This document contains text automatically extracted from a PDF or image file.

Formatting may have

been lost and not all text may have been recognized.
To remove this note, right-click and select "Delete table".
Pharmacological Research, Vol. 41, No. 1, 2000
Article No. phrs.1999.0560, available online at http:rrwww.idealibrary.com on
THE CANNABINOID AGONIST HU 210 MODIFIES RAT BEHAVIOURAL
RESPONSES TO NOVELTY AND STRESS
D. GIULIANI, F. FERRARIU and A. OTTANI
Department of Biomedical Sciences, Di¨ision of Pharmacology, Uni¨ersity of Modena,
¨ia G. Campi 287, I-41100, Modena, Italy
Accepted 7 June 1999
Experiments were performed on groups of rats after acute and sub-chronic treatment Ž once
daily for 9 days . with the cannabinoid agonist HU 210 Ž 25 100 g kg y1 , i.p. . as well as 24 h
and 7 days after the last drug injection. The animals underwent three behavioural tests in
novel environments. In the observation cages Ž Test 1 . , rat locomotor activity was found to
be dose-dependently reduced after acute and sub-chronic treatment at all doses and
virtually unchanged during abstinence; grooming was potently inhibited by acute treatment
but potentiated by the sub-chronic one at doses of 50 and 100 g kgy1, the effect of the
higher dose persisting after 24 h and 7 days abstinence. Vocalization in animals in response
to a tactile stimulus was highest after HU 210 at 100 g kgy1 in all experimental modes
except after 7 days abstinence. In the X-maze Ž Test 2 . , sub-chronic HU 210 dose-
dependently enhanced rat natural aversion for open arms, and this behaviour persisted
during abstinence after the highest dose. Grooming in the X-maze was completely absent in
rats acutely injected with HU 210 but potentiated in those sub-chronically treated or
abstinent. In the swimming test Ž Test 3 . rats sub-chronically treated at 50 and 100 pg kg
y1 displayed relevant wall-hugging and the same occurred 24 h after last injection. On the
whole, our results are indicative of an anxiogenic-like effect of sub-chronic HU 210 at high
doses and reflect the persistence of enhanced emotional response to novel environments
when the treatment is discontinued. 2000 Academic Press
K
EY WORDS
: cannabinoids, HU 210, anxiety-like state, rat.
INTRODUCTION
It is well established that the psychotropic effects of
9 -tetrahydrocannabinol Ž 9 -THC . , the major active
component of marijuana w 1 x , are linked to changes in
the activity of central neurotransmitter systems w 2 4 x .
The discovery of specific central cannabinoid bind-
ing sites Ž CB1 .w 5 8 x which seem to be primarily
involved in many of the behavioural and physiologi-
cal alterations produced in humans and animals by
9 -THC w 9 13 x , has led to the synthesis of a number
of new cannabinoids w 10, 12 x . Of those, HU 210 has
demonstrated a high degree of correlation between
its producing ability to in bind ¨i¨o to effects CB 1
receptors w 13 and its efficacy in
16 x . Several clinical
and experimental findings have promoted the pre-
sent study on HU 210. It has been reported that
anxiogenic reactions are elicited by marijuana or
U
Corresponding author.
9-THC in humans, particularly after high doses and
long-term exposure w 17 19 x ; similarly, an anxiety-like
state has been found in cannabinoid-treated rodents
subjected to different behavioural models w 16, 20,
21 x . Again, it has been demonstrated that 9 -THC
whether acutely or sub-chronically administered, po-
tently stimulates adrenocorticotropin hormone
Ž ACTH . secretion w 22 x , and that central corti-
cotropin-releasing factor Ž CRF . systems play a key
role in the mediation of cannabinoid-induced anxio-
genic effects w 16 x . ACTH and CRF vigorously en-
hance self-grooming in the rat w 23, 24 x and this
seems to represent a response to a state akin to
psychological human stress, since it manifests itself
as stereotyped behaviour in different stressful situa-
tions.
There appear to be few reports on the influence
of cannabinoids on grooming. Some researchers have
observed no significant modification w 25 x , while oth-
ers have demonstrated inhibition induced by acute
HU 210 w 15 x or 9 -THC w 26 x .
