0022-3565/97/2801-0384$03.00/0
THE JOURNAL OF PHARMACOLOGY AND EXPERIMENTAL THERAPEUTICS
Copyright © 1997 by The American Society for Pharmacology and Experimental Therapeutics
JPET 280:384 ––392, 1997
Vol. 280, No. 1
Printed in U.S.A.
Effects of a Novel Cholinergic Ion Channel Agonist SIB-1765F
on Locomotor Activity in Rats
FRÉDÉRIQUE MENZAGHI, KEVIN T. WHELAN, VICTORIA B. RISBROUGH, TADIMETI S. RAO and G. KENNETH LLOYD
SIBIA Neurosciences, Inc., La Jolla, California
Accepted for publication August 8, 1996
Conflicting reports have appeared in the literature showing stimulant or depressant effects of nicotine on locomotor
behavior in rats. This discrepancy may be explained by differences in age/weight, housing conditions, time of day or
strain; but, most importantly, may be attributed to the environmental experience of the rats before testing. Indeed, acute
administration of nicotine to rats naive to the test apparatus
reduces the high levels of exploratory locomotor activity seen
upon exposure to this novel environment. This effect seems to
be related primarily to side effects induced by nicotine and
may be followed by a slight increase in locomotor activity as
the rats recover and habituate to the test apparatus (Stolerman et al., 1973; Clarke and Kumar, 1983). Repeated administration of nicotine produces a rapid tolerance to the depressant locomotor effects seen in this setting (Morrison and
Stephenson, 1972; Stolerman et al., 1973, 1974). Alternatively, when the basal level of locomotor activity is low due to
habituation of the rats to the test apparatus, acute administration of nicotine increases locomotor activity after an insignificant depressant effect (O’Neill et al., 1991; Benwell and
Balfour, 1992). With low basal activity, sensitization to the
stimulant effect of nicotine develops rapidly after repeated
Received for publication April 26, 1996.
Furthermore, SIB-1765F produced a larger and longer-lasting
increase in locomotor activity when administered to rats familiar
with the test apparatus. Mecamylamine and dihydro-b-erythroidine but not hexamethonium blocked the increase in locomotor activity induced by SIB-1765F, suggesting that SIB1765F elicits this effect predominantly through the activation of
central NAChR. The SIB-1765F-induced increase in locomotor
activity was also attenuated by selective D1 and D2 dopamine
receptor antagonists, implying that this increase in locomotor
activity is mediated through the activation of dopamine receptors subsequent to the release of dopamine. Based on these
results, SIB-1765F appears to have a different locomotor activity profile than nicotine and epibatidine.
administration of the compound (Clarke and Kumar, 1983;
Ksir et al., 1987; Benwell and Balfour, 1992). Thus, nicotine
may decrease or increase locomotor activity, depending on
previous exposure of the animals to the drug and/or to the
test apparatus.
Interestingly, other NAChR agonists produce different effects on locomotor activity. For instance, lobeline, anatoxin
and isoarecolone decrease locomotor activity in rats not habituated to the test environment, but fail to increase locomotion in conditions where nicotine produces a stimulant effect
(Reavill et al., 1990; Stolerman et al., 1992, 1995; Whiteaker
et al., 1995). The different effects of NAChR agonists on
locomotor activity may be the result of differential effects on
DA release (Mirza et al., 1996). The locomotor stimulant
effect of nicotine is predominantly mediated through action
on the mesolimbic DA system, at the level of the cell bodies in
the ventral tegmentum area (Reavill and Stolerman, 1990;
Museo and Wise, 1990; Leikola-Pelho and Jackson, 1992)
and/or at the level of DA terminal fields in the nucleus
accumbens (Imperato et al., 1986, Damsma et al., 1989; Welzl
et al., 1990). The striatum also appears to be involved in
nicotine-induced locomotor activity (Richardson and Tizabi,
1994), as nicotine has been shown to release DA in the
striatum (Damsma et al., 1988). Differences in the effects of
ABBREVIATIONS: DA, dopamine; DHbE, dihydro-b-erythroidine; NAChR, nicotinic acetylcholine receptor(s); NE, norepinephrine; SIB-1765F
([6]-5-ethynyl-3-(1-methyl-2-pyrrolidinyl)pyridine fumarate).
