THE Jouan.i.
OF PHARMACOLOGY
Copyright C 1994 by The American
AND
EXPERIMENTAL
Society
VoL 269, No.2
in USA
THERAPEUTICS
for Pharmamiogj
and Experimental
Therapeutics
Printed
Antagonism of Nitrous Oxide Antinociception in the Rat
Hot Plate Test by Site-Specific Mu and Epsilon Opioid Receptor
Blockade13
BRADLEY
L. HODGES,4
MICHAEL
J. GAGNON,
THOMAS
A. GILLESPIE,45
DAVID
F. O’LEARY,7
SHUICHI
HARA8 and RAYMOND
M. QUOCK
of Biomedical
D,artment
Accepted
for publication
Sciences,
University
of Illinois College
of Medicine
rats.
was
was
Subanesthetic
duce a prominent
concentrations
of the anesthetic
gas N20 proanalgesic
effect in both human subjects
and
experimental
animals
(Finck,
1985; Marshall
and Longnecker,
1990). Inhalation
of 20% N20 in 02 reportedly
produces
an
1This
‘p.:
for publication
June
research
was supported
was approved
28, 1993.
by Public Health
by the Biologic
Service grant DE-06894.
Resource
Committee
of
the
Illinois
College
of Medicine
at Rockford
and was conducted
in
accordance
with the Guidefor
the Care and Use ofliiborotoryAnimois
as adopted
and promulgated
by the National
Institutes
of Health.
I Preliminary
reports of these results
were presented
at the 1992 annual
meeting of the Federation
of American
Societies
for Experimental
Biology
(O’Leary, et al., 1992), the 1992 fall meeting
of the American
Society
for Pharmacology
and Experimental
Therapeutics
(Hodges
and Quock,
1992) and the
Experimental
Biology 93 Meeting (Hodges et aL, 1993).
4Present address: University oflllinois at Urbana-Champaign,
Urbana-Chainpair’, IL
I Supported
by a High School Student
Summer Apprentice
Fellowship
from
the Rockford
Regional
Academic-Industrial
Consortium
(RAIC,
Rockford,
IL).
S Present
addresaC David Grant
U.S.A.F.
MediCal
Center,
Travis
Air Force
Base, CA.
7 Supported
by a High School Science Teacher Summer Research
Fellowship
from the Pierce
Chemical
Company
(Rockford,
IL). Present
address:
Crystal
Lake Central
High School, Crystal
Lake, IL
S Visiting
Scientist
from the DepartmentofForensic
Medicine,
Tokyo
MediCal
College, Tokyo,
Japan.
University
of
Illinois
,
of
was
2
Rockford,
R. BRENEISEN,6
January 31 1994
ABSTRACT
The an&gesic
property
the anesthetic
gas N20 has long been
known and used to treat pain in clinical medicine
and dentistry.
The present
study
cOndUCted
to ldenttfy by subtype and
possible
location
the brain opiold
receptors that mediate N2O
antinociception
in
A 5-mm exposure to 70% N20 consistently evoked an antinodceptive
effect in the hot plate test. This
drug effect
partly antagonized
in dose-related
fashion by
i.c.v. pretreatment
with naltrexone,
D-Phe-Cys-Tyr-D-Trp-OmThr-Pen-Thr-NH2
and -endorphin1.,
which block multiple,
mu
and epsilon oc1
receptors,
respectively.
However,
the N20evoked antinociception
unaffected
by i.c.v. pretreatment
with either the delta oploid antagonist
naftrindole or the kappa
Received
at Rockford,
JEFFREY
oploid antagonist
nor-binaltorphimine.
When D-Phe-Cys-Tyr-DTrp-Om-Thr-Pen-Thr-NH2
administered
intracerebrally
direcdy
Into the pedequeductal
gray, N20 antinociception
was
partly antagonized
dose-dependent
manner. The antinocicaptive
response
N2O was uninfluenced
by -endorphin1.
administered
into the petiaqueductal
gray. The findings
these
pharmacologicai
antagonism
studies
are consistent
with the
hypothesis that exposure
N2O causes a neuronal
release
$-endorphin.
