Mice that express the mutant form of the human amyloid precursor gene associated with early-onset familial Alzheimer&#x27;s disease demonstrate memory deficits and amyloid plaques. We show here that ciproxifan, a prototypical antagonist of H~3~-type histamine receptors, alleviates two types of learning and memory impairments in such mice. These data support the idea that modulation of H~3~ receptors represents a viable therapeutic strategy in the treatment of Alzheimer&#x27;s disease

Mark E. Bardgett

Molly S. Griffith

Natasha N. Davis

English

Nature Precedings

  Bardgett, Mark E. 1 
 
 
 
 
 
Ciproxifan, an H3 Receptor Antagonist, Improves Learning and Memory in the 
APP Mouse Model of Alzheimer’s Disease. 
 
Mark E. Bardgett, Natasha N. Davis, & Molly S. Griffith 
 
Department of Psychological Science, Northern Kentucky University 
 
Corresponding author: Mark E. Bardgett, Ph.D., Department of Psychological 
Science, Northern Kentucky University, 1 Nunn Drive, Highland Heights, KY  
41075, Phone: 859-572-5591, FAX: 859-572-6085 
 
Running Head: H3 antagonists and Alzheimer’s Disease 
 
 
 
  Bardgett, Mark E. 2 
 
Mice that express the mutant form of the human amyloid precursor gene 
associated with early-onset familial Alzheimer’s disease demonstrate 
memory deficits and amyloid plaques. We show here that ciproxifan, a 
prototypical antagonist of H3-type histamine receptors, alleviates two types 
of learning and memory impairments in such mice. These data support the 
idea that modulation of H3 receptors represents a viable therapeutic 
strategy in the treatment of Alzheimer’s disease.  
 