384
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ABSTRACT
SIB-1765F
([6]-5-ethynyl-3-(1-methyl-2-pyrrolidinyl)pyridine
fumarate) is a novel nicotinic acetylcholine receptor (NAChR)
agonist displaying a different in vitro pharmacological profile
than nicotine and epibatidine, suggestive of NAChR subtype
selectivity. Our study describes the effects of SIB-1765F on
locomotor activity in rats, which were compared to those observed for nicotine and epibatidine. The three NAChR agonists
decreased or increased locomotor activity in rats naive or habituated to the test apparatus, respectively. The transient reduction in locomotor activity induced by SIB-1765F was quantitatively similar to those induced by nicotine and epibatidine
but, unlike the effects of nicotine and epibatidine, was not
blocked by the NAChR antagonists mecamylamine and dihydro-b-erythroidine, suggesting different mechanisms of action.
1997
NAChR Agonists and Locomotor Activity
Materials and Methods
Animals. Male Sprague-Dawley rats (200–350 g) (Harlan, San
Diego, CA) were housed four per cage and maintained in a humidity(50–55%) and temperature- (22–24°C) controlled facility on a 12
hr:12 hr light/dark cycle (lights on at 6:30 A.M.) with free access to
food (Harlan-Teklad 4% rat diet 7001) and water. Rats were allowed
a 1-wk period of habituation to the animal room before testing. The
animals were handled once during this period.
Compounds. Mecamylamine hydrochloride, (-)-nicotine hydrogen
tartrate and hexamethonium bromide were obtained from Sigma
Chemical Co. (St. Louis, MO). DHbE, (6)-epibatidine di-hydrochloride, S-(-)-eticlopride hydrochloride and R(1)-SCH 23390 hydrochloride were purchased from Research Biochemicals International
(RBI, Natick, MA). SIB-1765F was synthesized at SIBIA as per
methods previously described (Cosford et al., 1996). Nicotine and
SIB-1765F were dissolved in saline and pH was adjusted to 7.0 by
addition of 10N NaOH. DHbE, epibatidine, eticlopride,
mecamylamine, hexamethonium and SCH 23390 were dissolved in
saline. Doses of epibatidine and nicotine are expressed in terms of
their free base concentrations. All compounds were administered s.c.
into the dorsal neck region in a volume of 1 ml/kg. Saline was used
as control.
Locomotor activity. Locomotor activity was assessed in photocell activity cages (San Diego Instruments, San Diego, CA). Each
cage consisted of a standard plastic rodent cage (24 3 45.5 cm)
surrounded by a stainless steel frame. Four infrared photocell beams
were located across the long axis of the frame, raised 3.2 cm above
the floor and spaced 9 cm apart. The number of cage crosses
(crossovers, i.e., consecutive interruptions of one beam followed immediately by interruption of an adjacent beam) was recorded and
used as a measure of locomotion. The number of crossovers was
recorded by a computer system during consecutive 5-min intervals.
Decreases in locomotor activity were evaluated in rats naive to the
test apparatus (hereafter referred to as naive rats). Rats received
injections with test compound or vehicle and were placed in the
photocell activity cages 5 min after injection. Locomotor activity was
recorded for a 30-min period.
Increases in locomotor activity were evaluated in rats previously
habituated to the test apparatus (hereafter referred to as habituated
rats). Rats were habituated to the photocell activity cages for 180
min, 24 hr before testing. This habituation period was required to
overcome the potentially stressful nature of the test apparatus and,
therefore, reduce the basal levels of activity of the rats. On the test
day, the rats were placed in the photocell activity cages for a habituation period of 90 min, after which they were removed, injected
with the test compound and were returned to the cages to be monitored for 120 to 360 min.
In studies evaluating the effects of NAChR antagonists on NAChR
agonists-induced decreases in locomotor activity, naive rats were
administered with antagonists 15 min before the injection of NAChR
agonists. After administration of NAChR agonists, locomotor activity
was recorded for 30 min.
In studies evaluating the effects of antagonists on NAChR agonists-induced increases in locomotor activity, habituated rats were
administered with nicotinic or dopaminergic receptor antagonists 15
or 30 min before the injection of NAChR agonists, respectively. After
administration of NAChR agonists, activity was recorded for 90 min.
Nicotinic and dopaminergic receptor antagonists were also tested
alone in a novel environment to determine potential intrinsic sedative or depressant effects. Naive rats received injections in their
home cages and were placed in the photocell activity cages 15 min
after injection of NAChR antagonists and 30 min after injection of
DA receptor antagonists. Locomotor activity was recorded for 30
min.