These results
indicate that supraspinal
mu and
epsilon
oc1
receptors
medte
N2O antinockeption
in the rat
hot plate paradIgm
and that one central site of such mu but not
epsilon oploid receptors
the periaqueductal
gray.
was
in a
to
of
to
of
is
analgesic
effect in human subjects
approximately
equivalent
to
that evoked
by 15 mg of morphine
sulfate
(Chapman
et aL,
1943; Parbrook
et aL, 1964).
The clinical
literature
includes
reports of the specific
application
of N20 analgesia
in dental
surgery (Dworkin
et aL, 1983; Hammond
and Full, 1984; Smith
and Beirne,
1985), in obstetrics
(Marx and Bassell,
1985), in
emergency
medicine
(Thai et at., 1979; Donen et at., 1982) and
in the treatment
of selected
types
of pain (Thompson
and
Locon,
1976; Fosburg and Crone, 1983; Henderson
et aL, 1990).
The mechanism
of this analgesic
effect of N20 has long been
attributed
to a nonspecific
depresaion
of CNS function
(Sonnenschein
et aL, 1948).
An opioid
mechanism
was initially
suggested
by Berkowitz
et aL (1976),
who first reported
that
N20 antinociception
in mice was sensitive
to antagonism
by
naloxone,
an opioid
receptor
antagonist.
This
finding
was
quickly
verified
by other studies,
both in experimental
animals
(Smith
and Bees, 1981; Lawrence
and Livingston,
1981) and
human subjects
(Chapman
and Benedetti,
1979; Giliman
et aL,
1980; Yang et at., 1980).
The discovery
of opioid receptor
multiplicity
and the differential sensitivity
of these receptor
subtypes
to various
opioid
ABBREVIAT1ONS
EOP, endogenous
o#{231}ioki
peptide; ME, methionk*.enkephaln;
-EP, -endorphi;
-EP1,
ndorptn1;
11-50,48811,
tra±)-3,4-dhloro-N-methy1-N-[2-(1-pyrrolldkiy1)cyclohexy1]-benzeneacetamIdemethane
sultonate; NTX, naltrexone; CTOP, D-Phe-Cys-Tyr-o-TrpOm-TIw-Pen-Tlw-NH2;
NTI, naltrindole; nor-BNI, nor-binaftorphETilne;
PAG, perlaqUedUCtal gray; CNS, central nervous system; l.c., Intracerebral.
596
N20 and Opioid Receptors
1994
(Lord et at., 1977) was followed
by the development
of newer and more selective
opioid receptor
antagonists.
The
introduction
of such drugs makes it possible
to identify
more
precisely
the central
opioid receptors
that mediate
N20 antinociception.
Another
objective
of the present
investigation
was
to shed some light on the possible
site of action of N20 in the
CNS.
antagonists
N20
more
than
N20.
The
cause
Drugs.
The
Belmont,
search
Biochemicals,
NTI
rats
(250-300
g) were
purchased
from Sasco Inc. (Omaha,
NE) and used in this research.
The animals
were housed in temperatureand humidity-regulated
quarters
with a
12-hr light/dark
cycle. Food and water were available
ad libitum.
Stereotaxic
surgery
and
central
microinjection
in rats. The
rats were anesthetized
with sodium
pentobarbital
(50 mg/kg
i.p.),
treated
with atropine
methylbromide
(4.0 mg/kg
i.p.) and then mounted
in a stereotaxic
headholder
(David Kopf Instruments,
Tujuga, CA). A
26-gauge
guide cannula
(Plastic
Products,
Roanoke,
and
testing
under
N20.
