracerebroventricularly) to affect the leukocyte pyrogen
(intracerebroventricular)-induced fever in rabbits despite its ability
to suppress the PGE2
(intracerebroventricular)-induced hyperthermia. Although this
discrepancy might be due to the difference of animal species, one may
discuss the different protocols of two experiments as the possible
causes, i.e., the timing of administration of SC-19220 in reference to
that of pyrogens, the dose of SC-19220, and the content of leukocyte
pyrogen. The difference in the timing of injection of SC-19220 in the
previous study and ours is unlikely to be the cause of different
results. Because an administration of SC-19220 either before or after
injection of rhIL-1 suppressed the fever in rats in the present
study, SC-19220 given immediately after injection of leukocyte pyrogen
could have affected the fever.
The amount of SC-19220 (15 µmol) administered into the LCV in the previous study (3) is much greater than the amount given (~330 nmol) in our study, although the doses per body weight are similar in both experiments. Because of a low water solubility of SC-19220, it was dissolved in DMSO in both experiments. However, the antagonist dissolved in DMSO, when it is injected into the LCV, is likely to precipitate more or less out of solution (3). If the formation of precipitate of SC-19220 reduces its access to PGE2, hyperthermia caused by PGE2 released in response to leukocyte pyrogen is less likely to be inhibited by the antagonist than that due to exogenously administered PGE2. Because it is suggested that the greater amount of SC-19220 produces the the greater precipitation, the precipitation might explain the previous negative finding (3) on the effect of SC-19220 to affect the leukocyte pyrogen-induced fever despite its antipyretic action on the PGE2-induced hyperthermia.
Finally, the possible involvement of cytokines other than IL-1
and/or PGs other than PGE2
in the leukocyte pyrogen-induced fever deserves attention as a cause
for the failure of PGE2
antagonists to affect it. Although the leukocyte pyrogen presumably
contains IL-1, other cytokines such as macrophage inflammatory
protein-1 and IL-8, which are suggested to produce fever independently
of the synthesis of prostanoids (12, 21), might contribute more to the
development of fever by the leukocyte pyrogen. The involvement of other
prostanoids such as PGF2
or
PGD2, which were reported to
exhibit hyperthermic actions in rabbits (13), might also be considered
in the leukocyte pyrogen-induced fever in rabbits. In rats, however,
PGD2 produces hypothermia (17),
and PGF2 is less potent in
inducing hyperthermia than PGE2
(6).
Although some findings support the view that IL-1 is produced in the
brain during the systemic inflammation (8, 11, 19, 20), it is not
completely known how much concentration of IL-1 is induced and to
what extent such centrally induced IL-1 contributes to the fever
during the peripheral inflammation. In this regard, the experimental
design (LCV injection of IL-1) in the present study does not seem to
be appropriate for determining what type(s) of EP receptors in the
brain mediate the fever during systemic inflammation. However, the
present result suggests the involvement of
EP1 receptors in the central
IL-1-induced fever that is associated with various types of brain
insults such as meningitis, brain injury, and cerebral ischemia
(7).
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ACKNOWLEDGEMENTS |
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This experiment was reviewed by the Committee of Ethics on Animal Experiments in the Faculty of Medicine, Kyushu University and was carried out under the control of the Guidelines for Animal Experiments of the Faculty of Medicine, Kyushu University, and the law (no. 105) and Notification (no. 6) of the government.
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FOOTNOTES |
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This work was supported by Grants-in-Aid for General Scientific Research (no. 09557006 and no. 10307001 to T. Hori and no. 09770040 to T. Oka) from the Ministry of Education, Science and Culture, Japan.
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. §1734 solely to indicate this fact.
Address for reprint requests: T. Hori, Dept. of Physiology, Kyushu Univ. Faculty of Medicine, Fukuoka 812-8582, Japan.
Received 26 March 1998; accepted in final form 5 August 1998.
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REFERENCES |
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Top
Abstract
Introduction
Materials & Methods
Results
Discussion
References
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, DMSO + rhIL-1
, SC-19220 + rhIL-1
, DMSO + saline (n = 5); 
, SC-19220 + saline (n = 5). Each point represents
mean ± SE. Tco, colonic
temperature; 
, change. ** Significant difference
(P < 0.01, one-way ANOVA followed by
Scheffé's test) compared with DMSO + saline-treated rats.
# Significant difference
(P < 0.05, one-way ANOVA followed by
Scheffé's test) compared with DMSO + rhIL-1