1043 6618r00r010047 07r$35.00r0 2000 Academic Press
Pharmacological 48
Research, Vol. 41, No. 1, 2000 The aim of our work was, therefore, to investigate
in more detail the influence exerted by HU 210 on
rat grooming and on general animals’ behaviour in
novel anxiogenic environments w 27 29 x . Rats’ be-
havioural patterns were recorded after acute and
sub-chronic treatment with the drug at different
doses, as well as during abstinence. In addition,
other parameters that fulfil the criteria for evaluat-
ing cannabimimetic activity, namely, locomotor ac-
tivity w 10 x and vocalization w 30 x , were evaluated in
some experimental conditions.
MATERIALS AND METHODS
Animals and general beha¨ioural procedure
The subjects were male SPF-Wistar rats Ž Harlan
Nossan, Udine, Italy . weighing 200 230 g at the
outset. They were housed in groups of four with
food and water ad libitum and on a 12-h light cycle,
from 07:00 to 19:00 h, for at least 1 week prior to the
start of the experiments. The experiments were per-
formed between 09:00 and 14:00 h in a soundproof,
air-conditioned room, with normal lighting condi-
tions, where the animals were monitored by trained
observers unaware of the treatment schedule. The
controls were handled in the same way as the treated
animals and received vehicle injections. Experimen-
tally-naive rats were randomly assigned to various
groups that were submitted to three tests in three
experimental conditions, namely, after acute treat-
ment with HU 210 Ž 25, 50 or 100 g kg y1 , i.p. . ,
sub-chronic treatment Ž once daily for 9 days at the
same doses . and 24 h after the last injection Ž absti-
nence . . Each rat performed one test only and was
used only in one experimental condition, thus avoid-
ing the risk that memory of the environment might
influence its behaviour. Having observed that, after
24 h abstinence, the rats injected with 100 g kgy1
behaved in a significantly different manner from the
controls in all tests, other naive animals were sub-
chronically treated at this dose and, 7 days after the
last injection, underwent Tests 1 and 2 Ž see below . .
The in-force regulations on the care of animals for
scientific purposes Ž CEE Concil 86r603; Italian D.L.
27-01-92 N 116 . were strictly complied with.
Test 1: locomotor acti¨ity, grooming and ¨ocaliza-
tion in the obser¨ation cages Behavioural evalua-
tions were carried out on groups of animals Ž homo-
geneous as regards treatment . which were trans-
ferred to glass observation cages Ž 40=30=34 cm
. 50 min after the i.p. injection of HU 210 or vehicle
Ž acute and sub-chronic . , as well as during absti-
nence. The test Ž 30 min . started immediately after
the animals had been placed in the observation
cages and the parameters considered were locomo-
tor activity, grooming and vocalization. Locomotor
acti¨ity was scored as described elsewhere w 31 x , each
rat being observed for 30 s at 5-min intervals and
rated on a scale 0 2 where: 0sabsent, 1s
discontinuous locomotor activity Ž exploratory be-
haviour, rearing and lateral movements . , 2 s
uninterrupted locomotor activity for at least 25 s.
This evaluation allowed the contemporaneous as-
sessment of grooming. Grooming was evaluated ac-
cording to Gispen et al. w 23 x . In brief, an observer
recorded at 15-s intervals whether or not each rat
displayed the phenomenon, defined as face and body
washing, scratching, licking paws or tail. If one of
these signs was observed a positive score was given.
Locomotor activity and grooming values for each
rat are the sum of all the scores attributed to the
animal for each parameter during the test period.
Immediately after their removal from the cages, the
same animals were tested for the presence or
absence of ¨ocalization. For this purpose, each rat
was gently pressed two to four times bilaterally
behind its forelimb, on the ventral aspect of the
frontal costal region, the experimenter using thumb
and forefinger w 30 x .
Test 2: X-maze A test was performed on animals
which were placed singly into the X-maze, 60 min
after the i.p. injection of HU 210 or vehicle Ž acute
and sub-chronic . , as well as during abstinence. The
apparatus was as previously described w 32 x . In brief,
it consists of a black Plexiglas floor in the form of a
regular cross, the arms of which are flanked by
transparent Plexiglas walls and meet at the intersec-
tion to form a central arena; one opposite pair of
arms is covered, while the other is left open. Each
rat was gently put into the central arena of the
apparatus, facing an open arm, and its subsequent
exploratory behaviour was timed over a 5-min pe-
riod. A rat was taken to have entered an arm when
all four paws were on it. The number of open- and
closed-arm entries, the time spent in each arm and
the number of total entries were recorded. Total
grooming displayed during the test was counted with
a stopwatch. At the end of each test, the animal was
removed and the maze floor was thoroughly cleaned.