These studies were based on independent subject design, with
each subject used only once. Twelve rats (individually caged) were
tested at one time. Testing was carried out between 8:00 A.M. and
5:30 P.M. each day (light cycle).
Statistics. Results were analyzed by Student’s t test or by oneand two-factor analyses of variance, with repeated measure methods
where appropriate (SigmaStat Software, Jandel, San Rafael, CA).
Dunnett’s test and Newman-Keul’s test were used for post hoc comparisons. P , .05 was considered significant.
Results
Effects of NAChR agonists on locomotor activity in
naive rats. When administered to naive rats, the three
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NAChR agonists on locomotor activity may also be related to
the presence of various NAChR subtypes in the central nervous system. Molecular and electrophysiological studies suggest that NAChR subtypes are structurally and functionally
diverse (for review, see Patrick and Luetje, 1993). Thus,
differences in locomotor effects may reflect different actions
of NAChR agonists at distinct subpopulations of NAChR or
differential activities at the same receptor(s) (i.e., partial
agonist vs full agonist). The unraveling of the pharmacology
of NAChR has been hampered by the limited number of
subtype-selective NAChR agonists and antagonists.
SIB-1765F is a novel NAChR agonist with an in vitro
pharmacological profile suggestive of NAChR subtype selectivity (Lloyd et al., 1995, Sacaan et al., accompanying manuscript). SIB-1765F displaces [3H]-cytisine binding to rat cortical membrane with high affinity but has a lower affinity
than nicotine at muscarinic cholinergic and a-bungarotoxin
binding sites. SIB-1765F is moderately more effective than
nicotine at releasing DA from rat striatal and olfactory tubercle slices in vitro (Rao et al., 1995; Sacaan et al., accompanying manuscript) and is as effective as nicotine at releasing [3H]-NE from rat thalamic and cortical slices. However,
unlike nicotine, SIB-1765F only slightly stimulates [3H]-NE
release from rat hipoccampal slices. Because the release of
striatal DA and hippocampal NE appears to be regulated by
distinct NAChR (Sacaan et al., 1995; Clarke and Reuben,
1996), differential effects of SIB-1765F on striatal DA and
hippocampal NE release suggest that SIB-1765F may preferentially activate specific neuronal NAChR subtypes as
compared to nicotine (Lloyd et al., 1995; Sacaan et al., accompanying manuscript).
Our experiments have been carried out to determine
whether the different pharmacological profile of SIB-1765F,
as compared to nicotine, translates into a different locomotor
profile. SIB-1765F was also compared to epibatidine, a potent
NAChR agonist that has recently been demonstrated to increase locomotor activity in rats (Sacaan et al., 1996). The
three NAChR agonists were administered to rats that were
naive or habituated to the test environment to compare the
locomotor effects of each compound. In addition, the mechanisms by which these NAChR agonists produced their effects
on locomotor activity were assessed using the peripherally
active NAChR antagonist hexamethonium, the centrally active noncompetitive NAChR antagonist mecamylamine
(Martin et al., 1993; Varanda et al., 1985) and the centrally
active competitive NAChR antagonist DHbE (Williams and
Robinson, 1984; Valera et al., 1992). Finally, the role of the
dopaminergic system in the stimulant effects induced by
nicotine and SIB-1765F was investigated using the D1 and
D2 dopamine receptor antagonists SCH 23390 and eticlopride, respectively.
385
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Menzaghi et al.
ity was not significantly different from the activity of vehicletreated animals.
Nicotine and epibatidine produced similar transient decreases in locomotor activity followed by mild increases in
locomotor activity [nicotine 3 time interaction: F(5, 50) 5
11.62, P , .0001; epibatidine 3 time interaction: F(5, 50) 5
39.42, P , .0001] (fig. 1, B-C, right panels). The magnitudes
of the depressant effects were similar for the three NAChR
agonists.
Effects of NAChR antagonists on NAChR agonistsinduced decreases in locomotor activity in naive rats.
Figure 2 shows the effects of mecamylamine, hexamethonium and DHbE on the decrease in locomotor activity induced by a single dose of each NAChR agonist in naive rats.