The
rats
were
ex-
the box through
an inflow
port at one end, circulated
and exited through an outflow port at the other end.
the box
in both
the
uously
monitored
(Criticare,
smaller
exposure
box
by using
Milwaukee,
which was covered with a heavy
of the gas. The levels of N20 and Q,
and the larger enclosure were contin-
a POET
WI).
The
II anesthetic
temperature
at
monitoring
the
surface
system
of
the
was 52.O’C. The response
time to paw licking or escape behavior
recorded. A maximum cutoff time of 60 sec was used.
Base-line
response times were determined
for all rats before antinociceptive
testing under 70% N2O. Then rats were pretreated
i.c.v. or
was
or one of several
interval,
the
doses
animals
of opioid
were
receptor
exposed
to
antagonists.
70%
N2O
injected
i.c.v. or i.e. with a dye marker
cannula
placement.
Preliminary
studies
3 weeks
for histological
determined
that
and
NTI
(Research
(Re-
Biochem-
Regent,
inhalation
prepared
Shirley,
described.
The
NY).
doses
drugs
albeit
i.c.v.
injections
of vehicle
analysis
control
1981).
To
and
various
were
used
saline).
of data.
In these
Before
and
the analysis
antinociception)
were
whether
or completely
conducted
by parametric
N20
to compare
treatment
group
and
was
the
pretest
its
response
significance
N2O
between
and
Murray,
by
one-
only
partly
antago-
a paired
time
time
data
transformation.
pretreatment,
response
fi-
received
the percentage
by arc-sin
by any drug
with
Murray,
statistics,
of
and
determined
(Tallarida
antinociception
abolished
the
to 70%
was
normalized
effects
animals
experiments,
t test
study
and were verified
Control
groups
Dunnett’s
i.c.v.,
U-50,488H
responsiveness
and
which
in this
antinociceptive
test.
(sterile
CTOP
solution,
administered
DPDPE,
pretreatment
of variance
determine
nized
the
(sufentanil,
in antinociceptive
analysis
(percent
drugs
NTX,
chloride,
drugs
saline
antagonists
not all in the rat hot plate
difference
way
sodium
peptide
sterile
of
agonist
the
The
earlier.
0.9%
based on doses found in the literature
opioid
EP),
as described
in sterile
of each
t test
animal
during
N20
the
response
in
exposure
1981).
A 5-mm
to hind
to
exposure
paw
lower
to 70% N20
prolonged
or, occasionally,
escape
concentrations
a consistent
Figures
with
licking
of N20
antinociceptive
1, 2 and 3 show
(25% and 50%)
effect.
the influence
time
Exposure
did not produce
of i.c.v.
pretreatment
receptor
antagonists
on the antinociceptive
response
to 70% N20.
Increasing
i.c.v. doses of NTX
significantly reduced
N2O antinociception
(fig. 1). Maximum
antagonism
NTX
various
attempt.
opioid
occurred
did not
at
10 g
produce
of
NTX.
greater
An
i.c.v.
antagonism
dose
of 20
of N20.
g
of
Further
z
-I-
80
0
LI
60
*
0
z
40
20
nI#{149}
5
10
20
5
-
time - base-line
response
time.
generally
used weekly
for 2 to
by
freshly
SALINE
for
mm and retested on the hot plate. The percent of antinociception
for
each rat was calculated
as 100 x test response
time
base-line
response
time/60-sec
cutoff
The rats were
nor-BNI
hot
plate
i.e. with vehicle
After a 15-mm
U.S.P.,
100
a Plexiglas
enclosure
(30 cm long x 30 cm wide x
which flowed the same concentration
of N2O in
testing,
the same concentration
of N2O in 02 was
circulated
through the enclosure,
Plexiglas lid to prevent the escape
N20,
Laboratories);
and
were made up in the same
0.01% Triton
X-100.
These
The exhausted gas was vented to a nearby fumehood.