Test 3: swimming-test This test was performed on
animals which were placed singly into a swimming-
pool, 60 min after the i.p. injection of HU 210 or
vehicle Ž acute and sub-chronic . , as well as during
abstinence. Swimming strategy was observed for 2
min in a circular tank Ž 130 cm in diameter, 60 cm in
depth . , constructed of white Plexiglas that, prior to
testing, had been filled to a depth of 45 cm with
water maintained at 30"2 C. The experimenters
recorded the episodes of wall-hugging, defined as a
complete turn of the pool in contact with the wall
w 29 x .
Drugs and treatments
HU 210 Ž Tocris-Cookson, Bristol, UK . was freshly
prepared as a suspension containing a drop of Tween
Pharmacological Research, Vol. 41, No. 1, 2000 49
80 Ž 0.1% . and distilled water at concentrations that
allowed the injection of 1 ml kgy1 intraperitoneally
Ž i.p. . . The doses of HU 210 and the pretreatment
time were chosen on the basis of preliminary experi-
ments w 29, 33 x .
Statistical e¨aluation
Data for locomotor activity and grooming, pre-
sented as means"
SEM
of the cumulative values of
the two parameters obtained for each rat in the test
period, were analysed using, as appropriate:
Kruskal Wallis test followed by Mann Whitney U-
test. As for vocalization, only its presence or absence
was noted and the percentage of animals vocalizing
was analysed using ANOVA followed by
Student Newman Keuls test.
Data for X-maze test are presented as means"
SEM
; the number of entries into the open arms is
shown as a percentage of the total number of arm
entries, and the time spent in the open arm is
similarly given as a percentage of the total time
spent in the arms. Grooming values are the means
"
SEM
of the cumulative seconds spent in the X-
maze. All parameters, as well as the number of total
entries, were analysed by the ANOVA followed by
Student Newman Keuls test and Student’ t-test.
Data for the swimming test are presented as num-
ber of animals displaying wall-hugging per group;
chi-squared test was used.
The level of significance was set at P-0.05. At
least six animals were used in each treatment group,
the exact number of rats is reported in the figures
and tables.
RESULTS
Test 1: locomotor acti¨ity, grooming and ¨o-
calization in the obser¨ation cages
Results obtained in Test 1 are reported in Fig. 1.
Acute and sub-chronic cannabinoid Ž 25 100 g
kg y1 , i.p. . dose-dependently decreased locomotor
activity w Fig. 1 Ž a .x , but the inhibitory effect obtained
with the highest dose was significantly lower after
sub-chronic than after acute treatment. At no dose
was any significant modification of this parameter
observed after 24 h abstinence with respect to the
controls. While acute HU 210 potently inhibited rat
grooming, sub-chronic treatment enhanced it, the
effect persisting after 24 h abstinence for the doses
of 50 and 100 g kg y1 w Fig. 1 Ž b .x . The percentage of
rats vocalizing increased dose-dependently in all
three experimental modes w Fig. 1 Ž c .x .
Table I shows locomotor activity, grooming and
vocalization displayed by rats sub-chronically treated
with HU 210 at 100 g kgy1, 7 days after the last
injection. While locomotor activity and vocalization
were similar to those of vehicle-treated animals,
grooming was found to be higher.

Fig. 1. Effect of HU 210 on locomotor activity, grooming

and vocalization in rats. HU 210 Ž HU 25 100 g kg y1. or
vehicle period Ž 30 were min i.p. . . injected 50 Each histogram min before the is the mean"
observation
SEM
of the
cumulative each rat scores Ž a, or the percentage locomotor activity; b, of animals vocalizing grooming Ž c, . vocal-
for
ization . . Number of rats used: acute treatments, eight
animals for all groups; sub-chronic treatments, vehicles6,
HU 8, HU 25s8, 25s8, HU HU 50s8, 50s11, HU 100s6; HU 100s7. abstinence, ŽU. Significantly
vehicles
different from the respective vehicle-treated rats; Ž '
. significantly different from acutely treated rats at the
same dose; Ž Kruskal Wallis followed by Mann Whitney
U-test . . Ž B . Significantly different from the respective
vehicle treated rats Ž ANOVA followed by Student
Newman Keuls test . .
Test 2: X-maze
Figure 2 Ž a c . shows the results obtained in rats
undergoing the X-maze test. Vehicle-injected rats
always exhibited an adversion for open arms w Fig.

