JNJ-10181457, a selective non-imidazole histamine H3receptor antagonist,
normalizes acetylcholine neurotransmission and has efficacy
in translational rat models of cognition
Ruggero Galici*, Jamin D. Boggs, Leah Aluisio, Ian C. Fraser, Pascal Bonaventure,
Brian Lord, Timothy W. Lovenberg
Johnson & Johnson Pharmaceutical Research & Development, L.L.C., 3210 Merryfield Row, San Diego, CA 92121, USA
a r t i c l e i n f o
Received 26 November 2008
Received in revised form
31 January 2009
Accepted 24 March 2009
a b s t r a c t
Histamine 3 (H3) receptors are distributed throughout the brain and regulate histamine as well as the
activity of other neurotransmitters including acetylcholine (ACh). Impaired ACh neurotransmission is
associated with deficits of cognitive-related functioning in many species including humans. The goal of
these studies was to evaluate the behavioral and neurochemical effects of JNJ-10181457, a selective non-
imidazole histamine H3receptor antagonist, in rats. The pharmacokinetic profile and receptor occupancy
of JNJ-10181457 were tested. The efficacy of JNJ-10181457 was evaluated, acutely, in the imetit-induced
water licking model, delayed non-matching to position (DNMTP) task and microdialysis studies. In
addition, the effects of repeated administration of JNJ-10181457 were evaluated in the reversal learning
task. A single administration of JNJ-10181457 (10 mg/kg, i.p.) resulted in significant plasma and brain
exposure and maximal H3receptor occupancy. In addition, JNJ-10181457 reversed imetit-induced water
licking, similarly to thioperamide (10 mg/kg, i.p.). In the DNMTP task, scopolamine (0.06 mg/kg, i.p.)
significantly decreased percentage correct responding. These effects were significantly reversed by JNJ-
10181457 (10 mg/kg, i.p.) and also by donepezil (1 mg/kg, i.p.), an acetylcholinesterase inhibitor, and
were associated with normalization of ACh neurotransmission in the cortex. Repeated administration of
JNJ-10181457 (10 mg/kg, i.p.) significantly increased percentage correct responding in the reversal
learning task. Treatment discontinuation was not associated with rebound effects on cognition. These
results indicate that selective blockade of histamine H3receptors might have therapeutic utility for the
treatment of working memory deficits and learning disorders, especially those in which ACh neuro-
transmission is compromised.
? 2009 Elsevier Ltd. All rights reserved.
Histamine mediates many important biological functions
including allergic reactions, gastric acid secretion, neurotransmitter
release and immunomodulation (Lovenberg et al.,1999; Thurmond
et al., 2008). The effects of histamine are mediated via four receptor
subtypes (i.e. H1, H2, H3and H4) which are uniquely distributed in
the central nervous system (CNS) and also in the periphery (Leurs
et al., 2005; Parsons and Ganellin, 2006). H3 receptors are
G-protein-coupled autoreceptors and regulate the release of hista-
mine in thebrain, particularly in thecortex, striatum, hippocampus,
amygdala and substantia nigra (Leurs et al., 1998; Lovenberg et al.,
neurotransmissionwhich is associated with increased wakefulness,
cognitive enhancement and suppression of food intake (Leurs et al.,
1998; Witkin and Nelson, 2004) and these effects are likely
mediated by post-synaptic histamine H1and H2receptors (Haas
et al., 2008; Pascoli et al., 2009). H3receptors are also hetero-
receptors; thus, their activation reduces, whereas blockade
increases, not only the release of histamine but also several other
neurotransmitters including acetylcholine
nor-epinephrine,GABAandglutamate(Leurset al.,1998;Haas etal.,
2008; Blandina et al., 1998).
Changes in ACh release are associated with cognitive deficits or
improvements depending on whether H3receptors are activated or
blocked, respectively. For example, imetit and R-alpha-methyl-
histamine, two H3agonists, reduce ACh release and these neuro-
chemical effects are associated with deficits in the passive
avoidance and object recognition tests (Blandina et al., 1996). In
addition, both H3agonists increase water intake and the reversal of
* Corresponding author. Tel.: þ1 858 353 6584.