The NAChR antagonists had no significant effect on locomotor activity when administered alone (fig. 2A-B). Mecamylamine (3 mg/kg) blocked the effects induced by nicotine (0.4
mg/kg) [interaction nicotine 3 mecamylamine: F(1, 20) 5
8.10, P , .009] and epibatidine (3 mg/kg) [interaction epiba-
Fig. 1. Decreases in locomotor activity induced by SIB1765F (A), nicotine (B) and epibatidine (C) in rats not
previously exposed to the test
apparatus (naive rats). Left
panel, Dose-related effects
presented as total crossovers
over a 10-min period beginning 5 min after administration
of the compounds (mean 6
S.E.M., n 5 6/group). *P , .05
vs vehicle-treated group, Dunnett’s test. Right panel, Time
course of effects presented as
crossovers taken at 5-min intervals beginning 5 min after
the administration of the compounds (n 5 6/group). Each
point represents mean 6
S.E.M. *P , .05 vs vehicletreated group, Newman-Keul’s
test.
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NAChR agonists, SIB-1765F, nicotine and epibatidine, decreased locomotor activity as illustrated by a decrease in the
number of crossovers (fig. 1, A-C, left panels). SIB-1765F and
epibatidine reduced locomotor activity in a dose-related manner [SIB-1765F: F(4, 25) 5 5.53, P 5 .0025; epibatidine: F(4,
25) 5 18.0, P , .0001] whereas nicotine decreased locomotor
activity in a non-dose-related manner during the first 10 min
of the session [F(4, 25) 5 6.60, P 5 .0009] (fig. 1).
Figure 1 (right panels) also shows the time course for the
effect of each NAChR agonist over the entire session (30
min). For clarity, only one dose of each compound is shown
and compared to its respective vehicle. There was a significant decline in activity for each vehicle-treated group as the
animals became habituated to the test environment. SIB1765F at a dose of 20 mg/kg produced a depressant effect that
decreased over time [SIB-1765F 3 time interaction: F(5, 50)
5 16. 78, P , .0001] (fig. 1A, right panel). This transient
decrease in locomotor activity was followed by a small increase in locomotor activity. However, this increase in activ-
Vol. 280
1997
387
tidine 3 mecamylamine: F(1,20) 5 17.3, P , .0005] but did
not attenuate the effect induced by SIB-1765F (20 mg/kg)
[interaction SIB-1765F 3 mecamylamine: F(1,20) 5 .14, P 5
.71] (fig. 2A). Similarly, DHbE (6 mg/kg) blocked the locomotor depressant effects induced by nicotine and epibatidine
but failed to attenuate the locomotor depressant effect induced by SIB-1765F (fig. 2B). Hexamethonium (10 mg/kg)
had no effect on the decrease in locomotor activity induced by
nicotine, epibatidine or SIB-1765F (fig. 2A).
Analysis of the time courses of the effects of mecamylamine
and DHbE over a period of 30 min revealed that these centrally acting NAChR antagonists failed to block the decrease
in locomotor activity induced by the administration of a 20mg/kg dose of SIB-1765F, but did block the mild increase in
locomotor activity that followed (fig. 3, A-B, left panels).
Under identical conditions, the decreases in locomotor activity induced by nicotine and epibatidine were both attenuated
by mecamylamine and DHbE and were insensitive to hexamethonium (fig. 3, A-B, center and right panels).
Effects of NAChR agonists on locomotor activity in
habituated rats. Figure 4 shows the effects of the three
NAChR agonists after administration to rats habituated to
the photocell activity cages. There was a significant difference between the levels of activity of vehicle-treated naive
and habituated rats over the first 30 min of exposure to the
cages (compare fig. 1A vs fig. 4A: right panels). Habituated
rats had a statistically significant lower level of activity than
naive rats (total crossovers/30 min: naive rats: 86.2 6 9.4,
habituated rats: 31.5 6 5.7; P , .05 Student’s t test). In
habituated rats, s.c. administration of SIB-1765F increased
the number of crossovers in a dose-related manner [F(5, 41)
5 20.4, P , .0001], with the maximum effect occurring with
a dose of 40 mg/kg (P , .05) (fig. 4A, left panel). Locomotor
Fig. 3. Time courses of the antagonism by mecamylamine (3 mg/kg) and hexamethonium (10 mg/kg) (A) and DHbE (6 mg/kg) (B) of the locomotor
effects induced by NAChR agonists in naive rats. Data are presented as average crossovers taken at 5-min intervals beginning 5 min after the
administration of SIB-1765F (20 mg/kg), nicotine (0.4 mg/kg) and epibatidine (3 mg/kg) (n 5 6/group). Doses used were chosen on the basis of
equal efficacy. S.E.M. were not shown for the clarity of the graphs. *P , .05 vs vehicle/vehicle-treated groups, #P , .05 vs vehicle/NAChR agonist,
Newman Keul’s test.