LOCOMOTOR ACTIVITY
Score 8 M
Vehicle
A
GROOMING
A
A
Vehicle HU 25 HU 50
(2)
Pharmacological 50
Research, Vol. 41, No. 1, 2000 Table I
Effect of HU 210 ( 100 g kg I1 ) on locomotor activity,
grooming and vocalization in test 1, 7 days after last
injection
Motor acti¨ity Grooming Vocalization
( score ) ( score ) ( %
) Vehicle 7.7"0.1 2.5"0.4 12
HU 210 7.5"0.2 8.0"1.5 25
HU 210 Ž 100 g kg y1. or vehicle were i.p. injected.
Each value for locomotor activity or grooming is the
mean"
SEM
of the cumulative scores for each rat during
the observation period Ž 30 min . ; the percentage of vocaliz-
ing animals is shown. Eight animals were used for each
group.
U P-0.05 ¨s vehicle Ž Mann Whitney U-test . .
2 Ž a,b .x . However, it was observed that the controls
tested after sub-chronic treatment and during absti-
nence spent more time in the open arms than those
treated Acute acutely HU 210 seemed Ž F 2,15
s12.9, to Ps0.000 .w Fig. reduce the permanence
2 Ž b .x .
into open arms, with respect to controls, only at the
dose of 25 g kg y1 w Fig. 2 Ž a,b .x , although signifi-
cance was not reached; no result was obtained with
the other two doses w Fig. 2 Ž a,b .x , probably owing to a
state of marked hypoactivity w Fig. 2 Ž c .x that ran-
domly froze the animals in the arms. After sub-
chronic treatment there was a significant reduction
in the number of open arm entries after doses of 50
and 100 g there was in kg the y1 time Ž F 3,20
spent s4.5, there Ps0.01 at .w Fig. 2 Ž a .x , as
all three dosage
levels abstinence, Ž F 3,20
s8.4, there Ps0.000 .w Fig. Ž 2b .x . was a reduction in the After 24 h
number of
open and 2 Ž b .x in in arm the the time entries animals spent Ž F treated 3,20
there s5.1, Ž with F 3,20
Ps0.009 s4, the .w Fig. 2 Ž a
.x Ps0.02 .w Fig.
cannabinoid at
the dose of 100 g kgy1. The number of total
entries w Fig. 2 Ž c .x decreased dose-dependently after
acute HU 210 and, to a lesser extent, treatment after Ž F sub-chronic 3,20
s16.8, Ps0.000
. treatment
Ž of F 3,20
vehicle-treated s5.8, Ps0.005 . but remained similar to that
rats during abstinence.
Comparison of the influence exerted on rats by
the same dose of HU 210 in each of the three
experimental modes Ž acute, sub-chronic or absti-
nence . , showed remarkable differences among the
groups. At the dose of 25 g kgy1, the time spent in
the open arms after sub-chronic treatment and dur-
ing abstinence was higher than after acute treatment
Ž ence F 2,15
s8.9, occurred Ps0.003 for 0.007 . w Fig. 2 Ž c .x : .w Fig. 2 Ž b .x . A similar differ-
total arm entries at the dose of 50 Ž F g 2,15
kg s7, y1 , Ps
total
arm entries during abstinence were significantly
highest than after acute and sub-chronic treatment
Ž Ž F F 2,15
2,15
Grooming s16.2, s63.8, Ps0.000 . , as they were at 100 g kg
y1 Ps0.000 . .
Ž Fig. 3 . , virtually non-existent in all but
a few of the acutely-treated controls, was completely

Fig. 2. Effect of HU 210 on X-maze behaviour in rats.

HU 210 Ž HU 25 100 g kg y1 or vehicle were i.p. injected
60 min before the test Ž 5 min . . Each histogram is the
mean"
SEM
of the percentage of open arm entries Ž a . ,
time ber spent there Ž b . of rats used: six or animals number for of all total groups; entries ŽU. Ž c . . Signifi-
Num-
cantly Ž .' different significantly from the respective vehicle treated rats;
different from acutely-treated rats at the
same the dose; Ž same dose . significantly Ž ANOVA different followed from abstinent by rats at
Student
Newman Keuls test . .
abolished by acute HU 210 at all Ps0.000 . but increased after sub-chronic doses Ž F 3,20
s8.2,
cannabi-
noid Ž F 3,20
s6.1, Ž F 3,20
s7.6, Ps0.004 Ps0.001 . . . Here, and during abstinence
too, the same dose of
HU 210 affected the rats differently, depending on
the experimental mode: after sub-chronic treatment
at all three doses grooming was higher than in the
other two Ps0.000 . Table II and groups shows Ž F 2,15
the Ž F s8.2, 2,15 X-maze s6.1, Ps0.004 performance Ps0.01 . , .Ž , respectively.