E-mail address: email@example.com (R. Galici).
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0028-3908/$ – see front matter ? 2009 Elsevier Ltd. All rights reserved.
Neuropharmacology 56 (2009) 1131–1137
this effect by H3antagonists is often used as in vivo functional
activity readout (Medhurst et al., 2007). For example, thioperamide
and clobenpropit two H3 antagonists, block H3 agonist-induced
drinking, increase ACh and have procognitive effects in several
behavioral assays (see Bacciottini et al., 2001 for a review).
Decreased ACh neurotransmission is associated with cognitive-
related pathologies such as Alzheimer’s disease, schizophrenia, and
attention deficit hyperactivity disorder (Leurs et al., 1998; Witkin
and Nelson, 2004). Thus, it has been suggested that normalization
of ACh neurotransmission, by blocking H3 receptors, should
ameliorate these disorders. Furthermore, expression of H3recep-
tors is still prevalent even in severe late stages of Alzheimer’s
disease, an important observation given the aim of targeting these
receptors as a potential novel therapeutic approach (Medhurst
et al., 2007).
Scopolamine is a non-selective muscarinic antagonist that
induces cognitive deficits by regulating brain concentrations of ACh
in many species, including rodents (Higgins et al., 2002; Robbins,
2002) and humans (Drachman and Leavitt, 1974; Bartus et al.,
1982). H3antagonists have been shown to reverse cognitive deficits
induced by scopolamine in different behavioral assays (Hancock
and Fox, 2004; Witkin and Nelson, 2004). However, there is
a paucity of data with regards to the effects of H3antagonists in
translational cognitive models. For example, in one study, thio-
peramide failed to reverse the scopolamine-induced deficits in the
delayed matching to position task (Kirkby et al., 1996). In two
separate studies, ciproxifan was evaluated in the 5-choice serial
reaction time task and it increased percent correct response only
when a very short stimulus duration was imposed (Ligneau et al.,
1998; Day et al., 2007).
JNJ-10181457 is a neutral, potentand selective non-imidazoleH3
antagonist. This compound increases NE and ACh concentrations in
rat frontal cortex and has efficacy similar to methylphenidate in the
7-trial repeated acquisition passive avoidance task (Bonaventure
et al., 2007), a behavioral assay conducted in spontaneous hyper-
tensive (SHR) pups and thought to measure memory acquisition.
The goal of these studies was two-fold: first, to evaluate the
effects of JNJ-10181457 in translational rat models of cognition.
Specifically, experiments were conducted using the delayed non-
matching to position (DNMTP) and the reversal learning tasks.
These are two behavioral procedures thought to measure reference,
working memory and learning related processes in rodents and
humans (Dunnett,1985; Clark et al., 2004). The second goal of these
studies was to evaluate the neurochemical effects of JNJ-10181457
on cortical ACh release.
Sprague–Dawley rats (Harlan, Indianapolis, Ind., USA) were used. For the
delayed non-matching to position task and reversal learning experiments, animals
were maintained at 80–90% of their free feeding weight and were trained to respond
for food in multiple trials per session. For imetit-induced drinking, pharmacoki-
netics and microdialysis studies animals had also ad libitum access to food. All
animals were single-housed in plastic cages under 12 h:12 h light:dark schedule and
humidity controlled rooms and had ad libitum access to water. All animal experi-
ments were carried out in accordance with the National Institutes of Health guide
for the care and use of laboratory animals (NIH Publications No. 8023, revised 1978)
and were approved by the Institutional Animal Care and Use Committee at Johnson
& Johnson Pharmaceutical Research & Development, L.L.C., San Diego.
2.2. Pharmacokinetics and receptor occupancy
Blood brain barrier and ex vivo autoradiography studies were performed as
previously described by Barbier et al. (2007). Briefly, rats received a single adminis-
administration. Dosing was followed by blood sampling via cardiac puncture over
a time course. Brains were removed from the animals and half of the brain was
homogenized for LC/MS–MS analysis, the other half was frozen on dry ice for ex vivo
sample with acetonitrile with vigorous mixing. These samples were incubated for
5 min, and then centrifuged at 14,000 rpm in a micro-centrifuge for 4 min. The
Samples were analyzed by LC-MS/MS. Tissue section for ex vivo receptor autoradi-
ography was prepared as previously described (Langlois et al., 2001).