Over the past decade, preclinical research has identified the H3 histamine 
receptor as a possible target for cognitive-enhancing drugs (1, 2). The H3 
receptor exists as a presynaptic autoreceptor that is expressed in relatively high 
densities in brain regions associated with memory function, such as the frontal 
cortex and hippocampus (3). Antagonism of the receptor leads to the release of 
histamine as well as neurotransmitters involved in learning and memory, such as 
acetylcholine and dopamine (4 - 6). Moreover, H3 antagonists can generate 
electrical activity in the brain that predicts new learning (7). On a behavioral level, 
drugs that act as H3 antagonists, such as the prototypical imidazole-containing 
compounds, thioperamide and ciproxifan, have been shown to improve memory 
function in several tasks – in normal rats and mice, as well as in animals treated 
with anti-cholinergic or anti-glutamatergic drugs (8 - 11). 
  Bardgett, Mark E. 3 
The ability of H3 antagonists to enhance memory in normal animals and in 
pharmacological models of memory impairment raises the possibility that such 
compounds may represent an effective treatment strategy for Alzheimer’s 
disease. As a way of addressing this possibility, we tested the effects of the H3 
antagonist, ciproxifan (8), on learning and memory deficits observed in the 
amyloid precursor protein (APP) transgenic mouse model of Alzheimer’s disease. 
Developed by Hsiao and colleagues (12), APP mice express a mutant form of the 
human APP gene associated with early-onset, familial Alzheimer’s disease. 
These mice exhibit a phenotype that includes deficits in spatial learning and 
memory and object recognition (12,13), as well as the formation of amyloid 
plaques with increasing age.  
In the first study, we tested the effects of ciproxifan on spatial memory and 
locomotor activity in APP mice and wild-type (WT) littermates of both genders at 
12-14 months of age. Approximately half of the mice of each genotype received 
intraperitoneal injections of ciproxifan (3 mg/kg) and the other half received 
injections of saline. The dose of ciproxifan chosen for study was based on 
previous research demonstrating the ability of this dose to improve memory in 
normal rats (9). Mice received daily injections over the course of three weeks and 
on behavioral testing days received such injections 30 minutes prior to testing. 
During the second week of treatment, all mice were tested for spatial 
learning in the swim maze task. Ciproxifan improved performance in APP mice 
over the course of the training trials (Fig. 1). There was a statistical interaction 
between ciproxifan treatment and APP genotype (Interaction term: F (1, 34) = 
  Bardgett, Mark E. 4 
6.93, p < .01) on the time to find the platform measure. APP mice treated with 
saline took longer to find the hidden platform in the test pool relative to wild-type 
mice treated with saline on days 1, 4, & 5 of training (Fig. 1a). On the latter two 
days, APP mice treated with ciproxifan took less time to find the platform than 
APP mice treated with saline (Fig. 1a). Also during the training trials, there was a 
treatment by genotype interaction (Interaction term: F (1, 34) = 4.46, p < .04) as 
well as an overall training day effect (F (4, 136) = 9.7, p < .0001) on the average 
distance from the platform measure. During the last four days of training, APP 
mice treated with saline tended to swim farther away from the platform in 
comparison to APP mice treated with ciproxifan and WT mice treated with saline 
(Fig. 1b). It should also be noted that there was trend towards a significant 
treatment x genotype interaction for the distance travelled before finding the 
platform (Interaction term: F (1, 34) = 4.46, p = .11 (two-tailed))(Supplemental 
Figure 1a). Finally, ciproxifan increased swimming speed in APP mice 
(Interaction term: F (1, 34) = 6.54, p < .02)(Supplemental Figure 1b). In this 
case, the most consistent group difference over the last four days of training was 
between the APP mice treated with ciproxifan and APP mice treated with saline. 
Unlike the results for the other measures, APP mice treated with saline did not 
differ from WT mice treated with saline during the last four days of training. This 
distinct pattern of data suggests that group differences in swim speed do not 
account completely for the significant differences observed on the other 
measures.   
  Bardgett, Mark E. 5 
One hour after the final training session, all mice were placed in the test 
pool for one minute without the platform. During this probe trial, APP mice treated 
with ciproxifan made as many crossings of the former platform site as did the WT 
mice treated with saline and significantly more than the APP mice treated with 
saline (Interaction term: F (1, 34) = 4.68, p < .04)(Fig. 1c). Moreover, when the 
amount of time spent in an area of the pool that included the former platform was 
measured, the same pattern of results emerged. APP mice treated with 
ciproxifan spent as much time as each group of WT mice in the zone that 
formerly contained the platform. In contrast to these groups, APP mice treated 
with saline spent significantly less time in this area (Interaction term: F (1, 34) = 
6.45, p < .02)(Fig. 1d). Mice were also tested 48 hours after the last training trial 
and the effects of drug and genotype were not significant at that time 
(Supplemental Fig. 2). 
A separate cohort of 12 – 14 month old APP and WT mice were tested on 
a novel object recognition (NOR) task. Ciproxifan or saline was injected into the 
mice 30 minutes prior to testing. Ciproxifan improved NOR in APP mice 
(Interaction term: F (1, 20) = 11.8, p < .003) (Fig. 2a). APP mice spent 
significantly less time exploring a novel object relative to a familiar one in 
comparison to all other groups, including the APP mice treatment with ciproxifan. 
The preference of this latter group of mice for the novel object did not differ from 
the WT mice treated with saline. These results were not due to changes in 
general exploration since neither genotype or drug treatment altered the total 
time spent exploring both objects during testing (Fig. 2b).  
  Bardgett, Mark E. 6 
Finally, we assessed locomotor activity in the same cohort of APP and WT 
mice used in the swim maze test. This testing was performed one week prior to 
swim maze testing. Mice were tested for one hour a day for five days. APP mice 
treated with saline were significantly more active than WT mice treated with 
saline (Genotype x treatment x time interaction: F (11, 385) = 2.0, p < .03)(Fig. 
2c). Ciproxifan reversed this effect. APP mice treated with ciproxifan were 
significantly less active than the APP mice treated with saline and were not 
different from the WT mice treated with saline during the first 25 minutes of 
testing.  
These experiments are the first to demonstrate the ability of an H3 
antagonist to alleviate memory deficits and hyperactivity in a transgenic mouse 
model of Alzheimer’s disease. The capacity for ciproxifan to improve cognitive 
and behavioral outcomes across multiple tests provides robust support for the 
pursuit of H3 antagonism as a therapeutic strategy in the treatment of Alzheimer’s 
disease. Current palliative treatments for Alzheimer’s disease include 
acetylcholinesterase inhibitors and NMDA antagonists, yet these treatments 
possess only limited efficacy and significant side effects. Receptor binding data 
has shown that H3 receptor densities are preserved in the brains of people with 
Alzheimer’s despite disease progression (5), indicating that the target for H3 
antagonists remains a viable one throughout the disease process. Therefore, 
antagonism of H3 receptors may represent an alternative pathway to cognitive 
enhancement or at least an approach that can be coupled with current 
treatments (14).  
  Bardgett, Mark E. 7 
It is likely that ciproxifan’s efficacy in the current study is related to its 
ability to enhance neurotransmitter release in the frontal cortex and 
hippocampus, and to generate electrophysiological activity predictive of learning. 
It will be important for future studies to test these ideas as well as to determine if 
longer term treatment with H3 antagonists can modify pathophysiological 
processes (e.g. plaque formation, synapse loss) observed in the APP mouse 
model (15). 
 