F 2,15
s13,
displayed
by rats sub-chronically treated with HU 210 at the
dose of 100 g kgy1, 7 days after the last injection.
While total entries were similar to those of vehicle-

ENTRIES 1N OPEN ARMS (a)

Vehicle HU 25
TIME SPENT IN OPEN ARMS
Vehicle HU 25
TOTAL ENTRIES
AA
Acute H Sub-chronic E Abstinence
Fig. 3. mean"
Effect of SEM
of the HU 210 on grooming in cumulative time displayed the X-maze. by each rat Animals during the test and period treatments
Ž 5 min as .Ž . ) in . Fig. Significantly 2. Each histogram is different from the
the
respective vehicle-treated different from abstinent rats; rats at the Ž ' . same significantly dose Ž ANOVA different followed
from acutely-treated by Student rats Newman at the Keuls same test . .
dose; Ž . significantly
treated rats, the number of open arm entries and
the time spent there were significantly reduced Ž ts
4.9, Ps0.000; ts4.9, Ps0.000, respectively . and
grooming was potentiated Ž ts4.8, Ps0.000 . .
Test 3: swimming test
Table III shows that, after acute treatments, all
groups behaved similarly in their swimming strategy;
in the sub-chronic mode, rat treated with 50 and 100
g kgy1 followed an enhanced thigmotactic pattern
Ž 2 s12.6; Ps0.007 . . The phenomenon was con-
firmed during 24 h abstinence Ž 2 s10.7; Ps0.01 . .
DISCUSSION
Our results shed further light on the behavioural
effects of HU 210 w 16, 13 x , in particular on the
anxiogenic-like properties of this potent synthetic
cannabinoid w 21 x . As expected, when acutely admin-
istered to rats, the drug dose-dependently produced
sedation and reduced locomotion, as also occurs
after 9 -THC and other cannabinoids w 20 x . In gen-
eral, tolerance, more or less complete, has been
Table II
Effect of HU 210 ( 100 g kg I1 ) on X-maze test, 7 days
after last injection
Treatments Open arms
Entries Time spent Total entries Grooming
( % ) ( % ) ( no. ) () s
Vehicle HU 210 37.5"2.7 13.8"3.5 U 23.5"2.6 7.7"1.9 U 11.5"1.0 0
9.5"0.7 21.8"4.5
U HU 210 or vehicle were i.p. injected. Each value is the
mean"
SEM
demonstrated for most of the pharmacological ef-
fects of cannabinoids w 15, 20 x , and in the present
study partial tolerance for depression of locomotor
activity was confirmed. In addition, comparison of
the inhibitory effects on this parameter in Test 1
Ž locomotor activity . and Test 2 Ž number of total
entries in the X-maze . showed that the phenomenon
of tolerance strictly depends on the dose and the
experimental model used. In fact, as already pointed
out w 34 x , it is practically impossible to obtain the
same results in behavioural models that elicit dif-
ferent emotional states in the animal. Although the
animals acutely injected with 50 and 100 g kgy1 of
HU 210 exhibited marked sedation, they were hy-
persensitive to tactile stimuli and vocalized strongly
when touched. Vocalization is considered a pointer
of cannabimimetic activity w 30 x , and it is elicited by
9-THC at doses much higher than those of HU 210.
Since, at the doses of 25 and 50 g kgy1 HU 210
has proved to be analgesic in rats Ž data not yet
published . the possibility that vocalization might be
due to pain can be excluded, rather, it seems to
reflect enhanced irritability in response to tactile
stimulus. In accordance with other authors w 20 x , we
Table III
Effect of HU 210 on swimming test
Treatment ( g kg
y1 ) Wall-hugging
Acute Sub-chronic Abstinence
Vehicle 2r8 1r8 1r8
HU 210, 25 1r8 0r8 1r8
RU 210, 50 4r8 HU 210, 100 4r8 5r8 6r8 U 5r8
6r8
U HU 210 or vehicle were i.p. injected. Data are pre-
sented as number of animals displaying wall-hugging dur-
ing U the P-0.05 test period ¨s of eight animals, during the observation pe-
riod U P-0.05 Ž 5 min . .