Ex vivo occupancy of histamine H3receptor binding sites, was measured in the
cortex of each individual rat. After thawing, the sections were dried under a stream
of cold air and then incubated at room temperature for 1 min with [3H] Methyl-
histamine (2 nM) in Na/K Phosphate Buffer (50 mM, pH 7.4). The sections were not
washed prior to incubation, to avoid dissociation of the drug–receptor complex.
Incubation was restricted to 1 min at room temperature to minimize dissociation of
the drug from the receptor. Three adjacent brain slices from the same animal have
been collected per slide. Two brain slices were used to measure the total binding,
and the third one was evaluated for nonspecific binding. After the incubation, the
slides werewashed in ice-cold buffer, followed bya quick rinse in ice-cold water. The
sections were then dried under a stream of cold air. Quantitative analysis was per-
formed as previously described (Langlois et al., 2001) using the b-imager (BioSpace,
Paris, France). Ex vivo H3receptor labeling was expressed as the percentage of
receptor/transporter labeling in corresponding brain areas of saline-treated animals.
The percentage of H3receptor occupancy was plotted against time.
2.3. In vivo microdialysis in freely moving rats
Dialysis experiments were conducted using a between subject design. A guide
cannula (Eicom, Kyoto, Japan) was implanted into the prefrontal cortex (incisor bar,
?3.5 mm, þ3.2 mm anterior, 0.8 mm lateral and 1 mm ventral to Bregma) (Paxinos
and Watson, 1997). Animals remained in their home cage throughout experiments.
Dialysis probes (Eicom, 4 mm active membrane length) were perfused with artificial
cerebral spinal fluid at a flow rate of 1 ml/min and implanted the afternoon prior to
sample collection. The following morning five 20-min periods of baseline samples
were collected into a 96-well plate (Sarstedt, 96 well multiply PCR) via a four-
channel fraction collector (Eicom) prior to drug injections. Then animals received
one injection of saline or JNJ-10181457 (10 mg/kg, i.p.) and 30 min later they
received a second injection of water (s.c.) or scopolamine (0.06 mg/kg, s.c.). There-
fore, each experiment included the following conditions: saline þ water,
saline þ scopolamine, JNJ-10181457 þ water and JNJ-10181457 þ scopolamine. This
experimental design was chosen to match the DNMTP conditions (see behavioral
procedures below). Following the second injection, samples were collected into the
96-well plate, every 20 min for up to 2 h, and maintained at 4?C containing 7.5 ml of
the antioxidant (0.1 M acetic acid, 1 mM oxalic acid and 3 mM L-cysteine in sterile
water). Samples were then analyzed for ACh by LC with MS/MS detection. A Thermo
Hypersil-Keystone, Hypurity Aquastar, 2.1 ? 50 mm, 5 mm column was used with
a mobile phase containing a mixture of 20 mM ammonium acetate in water (solvent
A) and 0.1% formic acid in acetonitrile (solvent B). The mobile phase was held with
an A:B ratio of 100:0 for 1 min. The linear gradient was then changed over the next
0.5 min to an A:B ratio of 5:95 and held for 1 min, then returned to the starting
conditions. The total run time was 3.5 min and flow rate was 0.6 mL/min (Shimadzu
LC-10AD VP with SCL-10A VP system controller). Tandem mass spectrometric (MS/
MS) detection was carried out on a PE Sciex API4000 in the positive ion mode (ESI)
by multiple reaction monitoring (MHþ/daughter was 146.1?87.1 m/z). The detection
limit for ACh was 0.025 pg/10 mL.
Imetit-induced water licking, DNMTP and reversal learning
experiments were conducted in sound attenuating operant cham-
bers (Med-Associates, St Albans, VT) equipped with a houselight,
two lights, one on the left and one on the right of the frontal panel,
two retractable levers positioned below the lights and a food
receptacle. Water bottles equipped with spouts were mounted
outside the chambers when imetit-induced water licking experi-
ments were conducted.
4.1. Imetit-induced water licking
Experiments were conducted using a between subject design.