After
a 5-mm exposure
period,
the animal
was quickly
removed
from
the box and dropped onto a hot plate analgesiometer
(I.I.T.C.,
Woodland Hills, CA) with
45 cm high) through
02. For 5 mm before
to
uncertain.
Results
in an enclosed
Plexiglas
(Abbott
Plastics,
Rockford,
IL)
box (25 cm long x 10 cm wide x 10 cm high) with a sliding lid. A
mixture
of 70% N2O and 30% 02 was delivered
into the box from a
standard
N2O/O2 dental sedation system (Porter, Hatfield, PA). The
rate of inflow
was
10 liters/mm
(7.0 liters/mm
N2O plus 3.0 liters/mm
through
were
(American
(Tallarida
to N20
02). Gas entered
(Peninsula
selective
each
rate of i.c.v. or i.c. infusion
was 1.0
solubility
and needed dose, the volwere generally
5 to 10 tl and 1.0
in rats
administered
Statistical
The drugs were administered
i.c.v. by using a 10-tl microsyringe
(Hamilton,
Reno, NV) mounted
in a model 341A syringe pump (Sage
Instruments,
Cambridge,
MA). A length of polyethylene
tubing connectedthe microsyringe
to a 33-gauge
internal
cannula
inserted
through
the guide cannula and projecting
1 mm beyond
the tip of the guide
Antinociceptive
of sensitivity
time
Welding,
Rockford,
IL); NTX
DE); CTOP (Peninsula
Labora-
MA);
to antagonize
was
posed
$-EP127
Wayland,
studies
VA) was implanted
experimentation.
cannula
into the target
site. The
d/min.
Depending
on the drug’s
umes of i.c.v. and i.e. microinjection
l, respectively.
in a loss
is at this
were used in this study:
in preliminary
7 days
before
was
as previously
were initially
ofthe
time
drugs
CA);
nor-BNI
fl-EP127
contained
directed
toward the lateral cerebral ventricle (stereotaxic
coordinates
0.0 mm anterior-posterior,
1.5 mm lateral and -3.0 mm dorsalventral,
Paxinos and Watson, 1982) or the PAG (-6.3
mm anteriorposterior,
0.2 mm lateral and -4.0 mm dorsal-ventral).
Cannulae
were
secured
to the calvarium
by using stainless
steel screws and dental
cement.
Each cannula
was plugged
with
a solid
33-gauge
dummy
cannula.
All i.c.v.- and i.c.-cannulated
rats were allowed a minimum
of
and
of recovery
resulted
of sensitivity
icals).
U.S.P.
Sprague-Dawley
times
loss
following
tories,
N2O/O2
Male
to times
and 0),, U.S.P.
(Rockford
Industrial
(DuPont
Laboratories,
Wilmington,
Methods
Animals.
two
of this
597
then
verification
of
exposure
to
Naltrexone
(tg/rat)
Fig. 1. Influence of icy. pretreatment
with increasing
doses of NTX or
vehicle on the antinociceptive
response to 70% N20 in rats. Each bar
indicates the mean antinociceptive response and each vertical line mdicates the S.E.M. of six to eight rats per group. Significance of difference:
*P < .05 compared with the vehicle pretreatment
control group.
598
Hodges
Vol. 269
et al.
respectively,
nociception
100
4 shows
Figure
z
80
on the
0
0.
LI
LI
60
0
z
I-
were generally
ineffective
(data not shown).
the
influence
antinociceptive
antagonism
administration
into
significantly
different
N20
of i.e. pretreatment
response
dose-related
in reducing
to 70%
N20.
with
CTOP
from
response
times
CTOP
produced
of N20 antinociception
PAG.
Pretest
response
the
anti-
after
times
under
N20
direct
were
at all i.e.
doses of CTOP.
On the other hand, similar
pretreatment
with
fl-EP127
(0.5-1.0
tg) directly
into the PAG had no appreciable
effect on N20 antinociception
(data not shown).