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Fig. 2. Effects of mecamylamine (3 mg/kg) (A), hexamethonium (10
mg/kg) (A) and DHbE (6 mg/kg) (B) on NAChR-agonists induced decreases in locomotor activity in naive rats. The NAChR antagonists
were administered 15 min prior to SIB-1765F (20 mg/kg), nicotine (0.4
mg/kg) and epibatidine (3 mg/kg). Data are represented as mean total
crossovers over a 10-min period beginning 5 min after the injection of
the agonists (mean 6 S.E.M., n 5 6). #P , .05 vs vehicle/vehicletreated group, *P , .05 vs vehicle/NAChR agonist-treated group, Newman-Keul’s test.
NAChR Agonists and Locomotor Activity
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Menzaghi et al.
Vol. 280
activity after administration of the 40 mg/kg dose of SIB1765F was nearly 15 times greater than that after administration of vehicle, over the 120-min period.
Nicotine also increased locomotor activity in a dose-dependent manner [F(5,40) 5 4.28, P , .003] but the maximum
effect was never more than 3.5 times that of vehicle (fig. 4B,
left panel). Epibatidine slightly increased locomotor activity
[F(5,40) 5 3.58, P , .009], but the total number of crossovers
produced by the most effective dose (3 mg/kg) was less than
twice that of vehicle (fig. 4C, left panel).
Figure 4 also shows the time courses for the effects of
the NAChR agonists on locomotor activity. A decline in crossovers for both vehicle- and drug-treated rats was observed as
the session proceeded. In the study of SIB-1765F (20 mg/kg),
there was a main effect of the compound [F(1, 14) 5 281.5,
P , .0001], a main effect of time [F(23, 322) 5 2.98, P ,
.0001] and a significant interaction between the two factors
[F(23, 322) 5 2.14, P 5 .0021] (fig. 4, A-C, right panels). The
increase in activity induced by SIB-1765F at doses of 20 and
40 mg/kg was observed within 5 to 15 min and lasted for 2.2
and 3.5 hr, respectively (fig. 5). The increase in locomotor
activity induced by nicotine at its maximally effective dose
(0.4 mg/kg) was observed within 5 min and lasted no more
than 80 min after administration [nicotine effect: F(1, 14) 5
20.42, P , .0006; time effect: F(23, 299) 5 8.23, P , .0001;
and no interaction nicotine 3 time: F(23, 299) 5 1.11, P 5
.33] (fig. 4B, right panel). Epibatidine, at a dose of 3 mg/kg,
increased locomotor activity within 20 min post-injection and
the effect lasted for less than 50 min [epibatidine effect: F(1,
15) 5 5.40, P 5 .03; time effect: F(23, 345) 5 5.58, P , .0001;
epibatidine 3 time interaction: F(23, 345) 5 1.66, P 5 .03]
(fig. 4C, right panel).
Effects of NAChR antagonists on NAChR agonistsinduced increases in locomotor activity in habituated
rats. Pretreatment with mecamylamine (3 mg/kg) or DHbE
(6 mg/kg) significantly attenuated the locomotor activity in-
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Fig. 4. Increases in locomotor activity induced by SIB1765F (A), nicotine (B) and epibatidine (C) in rats habituated
to the test cages. Left panel,
Dose-related effects presented as total crossovers
over a 120-min period beginning immediately after the administration of the compounds
(mean 6 S.E.M., n 5 7– 8/
group). *P , .05 vs vehicletreated group, Dunnett’s test.
Right panel, Time courses of
effects presented as crossovers taken at 5-min intervals
beginning immediately after
the administration of the compounds (n 5 7– 8/group). Each
point represents mean 6
S.E.M. The indications of statistical significance have been
omitted to enhance visual clarity.