Animals were placed into the operant chambers. The beginning of
R. Galici et al. / Neuropharmacology 56 (2009) 1131–11371132
the session was signaled by the illumination of the houselight.
Water was delivered when animals licked the spout. Each lick also
interrupted a circuit and generated a count. Total number of licks
was recorded over a period of 60 min. Saline, JNJ-10181457
(10 mg/kg, i.p.) or thioperamide (10 mg/kg, i.p.) was administered
60 min whereas water or imetit (3 mg/kg, s.c.) was administered
30 min prior to the beginning of the session.
4.2. DNMTP task
Experiments were conducted using a within subject design.
Each trial started with the illumination of the houselight and with
the random presentation of one lever. Initially, when the animal
pressed the lever it was retracted and immediately thereafter (i.e.
no delay) both levers were presented. Pressing the lever that was
not initially presented (i.e. correct response) resulted in the illu-
mination of the left and right lights for 1 s, retraction of the levers,
and the delivery of a 45 mg food pellet (Bio-Serv, Frenchtown, NJ,
USA), ended the trial and started the next one. Pressing the lever
that was initially presented (i.e. incorrect response) retracted both
levers, ended the trial and started a new one. Lack of response did
not have anyconsequence. Each session lasted 30 min. This training
phase was conducted until correct responding was >90% and
a stable baseline was obtained (i.e. correct responding did not
change by more than 20% of the mean value for 5 consecutive
sessions). When these criteria were met, a random delay was
introduced (0, 4, 8, 16 or 32 s) and animals had to complete the
same task described above. Under these conditions, the first test
was conducted when the following acquisition criteria (i.e. base-
line) were met: Total number of trials and % correct responding for
each delay did not vary by more than 20% of the mean value for 5
consecutive sessions and at least 5 trials/delay were completed.
Subsequent tests wereconducted when total number of trials and %
correct responding for each delay did not vary by more than 20%
from the baseline. However, at least two days always elapsed
between tests. Saline, donepezil (1 mg/kg, i.p.), thioperamide
(10 mg/kg, i.p.) or JNJ-10181457 (10 mg/kg, i.p.) was administered
60 min, whereas water or scopolamine (0.06 mg/kg, s.c.) was given
30 min prior to the beginning of the session.
4.3. Reversal learning
Experiments were conducted using a within subject design.
Animals responded (lever press) for food on a variable interval
schedule (5–20 s). At the beginning of each trial both levers were
available and a light above one of the levers was randomly illu-
minated. Pressing the lever associated with the light resulted in
the delivery of a 45 mg food pellet (i.e. correct response), ended
the trial and started a new one. Pressing the lever not associated
with the light (i.e. incorrect response) or lack of response (i.e.
omission) within 10 s terminated the trial and started a new one.
Daily sessions lasted 15 min. Training was conducted until
animals received 10 consecutive rewards within 5 min (acquisi-
tion criterion). After acquisition, 120 min test sessions occurred
under reversal learning by imposing a new rule. Specifically,
reward pellets were presented by pressing the lever not associ-
ated with the light. Pressing the lever associated with the light
did not have any consequence. A group of animals received water
and a separate group received JNJ-10181457 (10 mg/kg, i.p.)
30 min before the beginning of the session and for 3 consecutive
days (T1, T2 and T3). Thereafter, treatment was discontinued and
percent correct responding was evaluated for an additional 4 days
(W1, W2, W3, W4).
pholine and thioperamide hydrochloride were synthesized at
Johnson & Johnson Pharmaceutical Research & Development, L.L.C.,
La Jolla, CA. Donepezil hydrochloride was purchased from Tocris
Bioscience (Ellisville, MO) and scopolamine hydrochloride was
purchased from Sigma Aldrich (St. Louis, MO). All compounds were
dissolved in saline or water. Volume of injection was 1 ml/kg. Dose
is expressed as free base.