40
z
20
Discussion
0.5g
SALINE
1.0ig
In the present
antinociceptive
5.0g
generally
Dose
of
CTOP (tg)
study, exposure
of rats to 70% N20
response
in the rat hot plate test.
consistent
with
the
results
of earlier
evoked an
This was
studies
that
used
oploid
antagonmst CTOP or vehicle on the antinocmceptive response to
70% N20 in rats. Each bar indicates the mean antinociceptive
response
and each vertical line indicates the S.E.M. of six to eight rats per group.
Significance of difference:
< .05 compared
with the vehicle pretreat-
paradigm
(Smith and Rees, 1981; Novelli
et aL, 1983).
to 25% and 50% N20 did not produce
consistent
antinociception,
which suggests
a steep dose-response
curve for
N20 antinociception
in this paradigm;
this was in agreement
with the findings
of Novelli
et at. (1983). Zuniga
et at. (1987a)
ment control group.
also
Influence of cv. pretreatment
Fig. 2.
with increasing
doses of the mu
the same
Exposure
observed
an unexpectedly
tive response
The
100
first
that
N20
antinociception
N20
antinociception
was, at least in part,
Berkowitz
et at. (1977)
in the
rat tail
flick
test
was
kg of naloxone.
The antinociceptive
40
z
20
to 70%
N20
was
sensitive
to
100
0
SALINE
1.0
3.0
Dose
5.0
of P-EP127
7.0
10
20
z
(.tg)
control
80
0
Fig. 3. Influence of cv. pretreatment
with increasing doses of the
epsilon
oploid antagonist
fl-EP1v
O’ vehicle on the antinociceptive
response to 70% N20 in rats. Each bar indicates the mean antinociceptive
response and each vertical line indicates the S.E.M. of six to eight rats
per group. Significance
of difference:
*)
< .05 compared
with the vehicle
pretreatment
response
antagonism
by i.c.v.-administered
NTX
in a dose-dependent
manner.
However,
although
relatively
selective
for mu opioid
receptors
at lower doses,
antagonists
such as naloxone
and
NTX lose their selectivity
at higher doses and may occupy all
opioid receptor
subtypes
(Lord et at., 1977). Nonetheless,
maximum antagonism
was achieved
by an i.c.v. dose of 10 ig of
NTX and doubling
the dose to 20 g did not antagonize
further
0
I-
for
60
U
z
basis
of antinocicep-
80%.
partly antagonized
by 5.0 to 30 mg/kg of naloxone.
Zuniga et
aL (1987a)
likewise
reported
that N20 antinociception
in the
rat tail flick and foot flick tests was partly reduced
by 10 mgI
a-
U
of an opioid
below
80
0
UJ
inkling
variability
of N20
emerged
from observations
that N20
antagonized
by opioid receptor
blockade.
reported
z
large
at concentrations
0.
uJ
LI
LI
60
0
z
*
40
z
group.
*
20
analysis
showed
significant
differences
between
pretest
re-
sponse times and those during N20 exposure
in both of these
pretreatment
groups (10 and 20 g of NTX).
Increasing
i.c.v.
doses of the selective
mu opioid
antagonist
CTOP
resulted
in
partial antagonism
of N20 antinociception,
which was maximal
at 1.0 zg (fig. 2). Increasing
i.c.v. doses of the epsilon
opioid
antagonist
-EP127
resulted
initially
in a dose-dependent
antagonism
between
1.0 and 5.0 tg followed
by reversal
antagonism
at higher
doses
(7-20
zg) of fl-EP127
(fig.
i.c.v. pretreatments
with NT!
(5-100
g)
and nor-BNI
Lg),
which
are selective
for delta
and
kappa
opioid
of the
3). The
(20-50
receptors,
0
SALINE
0.2
0.5
Dose of
Fig. 4. Influence
of ic. pretreatment
CFOP
.0
(tg)
in the PAG with increasing
doses of
the mu opiold antagonist CTOP or vehicle on the antinociceptive
response to 70% N20 in rats. Each bar indicates the mean antinociceptive
response and each vertical line indicates the S.E.M. of six to eight rats
per group. Significance
of difference:
< .05 compared
with the vehicle
pretreatment control group.