¨s vehicle Ž Student t-test . .
Ž 2 min . .
vehicle Ž chi-squared test . .

Pharmacological Research, Vol. 41, No. 1, 2000 51

Seconds
GROOMING
Pharmacological 52
Research, Vol. 41, No. 1, 2000 therefore hypothesize that vocalization, like aggres-
siveness, which is frequently reported in cannabi-
noid-treated rodents subjected to stress-inducing
procedures w 16, 20 x , is a behavioural expression of
heightened emotionality associated with a state of
fear.
The correlation between cannabinoids and stress
has long been proposed w 35, 36 x and supported by
experimental findings on animals, namely, that
cannabinoids induce a potent secretion of ACTH
w 22 x and CRF w 16 x , which, as is known, play a key
role in stress w 37, 23 x . Moreover, the attenuation
exerted by the CRF antagonist
D
been used to highlight anxiogenic-like effects of
several cannabinoids, and acute HU 210, in particu-
lar, has been found to intensify animals’ aversion to
the open arms w 21 x . No significant change in X-maze
behaviour has been seen by the authors during with-
drawal, for any of the compounds tested.
In the present study sub-chronically treated and
abstinent rats were consistently different in their
behaviour from vehicle-injected animals, and the
parameters that were found to be modified Ž groom-
ing and exploration of novel environments . an-
thropomorphically reflect an anxiogenic response.
-phenyl CRF12 41
on rat anxiogenic responses to HU 210 w 16 x strongly
suggests the mediation of endogenous CRF systems
in these effects.
Increased rat grooming, which has been recog-
nized as occurring under certain mildly-stressful
events, has been found to be markedly potentiated
by central administration of CRF w 24 x . CRF releases
ACTH from the pituitary and ACTH also stimulates
grooming w 23 x . Our findings that grooming is en-
hanced after sub-chronic HU 210 at 50 and 100 g
kgy1 is therefore not surprising. In addition, we
observed a grooming syndrome during abstinence
after the highest dose of the compound. In contrast
with the present data, which suggest a behavioural
sensitization, grooming has been found to be de-
pressed by acute HU 210 while remaining unaf-
fected in sub-chronically-treated rats, in which the
drug merely developed a behavioural tolerance to
this decrease w 15 x .
The hypothesis that the grooming enhancement
observed after sub-chronic treatment simply de-
pends on a phenomenon of tolerance to the sedative
effects of the drug is not tenable. In fact, sub-chronic
treatment at 25 and 50 g kgy1 did not produce an
Aversion for open spaces and wall-hugging, as per-
sistence in trying to escape along the walls of the
pool, have been attributed to an anxiety-like state
w 27 29 x .
Our data on vocalization, in line with those re-
ported for 9 THC w 25 x , apparently contradict a sup-
posed enhancement of fear response in tolerant and
abstinent animals, for vocalization was slightly higher
after acute HU 210 injection than in the other two
experimental modes. At present we are not in a
position to say whether tolerance is responsible for
this phenomenon. It might be simply ascribable to
the fact that our acutely-injected animals were un-
used to any handling, unlike those in the other two
groups, for adaptation to handling seems to be the
main explanation for the less anxious behaviour of
subchronically-treated control rats in the X-maze
test.
In conclusion, in view of the number of similari-
ties between animal and human behaviour after
cannabinoids, our study suggests an anxiogenic activ-
ity of chronic the potent treatment at CB high 1
agonist HU 210, after sub-
doses, and the persistence
of enhanced emotional response to novel environ-
ments when the drug is discontinued.
effect significantly different from that after acute
treatment on locomotor activity, which remained
significantly different from that of vehicle-treated
ACKNOWLEDGEMENTS
rats; however, in these two groups enhancement of
grooming was evident. At the dose of 25 g kgy1,
grooming was higher than that after acute treat-
This work was supported by grants from Ministero
della Pubblica Istruzione Ž 60% . and by CNR.
ment; at that of 50 g kgy1 it was also higher than
that observed in vehicle-treated rats. The dose of
100 g kgy1 produced a partial tolerance to the
REFERENCES
inhibition of locomotor activity, which remained
lower than that of vehicle-treated rats. In this case
too, grooming was much higher than that of
1. Gaoni Y, Mechoulam R. Isolation, structure and
partial synthesis of an active constituent of hashish. J
Am Chem Soc 1964; 86: 1646 7.
vehicle-treated rats and not simply the same, as
would occur if the phenomenon were strictly depen-
dent on tolerance to sedative effects.