4.5. Statistical analysis
For imetit-induced water licking, data were analyzed with one-
way analysis of variance (ANOVA) followed by Bonferroni post hoc
test. For the DNMTP task, percentage correct responding was
analyzed with two-way repeated measures ANOVA followed by
Bonferroni post hoc test to determine treatment, time effects and
interaction time ? treatment. Number of trials was analyzed with
one-way repeated measures ANOVA. For the reversal learning task,
cumulative correct choices and percent correct responding were
analyzed with two-way repeated measures ANOVA.
For in vivo microdialysis data, percent change from baseline
values was calculated from the mean basal value of ACh for each
animal and plotted in 20 min bins. Area under the curve (AUC)
values were then calculated by the summation of the difference
between ACh post drug administration and the mean percent of
basal release value between 20 and 80 min. This period of time was
chosen because it coincides with the time of behavioral measure-
ment. One-way ANOVA was conduced on AUC data. All data anal-
yses were conducted with Prism Graph Pad (Prism, San Diego). The
level of significance was p < 0.05.
5.1. Pharmacokinetics and receptor occupancy of JNJ-10181457
We first evaluated plasma and brain exposure and receptor
occupancy. A single administration of JNJ-10181457 resulted in
robust plasma and brain concentrations (Fig.1, left axis). The brain–
plasma ratio was 15 ? 3. This robust exposure was associated with
maximal H3receptor occupancy in cortex for up to 90 min (Fig. 1,
0 3060 90
% Receptor Occupancy
Fig. 1. JNJ-10181457 pharmacokinetics and receptor occupancy. Brain and plasma
concentrations (left ordinate) and % receptor occupancy (right ordinate) after a single
administration of JNJ-10181457 (10 mg/kg, i.p.) are plotted as function of time in
minutes. Data are expressed as mean ? S.E.M. N ¼ 6/time point.
R. Galici et al. / Neuropharmacology 56 (2009) 1131–1137 1133
5.2. Effects JNJ-10181457 on imetit-induced water licking
One-way ANOVA indicated that there was a main treatment
effect [F (3,31) ¼ 10.4, Fig. 2]. Specifically, imetit significantly
increase number of water licks compared to control (p < 0.05). JNJ-
10181457 and thioperamide significantly reversed the effects of
imetit (p < 0.05).
5.3. Effects of JNJ-10181457 in the DNMTP task
Two-way repeated measures ANOVA indicated that there was
a main treatment effect in the donepezil [F (3,20) ¼ 14.7, p < 0.05,
Fig. 3a)], thioperamide [F (3,20) ¼ 15.8, p < 0.05, Fig. 3b)] and JNJ-
10181457 [F (3,20) ¼ 16.9, p < 0.05, Fig. 3c)] studies. Post hoc
analysis indicated that scopolamine induced a significant decrease
in percent correct responding and these effects were significantly
reversed by donepezil and JNJ-10181457 but not by thioperamide.
None of the compounds increased percent correct responding by
itself. In addition, there was a delay-dependent effect in the
donepezil [F (4,80) ¼ 10.6, p < 0.05, Fig. 3a)], thioperamide [F
(4,80) ¼ 6.6, p < 0.05, Fig. 3b)] and JNJ-10181457 [F (4,80) ¼ 6.8,
p < 0.05, Fig. 3c)] studies but there was not a treatment ? delay
interaction for donepezil [F (12,120) ¼ 0.8, p > 0.05], thioperamide,
[F (12,120) ¼ 0.9, p > 0.05] and JNJ-10181457 [F (12,120) ¼ 0.7,
p > 0.05]. One-way repeated measures ANOVA indicated that there
was no change in number of trials for the donepezil [F (3,23) ¼ 2.9,
p > 0.05], thioperamide [F (3,23) ¼ 2.7, p > 0.05] and JNJ-10181457
[F (3,23) ¼ 2.8, p > 0.05] studies (Table 1).
5.4. Effects JNJ-10181457 on ACh release
One-way analysis of variance (ANOVA) indicated that there was
a main treatment effect [F (3,23) ¼ 5.6, Fig. 4a and b]. Post hoc
whereas scopolamine decreased, ACh release in the cortex,
compared to control. Furthermore, although JNJ-10181457 did not
significantly reverse scopolamine-induced deficits, it normalized
ACh neurotransmission since the combination of JNJ-10181457 and
scopolamine was not significantly different from control (i.e.
saline þ water).