1994
N20 and Oplold
the residual
antinociceptive
response.
The response
times of
N20-exposed
rats after pretreatment
with 10 and 20 g of NTX
continued
to be significantly
greater than the pretest
response
times. This might suggest
a NTX-resistant
or nonopioid
cornponent
to N20 antinociception.
Which
receptor
subtypes
are
then involved
in mediating
the opioid component
of the antinociceptive
effect of N20 in the rat hot plate test?
The results
of the present
study suggest
that N20 antinociception
is mediated
primarily
by supraspinal
mu and epsilon
opioid receptors
in the brain. The i.c.v. pretreatment
with the
somatostatin
analog CTOP
produced
a partial
antagonism
of
N20 antinociception.
CTOP
has been shown
to bind selectively
to mu opioid receptors
(Kramer
et at., 1989)
and antagonize
selectively
the effects
of morphine
and DAMGO
but not
DPDPE
or U-50,488H
(Fanselow
et at., 1989; Tseng and Wang,
1992). $-EP127,
which is a putative
epsilon
opioid antagonist
(Suh et aL, 1988), produced
dose-related
antagonism
of N20
antinociception
only at i.c.v. doses up to 5.0 g. At i.c.v. doses
exceeding
5.0 big, there was a reversal
of -EP1.27
antagonism
of N20 antinociception.
One possible
explanation
for this biphasic influence
of -EP127
is that fi-EP127 has partial agonist
properties
at higher doses (Haminonds
et al., 1984; Suh et at.,
1988). The wide range of individual
antinociceptive
responses
to 70% N2O in rats pretreated
with 7.0 tg of fl-EP1..27 indicated
that this dose might be near the threshold
concentration
that
separates
the antagonist
and partial
agonist
activities
of 3EP127.
However,
the data presented
in figure 3 alone do not
distinguish
between
a loss of epsilon
opioid antagonist
activity
or an increase
in (or modulation
of) epsilon
opioid
agonist
activity.
Supraspinal
delta and kappa opioid receptors
appear not to
be involved
in N20 antinociception
in the rat hot plate model.
This is strongly
suggested
by the failure of i.c.v. pretreatment
with several doses ofNTI
and nor-BNI
to exert any appreciable
effect on the antinociceptive
effect of 70% N20. The initial
pretreatment
doses of these opioid antagonists
were sufficient
to attenuate
the antinociceptive
effects
of i.c.v.-adininistered
DPDPE
in the rat hot plate test and i.c.v.-adniinistered
U50,488H
in the rat abdominal
constriction
test (unpublished
observations).
This demonstration
of mu and epsilon
opioid
receptor
involvement
in N2O antinociception
is in contrast
to
the opioid
receptor
subtypes
that apparently
mediate
N2O
antinociception
in other rat paradigms
and other species.
Earlier, we found that i.c.v. pretreatment
with the kappa and mu
opioid receptor
antagonist
(-)-5,9-diethyl-a-5,9-dialkyl-2-hydroxy-6,7-benzomorphan,
although
not the selective
mu opioid
receptor
antagonist
9-funaltrexamine,
significantly
reduced
N20 antinociception
in the warm water tail withdrawal
test in
rats (Quock
et at., 1990b).
In other
research,
we demonstrated
that kappa opioid receptors
mediate
N2O antinociception
in the
mouse
1988;
abdominal
constriction
paradigm
(Quock
and
Graczak,
et aL, 1990a; Quock
and Mueller,
1991).