While caution must be exercised in attributing
2. Bloom AS. Effect of 9-tetrahydrocannabinol on the
synthesis of dopamine and norepinephrine in mouse
brain synaptosomes. J Pharmacol Exp Ther 1982; 221:
97 103.
3. Poddar MK, Biswas B, Ghosh JJ. 9-THC and brain
emotional responses to specific animal behavioural
biogenic amines. In: Drugs and central synaptic trans-
patterns, our results in the X-maze test also confirm
that, after sub-chronic treatment with certain doses
of HU 210 and during abstinence, the rats displayed
an anxiety-like state. The X-maze test has already
mission. Bradley PB, Dhawan BN eds. London:
MacMillan, 1976: 193 9.
4. Tripathi HL, Vocci FJ, Dewey WL. Effect of
cannabinoids on cholinergic systems in various re-
gions of the mouse brain. Fed Proc 1979; 38: 590.
Pharmacological Research, Vol. 41, No. 1, 2000 53
5. Devane WA, Dysarz FAIII, Johnson MR, Melvin LS,
Howlett AC. Determination and characterization of
a cannabinoid receptor in rat brain. Mol Pharmacol
1988; 34: 605 13.
6. Harris LS, Carchman RA, Martin BR. Evidence for
the existence of specific cannabinoid binding sites.
Life Sci 1978; 22: 1131 8.
7. Herkenham M, Lynn AB, Johnson MR, Melvin LS,
De Costa BR, Rice KC. Characterization and locali-
sation of cannabinoid receptors in rat brain: a quan-
titative in ¨itro autoradiographic study. J Neurosci
1991; 11: 563 83.
8. Nye JS, Seltzman HH, Pitt CG, Snyder SH. High-
affinity cannabinoid binding sites in brain mem-
branes labelled with w3 H x -5 -trimethylammonium
Ž 8 -THC . . J Pharmacol Exp Ther 1985; 234: 784 91.
9. Collins DR, Pertwee RG, Davies SN. The action of
synthetic cannabinoids on the induction of long-term
potentiation in the rat hippocampal slice. Eur J Phar-
macol 1994; 259: R7 8.
10. Compton DR, Johnson MR, Melvin LS, Martin B.
Pharmacological profile of a series of bicyclic
cannabinoid analogs: classification as canna-
bimimetic agents. J Pharmacol Exp Ther 1992; 260:
201 9.
11. Compton DR, Rice KC, De Costa BR, Razdan RK,
Melvin LS, Johnson MR, Martin BR. Cannabinoid
structure activity relationship: correlation of recep-
tor binding and in ¨i¨o activities. J Pharmacol Exp
Ther 1993; 265: 218 26.
12. Little PJ, Compton DR, Johnson MR, Melvin SL,
Martin BR. Pharmacology and stereoselectivity of
structurally novel cannabinoids in mice. J Pharmacol
Exp Ther 1988; 247: 1046 51.
13. Little PJ, Compton DR, Mechoulam R, Martin
BR. Stereochemical effects of 11-OH- 8-THC-
dimethyleptyl in mice and dogs. Pharmacol Biochem
Beha¨ 1989; 32: 661 6.
14. Howlett AC, Champion TM, Wilken GH, Mechou-
lam R. Stereochemical effects of 11-OH-delta 8-
tetrahydrocannabinol-dimethylheptyl to inhibit
adenylate cyclase and bind to the cannabinoid recep-
tor. Neuropharmacology 1990; 29: 161 5.
15. De Fonseca FR, Calderon JL, Mechoulam R, Navarro
M. enhances Repeated Ž y stimulation . -11-hydroxy-D8-tetrahydrocannabinol-
of D1 dopamine receptors
dimethylheptil-induced catalepsy in male rats. Neu-
roreport 1994; 5: 761 5.
16. De Fonseca FR, Rubio P, Menzaghi F, Merlo-Pich
E, Rivier J, Koob GF, Navarro M. Corticotropin-
releasing factor Nle21,38, CaMeLeu37 Ž CRF x CRF . antagonist w D-PHE12,
attenuates the acute ac-
tions of the highly potent cannabinoid receptor ago-
nist HU 210 on defensive-withdrawal behaviour in
rats. J Pharmacol Exp Ther 1996; 276: 56 64.