5.5. Effects of JNJ-10181457 in the reversal learning task
Cumulative correct choices were not statistically different in
animals that were subsequently treated (i.e. between T1 and T3)
with water (pre water-treated) and with the compound (pre JNJ-
10181457-treated) [F (1,13) ¼ 0.5, p > 0.05, Fig. 5a]. In addition,
there was a time [F (9,117) ¼ 61, p < 0.05] but not a time ? treat-
ment interaction [F (9,117) ¼ 0.6, p > 0.05]. Percent correct
responding prior to switching to the new rule was 91.2 ? 1.1 and
92.1 ? 1.9 for the pre water-treated and pre JNJ-10181457-treated
groups, respectively (T0, Fig. 5b). When the new rule was imposed
(T1) there was a significant reduction in percent correct responding
compared to the day before (T0, Fig. 5b) for pre water-treated [F
(1,13) ¼ 894, p < 0.05] and pre JNJ-10181457-treated [F (1,13) ¼ 982,
p < 0.05] groups. Repeated administration of JNJ-10181457
increased percent correct responding compared to water-treated
animals and this difference was statistically significant in the third
day (T3) of treatment [F (1,13) ¼ 6.5, p < 0.05]. Furthermore,
percent correct responding increased in both groups of animals
after discontinuation. However, performance was significantly
higher in animals treated with JNJ-10181457 up to three days after
discontinuation and returned to control values on day 4 (W4,
[F (1,13) ¼ 3.3, p > 0.05].
Fig. 2. Effects of JNJ-10181457 and thioperamide on imetit-induced water licking.
Saline, JNJ-10181457 (10 mg/kg) or thioperamide (10 mg/kg) was administered i.p.
60 min before the session. Water or imetit (3 mg/kg) was administered s.c. 30 min
before the session. *p < 0.05 indicates statistical difference from Saline þ Water;
#p < 0.05 indicates statistical difference from saline þ imetit. Data are expressed as
mean ? S.E.M. N ¼ 8/condition.
04812 16 20 24 28 32
04812 16 20 24 28 3204812 16 20 24 28 32
Delay (sec)Delay (sec)
Saline + Scopolamine
Donepezil + Scopolamine
Donepezil + Water
Saline + Water
% Correct Responding
% Correct Responding
% Correct Responding
Saline + Water
Saline + Scopolamine
Thioperamide + Scopolamine
Thioperamide + Water
Saline + Scopolamine
JNJ-10181457 + Scopolamine
JNJ-10181457 + Water
Saline + Water
Fig. 3. Effects of donepezil, thioperamide and JNJ-10181457 in the DNMTP task. Saline, donepezil (1 mg/kg, panel A), thioperamide (10 mg/kg, panel B) or JNJ-10181457 (10 mg/kg,
Panel C) was administered i.p. 60 min. Water or scopolamine was administered s.c. 30 min before the session. *p < 0.05 indicates statistical difference from Saline þ Water;
#p < 0.05 indicates statistical difference from saline þ scopolamine. Data are expressed as mean ? S.E.M. N ¼ 6/condition.
R. Galici et al. / Neuropharmacology 56 (2009) 1131–11371134
The results of these studies indicate that JNJ-10181457 improves
learning and reverses the cognitivedeficits induced byscopolamine
in rats. Furthermore, the behavioral effects of JNJ-10181457 are
associated with normalization of ACh neurotransmission release at
a dose that maximally occupies the H3 receptor and results in
significant brain exposure.
In these studies, we first determined the time course of brain
exposure and H3receptor occupancy for JNJ-10181457. A dose of
10 mg/kg was tested based on previous studies (Bonaventure et al.,
2007). A single administration of JNJ-10181457 resulted in
a significant brain exposure and maximal H3receptor occupancy
for at least 90 min. This time frame was chosen to parallel the time
required to evaluate the behavioral and neurochemical effects of
the compound. Specifically, based on these results we evaluated the
same dose of JNJ-10181457 in a translational assay of reference and
working memory requiring a similar time frame for completion. In
the DNMTP task, scopolamine decreased percent correct respond-
ing without affecting the number of trials. These results indicate
that the effects of scopolamine are likely related to reference and
working memory and are consistent with other studies reported in
the literature (Dunnett,1985; Kirkby et al.,1996). There was a trend
for JNJ-10181457 to increase percent correct responding by itself.