1997
duced by SIB-1765F (20 mg/kg) [mecamylamine 3 SIB1765F interaction: F(1,20) 5 9.85, P 5 .005; DHbE 3 SIB1765F interaction: F(1,20) 5 6.63, P 5 .01] (fig. 6A, left and
right panels). In contrast, hexamethonium (10 mg/kg) had no
effect on locomotor activity induced by SIB-1765F [SIB-
389
1765F 3 hexamethonium interaction: F(1,20) 5 .46, P 5 .50]
(fig. 6A, center panel). The locomotor activity induced by
nicotine (0.4 mg/kg) was also blocked by mecamylamine and
DHbE as shown by a significant agonist and antagonist
interaction [nicotine 3 mecamylamine: F(1,28) 5 20.6, P ,
.0001 nicotine 3 DHbE: F(1,20) 5 6.34, P 5 .02] and was
insensitive to hexamethonium [nicotine 3 hexamethonium:
F(1,27) 5 1.69, P 5 .20] (fig. 6B). Epibatidine- (3 mg/kg)
induced locomotor activity was blocked by mecamylamine
[epibatidine 3 mecamylamine: F(1,20) 5 36.0, P , .0001] but
was insensitive to DHbE [epibatidine 3 DHbE: F(1,34) 5 .04,
P 5 .85] or hexamethonium [epibatidine 3 hexamethonium:
F(1,20) 5 2.81, P 5 .11] (fig. 6C).
Effect of selective D1 and D2 DA receptor antagonists on NAChR agonists-induced locomotor activity
in habituated rats. To determine if the locomotor activity
induced by SIB-1765F and nicotine were mediated through
the activation of the dopaminergic system, the selective D1
receptor antagonist SCH 23390 and the selective D2 receptor
antagonist eticlopride were administered before the test compounds.
SCH 23390 reduced SIB-1765F-induced locomotor activity
in a dose-dependent manner [SCH 23390 3 SIB-1765F in-
Fig. 6. Effects of mecamylamine, hexamethonium and DHbE on the locomotor activity induced by SIB-1765F (20 mg/kg) (A), nicotine (0.4 mg/kg)
(B) and epibatidine (3 mg/kg) (C) in rats habituated to the test cages. The antagonists were administered 15 min before the NAChR agonists. Data
are represented as mean total counts taken over a 90-min period after the injection of vehicle (white bar) or NAChR agonists (gray bar) (mean 6
S.E.M., n 5 6 –9). #P , .05 vs vehicle/vehicle-treated group, *P , .05 vs vehicle/NAChR agonist-treated group, Newman-Keul’s test.
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Fig. 5. Duration of the locomotor activity induced by SIB-1765F in
rats habituated to the test cages. Data are presented as average of
crossovers taken at 5-min intervals beginning immediately after the
administration of the compound (n 5 8/group). S.E.M. were not shown
for the clarity of the graph.
NAChR Agonists and Locomotor Activity
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Discussion
Depressant or stimulant locomotor effects induced by
NAChR agonists can be revealed by exposing rats to a novel
or a familiar testing environment. Acute s.c. administration
of nicotine, epibatidine or the novel NAChR agonist SIB1765F decreased locomotor activity in experimentally naive
rats and increased locomotor activity in rats habituated to
the test apparatus. The decreases in locomotor activity induced by these three NAChR agonists were quantitatively
similar. In contrast, the three NAChR agonists differ considerably in their stimulatory effects, with SIB-1765F producing
a greater increase in locomotor activity than nicotine or epibatidine.
The decreases in locomotor activity observed after administration of SIB-1765F, nicotine and epibatidine in experimentally naive rats are similar to those observed previously
with other NAChR agonists (Reavill et al., 1990; Garcha et
al., 1993; Stolerman et al., 1995). These effects have a short
duration (,15 min).
The noncompetitive NAChR antagonist mecamylamine
(Varanda et al., 1985; Martin et al., 1993) and the competitive
NAChR antagonist DHbE (Reavill et al., 1988, Williams and
Robinson, 1984; Valera et al., 1992, Damaj et al., 1995) completely reversed the locomotor depressant effects induced by
nicotine and epibatidine, implying that these effects were
mediated through the activation of NAChR. The peripherally
active NAChR antagonist hexamethonium failed to block the
decrease in locomotor activity induced by nicotine or epibatidine. Thus, the decreases in locomotor activity induced by
nicotine and epibatidine appear to be mediated through the
activation of central and not peripheral NAChR. Interestingly, hexamethonium, mecamylamine and DHbE failed to
block the decrease in locomotor activity produced by SIB1765F in naive rats. This suggests that SIB-1765F acts on
NAChR subtype(s) insensitive to mecamylamine and DHbE
or reduces locomotor activity by a nonnicotinic mechanism.