17. Fishman MW, Rosenbaum JF, Yabusaki DI, Carr
DB. Marijuana related anxiety: a questionnaire-based
pilot study of normal and psychiatric populations.
Res Commun Subst Abuse 1988; 9: 219 26.
18. Weil AT, Zinberg NE, Nelsen JM. Clinical psycho-
logical DC ) 1968; effects 162: of 1234 marijuana 42.
in man. Science ( Wash.
19. Zuardi AW, Shirakawa I, Finkelford E, Karniol IG.
Action of cannabinol on the anxiety and other effects
produced by 9-THC in normal subjects. Psychophar-
macolology 1982; 76: 245 50.
20. Dewey WL. Cannabinoid pharmacology. Pharmacol
Re¨ 1986; 38: 151 78.
21. Onaivi ES, Green MR, Martin BR. Pharmacological
characterization of cannabinoids in the elevated plus
maze. J Pharmac Exp Ther 1990; 253: 1002 9.
22. Dewey WL, Peng TC, Harris LS. The effect of 1-trans-
9THC on the hypothalamohypophyseal-adrenal axis
of rats. Eur J Pharmacol 1970; 12: 382 4.
23. Gispen WH, Wiegant HM, Greven HM, De Wied D.
The induction of excessive grooming in the rat by
intraventricular application of peptides derived from
ACTH-structure activity studies. Life Sci 1975; 17:
645 50.
24. Morley JE, Levine AS. Corticotrophin releasing fac-
tor, grooming and ingestive behaviour. Life Sci 1982;
31: 1459 64.
25. Jarbe TUC, Hiltunen AJ. Cannabimimetic activity of
cannabinol in rats and pigeons. Neuropharmacology
1987; 26: 219 28.
26. Romero J, Garcia L, Cebeira M, Zadrozny D, Fer-
nandez-Ruiz JJ, Ramos JA. The endogenous
cannabinoid receptor ligand, anandamide, inhibits
the motor behaviour: role of nigrostriatal dopaminer-
gic neurons. Life Sci 1995; 56: 2033 40.
27. Pellow S, Chopin P, File SE, Briley M. Valuation of
open-closed arm entries in an elevated plus-maze as
a measure of anxiety in the rat. J Neurosci Meth 1985;
14: 149 67.
28. Sutherland RJ, Rudy JW. Place learning in the Mor-
ris place navigation task is impaired by damage to
the hippocampal formation even if the temporal de-
mands are reduced. Psychobiology 1988; 16: 157 63.
29. Ferrari F, Ottani A, Giuliani D. Learning impair-
ment produced in rat by the cannabinoid agonist HU
210 in a water maze task. Pharmacol Bioch Beha¨
1999: in press.
30. Henriksson BG, Jarbe TUC. Cannabis-induced vocal-
ization in the rat. J Pharm Pharmacol 1971; 23:
457 8.
31. Ferrari F, Giuliani D. Influence of eticlopride on
cocaine- fects in and rats. D Pharmacol A D 2
agonist-induced Biochem behavioural Beha¨ 1996; ef-
53:
523 30.
32. Ferrari F, Tartoni PL, Mangiafico V. B-HT 920 an-
tagonizes rat neophobia in the X-maze test: a com-
parative study with other drugs active on adrenergic
and dopaminergic receptors. Arch Int Pharmacodyn
Ther 1989; 298: 7 14.
33. Ferrari F, Ottani A, Giuliani D. Cannabinoid activity
in rats and pigeons of HU 210, a potent anti-emetic
drug. Pharmacol Biochem Beha¨ 1999; 62: 75 80.
34. Ferrari F, Giuliani D. Effect on rat feeding be-
haviour of two selective iol Beha¨ 1994; 56: 921 D 6.
2
dopamine agonists. Phys-
35. Kumar MS, Chen CL. Effect of an acute dose of
9-tetrahydrocannabinol on hypothalamic luteinizing
hormone releasing hormone and met-enkephalin
content and serum levels of testosterone and corti-
costerone in rats. Subst Alcohol ActionsrMisuse 1983;
4: 37 43.
36. Maclean KI, Littleton JM. Environmental stress as a
factor in the response of rat brain catecholamine
metabolism to 9THC. Eur J Pharmacol 1977; 41:
171 82.
37. Dunn AJ, File SE. Corticotropin-releasing factor has
an anxiogenic action on the social interaction test.
Horm Beha¨ 1987; 21: 193 202.