However, these effects were not statistically significant. JNJ-
10181457 significantly reversed scopolamine-induced deficits at
a dose that was associated with a robust plasma and brain exposure
and maximally occupied the receptors. Furthermore, the effects of
JNJ-10181457 were similar to donepezil, an acetylcholinesterase
inhibitor. These results are also consistent with the literature. For
example, acetylcholinesterase inhibitors such as tacrine and
donepezil have been shown to reverse the effects of scopolamine in
several behavioral assays (Hancock and Fox, 2004; Witkin and
Nelson, 2004) including translational procedures. For example, in
one study, tacrine reversed the effects of scopolamine in the
delayed match to position (Kirkby et al., 1996). In the same study
however, 10 mg/kg of thioperamide did not significantly reverse
scopolamine-induced deficits. This is also consistent with our
results indicating a partial but not significant effect. It is unlikely
that the lack of efficacy of thioperamide is related to the dose used,
since in this study 10 mg/kg completely reversed imetit-induced
drinking. One possible explanation is that thioperamide is not
a very selective compound. For example it has affinity for the H4
and 5-HT3 receptors (Leurs et al., 1995) and interacts with the
neuronal GABA transporter (Yamamoto et al., 1997). Thus, such
a complex interaction with other receptors might have determined
lack of effects in this procedure. DNMTP is a complex behavioral
procedure that requires a significant cortical ACh participation
(Sloan et al., 2006; Touzani et al., 2007). Thus, another possible
explanation for the lack of effects of thioperamide is that, under the
condition of this and other studies (Kirkby et al., 1996), it did not
sufficiently enhance ACh release in cortical regions to counteract
the scopolamine-induced deficits.
We wanted to extend our results using an additional trans-
lational assay that did not depend on pharmacological manipula-
tions (i.e. scopolamine-induced deficits). Therefore, the effects of
JNJ-10181457 were tested in the reversal learning task. One
advantage of this procedure is that it allows for repeated admin-
istration of compounds and therefore loss or improvement of effi-
cacy can be evaluated over time. Our results indicate that repeated
responding compared to vehicle-treated animals suggesting that it
improves learning. It is highly unlikely that the effects of JNJ-
10181457 are due to better performer rats compared to vehicle-
treated animals, since both groups had a similar rate of acquisition
in the initial phase of the task (i.e. prior to switching the rule). In
addition, when the new rule was imposed, there was a significant
and similar decrease in correct responding for both groups and
Number of trials completed in the DNMTP task.
Saline þ water
Saline þ Scopolamine
Donepezil þ water
Donepezil þ Scopolamine
Thioperamide þ water
Thioperamide þ Scopolamine
JNJ-10181457 þ water
JNJ-10181457 þ Scopolamine
39.1 ? 6.1
32.8 ? 4.1
42.1 ? 4.8
54.1 ? 4.1
47.3 ? 2.5
37.3 ? 4.3
48.6 ? 4.0
40.8 ? 5.4
-60 -300 30 6090 120
Saline + Scopolamine
JNJ-10181457 + Scopolamine
JNJ-10181457 + Water
of ACh Basal Release
Saline + Water
Saline + Scopolamine
Ach AUC (60 min)
Fig. 4. Effects of JNJ-10181457 on ACh release in the cortex. Saline or JNJ-10181457 (10 mg/kg) was administered i.p. Water or scopolamine (0.06 mg/kg) was administered s.c.
30 min later. Percent Change of ACh release is plotted as function of time in panel A. ACh AUC is plotted in panel B. Data are expressed as mean ? S.E.M. N ¼ 6/condition. *p < 0.05
indicates statistical difference from Saline þ Water.