Where
is the location
of these opioid receptors
involved
in
mediating
the antinociceptive
effect of N20 in the rat hot plate
test? When administered
directly
into the PAG, CTOP caused
a dose-related
antagonism
of N20 antinociception,
again leaving a residual
antinociceptive
effect. The PAG is a site of action
of morphine,
which acts on mu opioid receptors
(Yaksh
et at.,
1976). High levels of$-EP
are found in the PAG, which indicate
a functional
role there (Palkovits
and Eskay,
1987).
On the other hand, -EP127
was not effective
in reducing
N2O antinociception
when administered
directly
into the PAG.
Quock
599
Receptors
site of the epsilon
opioid involvement
in N20 antinociception remains
unknown,
but possible
sites might
include
the
raphe obscurus
nucleus
and raphe pallidus
nucleus in the caudal
medulla
oblongata
or the medial posterior
nucleus
accumbens,
medial
preoptic
area and arcuate
hypothalamic
nucleus
in the
forebrain,
where i.e. microinjection
of f-EP
produces
significant inhibition
of the tail flick response
in rats (Tseng
et at.,
The
1990;
Tseng
and
Wang,
1992).
How does N20 interact
with these brain opioid receptors?
Berkowitz
et at. (1979)
observed
a unilateral
cross-tolerance
between
N20 and morphine,
in which morphine-tolerant
animals were nonresponsive
to N2O but N20-tolerant
animals
were still sensitive
to morphine.
They suggested
that the antinociceptive
effect of N20 might be secondary
to N2O-stimulated
release
of EOPs
from central
opioid
neurons.
Accordingly,
chronic
exposure
to N20 resulted
in excessive
depletion
of
EOPs
(N20 tolerance)
in the absence
of down-regulation
of
post8ynaptic
opioid receptors
(lack of cross-tolerance
to morphine).
In earlier studies,
we reported
that exposure
to N20
increased
levels of ME but not leucine-enkephalin
or $-EP in
fractions
of artificial
cerebrospinal
fluid collected
from ventricular cisternally
perfused
rats (Quock
et at., 1985).
Increases
in
ME and ME-Arg-Phe7
but not leucine-enkephalin,
fi-EP or
dynorphin
A have also been
found in cerebrospinal
fluid collected from the third ventricle
of dogs exposed
to 66% to 75%
N2O (Finck et at., 1990). Exposure
of rats to N20 also increased
regional
brain levels of ME (Quock
et at., 1986; Silverstein
et
at., 1992ab)
and f-EP
(Zuniga
et at., 198Th; Silverstein
et at.,
1992b).
Zuniga et at. (1987c)
also showed
that N20 stimulated
the secretion
of fl-EP from basal hypothalamic
cells attached
to microcarrier
beads in an in vitro perfusion
system.
If N20 does indeed induce neuronal
release of ME and/or fiEP, which EOP is responsible
for the antinociceptive
effect of
N2O in the rat hot plate test or are both EOPs involved
in N2O
antinociception?
Because
-EP
has been shown to exert agonist
activity
at mu and epsilon
opioid receptors
(Suh et at., 1988;
Shook
et at., 1988; Bals-Kubik
et at., 1990),
our results would
appear to support
the -EP
hypothesis
(Zuniga et at., 198Thc).
This
tion
is also
in rats
consistent
is blocked
with the
by i.c.v.
finding
that N2O antinociceppretreatment
with
antiserum
(Hara et at., 1993; Hara
et at.,
against
$-EP but not ME
in
press).
In conclusion,
the findings
of the present
research
indicate
that
central
mu and epsilon
opioid
receptors
mediate
N20
antinociception
in the rat hot plate test. One brain
locus of
participating
mu opioid
receptors
is the PAG; the involved
epsilon
opioid receptors
appear
to be located
elsewhere
in the
CNS.
However,
delta and kappa opioid receptors,
at least supraspinally,
do not appear
to be involved
in mediating
N20
antinociception
in this paradigm.
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PhD.,
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basal hypothalsystem.
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