Mecamylamine also failed to block the decrease in locomotor
activity induced by the NAChR agonists lobeline and ana-
Fig. 7. Effects of SCH 23390
and eticlopride on the locomotor activity induced by SIB1765F (20 mg/kg) (A) and nicotine (0.4 mg/kg) (B) in rats
habituated to the test cages.
SCH 23390 and eticlopride
were administered 30 min before the NAChR agonists.
Data are presented as crossovers taken over a 90-min period after the injection of saline
(white bar) or NAChR agonists
(gray bar) (mean 6 S.E.M., n 5
6). #P , .05 vs vehicle/vehicletreated group, *P , .05 vs
vehicle/NAChR agonist-treated
group, Newman-Keul’s test.
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teraction: F(3,40) 5 13.7, P , .0001] (fig. 7A, left panel). SCH
23390 also attenuated the locomotor activity induced by nicotine [SCH 23390 3 nicotine interaction: F(3,40) 5 4.14, P 5
.01] (fig. 7B, left panel). SCH 23390 did not have a significant
intrinsic effect on activity when administered to habituated
rats; however, it reduced spontaneous locomotor activity in
experimentally naive rats in a dose-dependent manner [SCH
23390: F(3,20) 5 9.82, P 5 .0003] (fig. 8A, left panel). Analysis of the time course showed a main effect of SCH 23390
[F(3,20) 5 6.36, P , .003], a main effect of time [F(5,143) 5
47.93, P , .0001] and a significant interaction between the two
factors [F(15,143) 5 5.66, P , .0001] (fig. 8A, right panel).
The selective D2 receptor antagonist eticlopride also produced a significant reduction in the effect of SIB-1765F on
locomotor activity [eticlopride 3 SIB-1765F interaction:
F(3,40) 5 14.9, P , .0001] (fig. 7A, right panel). Analysis of
the time course showed a main effect of eticlopride [F(3,20) 5
12.63, P , .0001], a main effect of time [F(5,143) alone 5
112.68, P , .0001], and a significant interaction between the
two factors [F(15,143) 5 2.95, P , .0007] (fig. 8B, right
panel). Low doses of eticlopride (5–10 mg/kg) that had no
significant effect on spontaneous activity when tested in naive rats (fig. 8B) did attenuate SIB-1765F-induced locomotor
activity (fig. 7A: right panel). Eticlopride at a 20-mg/kg dose
also attenuated the effect of nicotine on locomotor activity
although there was no significant interaction between the
drugs [eticlopride effect: F(3,40) 5 2.79, P 5 .05; nicotine
effect: F(1,40) 5 23.76, P , .0001; eticlopride 3 nicotine
interaction: F(3,40) 5 2.07, P 5 .12] (fig. 7B, right panel).
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1997
NAChR Agonists and Locomotor Activity
391
toxin in naive rats (Stolerman et al., 1992, 1995). Therefore,
the depressant effect induced by SIB-1765F in experimentally naive rats is similar in magnitude to those induced by
nicotine and epibatidine, but appears to be mediated through
a different mechanism.
The locomotor stimulatory effects induced by SIB-1765F
and nicotine in habituated rats were blocked by
mecamylamine and DHbE but not by hexamethonium, indicating that these NAChR agonists elicit their stimulant effects predominantly through the activation of central
NAChR. This contrasts with the lack of effect of
mecamylamine and DHbE on SIB-1765F-induced decrease in
locomotor activity in experimentally naive rats, and suggests
that the stimulant and depressant effects of SIB-1765F are
likely to be mediated through different mechanisms. Moreover, epibatidine-induced increase in locomotor activity was
attenuated by mecamylamine but not by DHbE whereas
epibatidine-induced decrease in locomotor activity was attenuated by both centrally active NAChR antagonists. These
data suggest that the depressant and stimulant effects of
epibatidine are also likely to be mediated through different
mechanisms.