R. Galici et al. / Neuropharmacology 56 (2009) 1131–1137 1135
learning differences emerged only after repeated administration of
JNJ-10181457. Treatment was suspended on the third day because
a significant effect was obtained. Furthermore, discontinuation of
the treatment did not disrupt learning suggesting lack of with-
drawal andrebound effects.
responding was significantly increased in animals previously
treated with JNJ-10181457 compared to vehicle-treated rats. Since
the acquisition curve under the new rule appears to be shifted
leftward, these results indicate that JNJ-10181457 significantly
increased learning. Eventually, six days after discontinuation,
percent correct responding was similar in both groups of animals.
It is difficult to separate the cognitive from the wake-promoting
effects of H3 receptor antagonists. For example, JNJ-10181457
promotes wakefulness in rats up to 6 h and these effects are no
longer present 24 h later (Bonaventure et al., 2007). The wake-
promotingeffects of H3antagonists are characterized bya quiet and
alert waking (i.e. locomotor hyperactivity is absent) and lack of
sleep rebound (Parmentier et al., 2007). This is in contrast with the
effects of psychostimulants such as amphetamine and caffeine
which have wake-promoting effects at doses that increase loco-
motor activity and result in significant sleep rebound, often char-
acterized by an increased non-REM duration, intensity of sleep and
cognitive deficits (Parmentier et al., 2007). Thus, one important
conclusion from the current studies is that the wake-promoting
effects of JNJ-10181457 did not prevent the compound from
improving learning and reversing scopolamine deficits. For
example, any rebound effect orlong lasting wake-promotingeffects
would have likely negatively impacted learning under the condi-
tion of these studies.
Furthermore, H3 antagonists have been shown to promote
wakefulness and vigilance through the action of many neuro-
transmitters including ACh and nor-epinephrine and both could
also ultimately improve cognitive performance (Haas et al., 2008).
However, the relative contribution of H3 mediated neurotrans-
mitter (i.e. histamine vs nor-epinephrine vs ACh) release to each
behavior (i.e. vigilance vs cognition) is not fully understood. We
tested the hypothesis that H3 antagonists improve cognitive
processes via cholinergic systems. Specifically, experiments were
designed to evaluate whether the reversal of scopolamine-induced
cognitive-related deficits was associated with changes in ACh
neurotransmission in the cortex. The mechanism by which H3
antagonists increase ACh release in the cortex is not fully under-
stood. However, H3 receptors are expressed pre-synaptically in
non-histamine-containing neurons (Leurs et al., 1998). Thus, H3
antagonists can increase ACh outflow in areas of the brain rich in
cholinergic neurons such as the cortex. Alternatively, release of
For example, percentcorrect
histamine by H3antagonists could stimulate post-synaptic hista-
mine H1and or H2receptors which are expressed on cholinergic
neurons and, therefore, could increase the release of ACh (Haas
et al., 2008; Pascoli et al., 2009).
The results of our studies indicate that JNJ-10181457 signifi-
cantly increased ACh release when administered alone. Further-
more, scopolamine significantly reduced ACh release and this likely
resulted in the cognitive-related deficits described in this study.
Interestingly, JNJ-10181457 did not significantly reverse scopol-
amine-induced ACh release deficits. However, the co-administra-
tion of JNJ-10181457 and scopolamine was not statistically different
from control (i.e. saline þ water). These results indicate that JNJ-
10181457 normalized ACh neurotransmission in the cortex and
suggest that selective H3non-imidazole antagonists could be very
effective in conditions in which ACh release is decreased as those
found in cognitive disorders such as Alzheimer’s disease, dementia,
autism and schizophrenia. However, alterative non-cholinergic
mechanisms could also account for the procognitive effects
obtained in these studies (Haas et al., 2008; Pascoli et al., 2009;
Dere et al., 2008).
Collectively, JNJ-10181457 improved learning and memory in
rats by selectively blocking H3 receptors and normalizing ACh
neurotransmission. These results provide further in vivo evidence
of the heterologus nature of this receptor system and support the
hypothesis that H3 antagonists may have clinical efficacy in
cognitive-related disorders, especially those in which ACh neuro-
transmission is compromised.
The authors acknowledge the assistance of Kevin Sharp and his
staff at Johnson & Johnson PRD L.L.C.
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% correct responding
Fig. 5. Effects of JNJ-10181457 in the reversal learning task. Cumulative correct choices are plotted as function of time (panel A) for the last training session prior to treatment with
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