NAChR agonists are known to increase locomotor activity
through an interaction with the dopaminergic system
(Reavill and Stolerman, 1990; Museo and Wise, 1990; Imperato et al., 1986). Therefore, the effect of DA antagonists on
the increases in locomotor activity induced by nicotine and
SIB-1765F were evaluated. SCH 23390, a selective D1 DA
receptor antagonist, produced a dose-dependent decrease in
spontaneous locomotor activity in naive animals, but did not
affect locomotor activity in habituated animals. These results
contradict a previous report from O’Neill et al. (1991) showing that SCH 23390 at doses five times higher than those
used in our study failed to attenuate spontaneous locomotor
activity in naive rats. In habituated rats, SCH 23390 attenuated SIB-1765F induced locomotor activity at doses that did
not alter spontaneous locomotor activity in naive rats, suggesting that D1 receptor activation may in part contribute to
the stimulant effect of SIB-1765F. Previously, SCH 23390
was shown to reduce the locomotor stimulant effects induced
by nicotine and epibatidine (O’Neill et al., 1991; Sacaan et al.,
1996). The D2 DA receptor antagonist eticlopride also attenuated the increase in locomotor activity induced by SIB1765F at doses (5 and 10 mg/kg) that did not significantly
reduce spontaneous activity in naive rats. Nicotine-induced
locomotor activity was blocked by a 20-mg/kg dose of eticlopride that also attenuated spontaneous activity in naive rats.
Nicotine-induced locomotor activity is known to be sensitive
to other D2 antagonists such as raclopride (O’Neill et al.,
1991) and haloperidol (Arnold et al., 1995). We recently reported that the stimulant effect of epibatidine was also attenuated by eticlopride (Sacaan et al., 1996). These data
provide additional evidence that the acute increases in locomotor activity induced by NAChR agonists are mediated, at
least in part, through the activation of both D1 and D2 DA
receptors subsequent to the release of DA.
SIB-1765F had substantial effects on locomotor activity in
habituated rats as compared to nicotine or epibatidine. Over
a period of 90 min, a maximally effective dose of SIB-1765F
was approximately four times more efficacious and lasted
two to three times longer than nicotine or epibatidine in
increasing locomotor activity. In vitro studies have shown
that SIB-1765F releases [3H]-DA from rat striatal and olfactory tubercle slices, regions containing terminal fields of the
nigrostriatal and mesolimbic dopaminergic pathways, respectively (Rao et al., 1995; Sacaan et al., accompanying
manuscript). In these assays, SIB-1765F tends to be more
effective than nicotine at releasing [3H]-DA. Microdialysis
Downloaded from jpet.aspetjournals.org at ASPET Journals on June 16, 2017
Fig. 8. Effects of SCH 23390 (A) and eticlopride (B) on locomotor activity of naive rats. Left panel, Dose-related effects
presented as total crossovers taken over
a 10-min period beginning 30 min after
the administration of the compounds
(mean 6 S.E.M., n 5 6/group). *P , .05
vs vehicle-treated group, Dunnett’s test.
Right panel, Time course of effects presented as average crossovers taken at
5-min intervals beginning 30 min after
the administration of the compounds
(n 5 6/group). S.E.M. were not shown for
the clarity of the graph. *P , .05 vs vehicle-treated group.
392
Menzaghi et al.
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Downloaded from jpet.aspetjournals.org at ASPET Journals on June 16, 2017
studies also indicate that SIB-1765F is considerably more
efficacious than nicotine at releasing DA from both rat striatum and nucleus accumbens in vivo (Menzaghi et al., 1995;
Sacaan et al., accompanying manuscript). Furthermore, SIB1765F is more efficacious than nicotine and epibatidine at
inducing ipsilateral turning in unilaterally 6-OHDA-lesioned
rats (Lloyd et al., 1995; Cosford et al., 1996). Thus, the
greater effect of SIB-1765F on DA release in vivo, as compared to nicotine, may be, at least in part, responsible for a
greater increase in locomotor activity. In addition, activation
of different NAChR subtypes and differences in pharmacokinetics may also contribute to these differences in profiles.
In summary, these results demonstrate that the novel
NAChR agonist SIB-1765F both increased and decreased
spontaneous locomotor activity in rats depending on the experimental paradigm. These effects can be demonstrated by
testing animals in a novel or a familiar environment. Similar
effects were seen with the NAChR agonists nicotine and
epibatidine. Interestingly, the decreases in locomotor activity
induced by these three nicotinic agonists were similar in
magnitude but appeared to be mediated through different
mechanisms. Furthermore, our data suggest that the stimulant and depressant effects induced by SIB-1765F or epibatidine are likely to be mediated through different mechanisms. SIB-1765F produced a larger and longer-lasting
increase in locomotor activity. This effect is dependent on D1
and D2 DA receptor activation subsequent to DA release.
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