ACKNOWLEDGEMENTS: This work was supported by National Center for 
Research Resources Grant 2P20 RR16481 and National Institute of Mental 
Health Grant R15 MH076788 to M.E.B. We appreciate the assistance of Dr. 
Patrick J. Schultheis with the genotyping of the mice. 
 
AUTHOR CONTRIBUTIONS: M.E.B. conceptualized and designed the project 
and wrote the manuscript. N.N.D. helped in designing the object recognition 
experiment and conducted the experiment. M.S.G. helped to design the entire 
project, genotyped the mice, conducted the swim maze and amyloid plaque 
experiments, and helped to write the manuscript.  
 
 
1. Esbenshade, T.A., Fox, G.B., & Cowart, M.D. Mol Interv, 6, 77-88 (2006).  
  Bardgett, Mark E. 8 
2. Bonaventure, P. et al. Biochem Pharmacol 73, 1084-1096 (2007).  
3. Pillot, C. et al. Neuroscience 114, 173-193 (2002). 
4. Fox, G.B. et al. J Pharmacol Exp Ther 313, 176-190 (2005). 
5. Medhurst, A.D. et al. J Pharmacol Exp Ther 321, 1032-1045 (2007). 
6. Ligneau, X. et al. J Pharmacol Exp Ther 320, 365-375 (2007).  
7. Hajós, M. Siok, C.J., Hoffmann, W.E., Li, S. & Kocsis, B. J Pharmacol Exp 
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8. Ligneau, X. et al. J Pharmacol Exp Ther 287, 658-666 (1998). 
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10. Bernaerts, P., Lamberty, Y., & Tirelli, E. Behav Brain Res 154, 211-219 
(2004).  
11. Galici, R. et al. Neuropharmacology 56, 1131-1137 (2009). 
12. Hsiao, K. et al. Science 274, 99-102 (1996). 
13. Taglialatela, G., Hogan, D., Zhang, W.R., & Dineley, K.T. Behav Brain Res 
200, 95-99 (2009). 
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15. Dong, H. Martin, M.V., Chambers, S., & Csernansky, J.G. J Comp Neurol 
10, 311-321 (2007). 
 
  Bardgett, Mark E. 9 
 
 
Figure legends: 
Figure 1 Ciproxifan alleviates learning deficits exhibited by the APP mice in the 
swim maze task. (a) APP mice treated with saline took longer to find the escape 
platform than saline-treated WT mice on days 1, 4, & 5 of training, as indicated 
by *, and APP mice treated with ciproxifan (3.0 mg/kg) on days 4 & 5 of testing, 
as indicated by #. (b) APP mice swam, on average, a greater distance away from 
the platform in comparison to saline-treated WT mice on days 2, 3, & 5 of 
training, as indicated by *, and APP mice treated with ciproxifan on training days 
2 – 5, as indicated by the #. There was an overall training day effect, indicative of 
learning across training days. (c) APP mice treated with saline displayed fewer 
crossings of the former platform location during the first probe trial in comparison 
to the APP mice treated with ciproxifan as indicated by #. (d) APP mice treated 
with saline spent less time near the former location of the platform during the 
probe trial in comparison to all other groups including the APP mice treated with 
ciproxifan as indicated by +. All data represent means + s.e.m. All symbols 
represent statistically significant differences (p < .05, two-tailed).  
 
Figure 2 Ciproxifan alleviates the novel object recognition deficit and 
hyperactivity observed in APP mice. (a) APP mice treated with saline spent less 
  Bardgett, Mark E. 10 
time exploring a novel object relative to the time spent exploring a familiar one 
when compared to all other groups, including the APP mice treated with 
ciproxifan as indicated by +. (b) The total time spent exploring both objects 
during the test trial did not differ between groups. (c) APP mice were significantly 
more active during the first 25 minutes of activity testing relative to all of the other 
groups, including the APP mice treated with ciproxifan as indicated by +. All data 
represent means + s.e.m. All symbols represent statistically significant 
differences (p < .05, two-tailed).  




Ciproxifan, an H~3~ Receptor Antagonist, Improves Learning and Memory in the APP Mouse Model of Alzheimer&#x27;s Disease

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Ciproxifan, an H~3~ Receptor Antagonist, Improves Learning and Memory in the APP Mouse Model of Alzheimer's Disease

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Ciproxifan, an H~3~ Receptor Antagonist, Improves Learning and Memory in the APP Mouse Model of Alzheimer&#x27;s Disease

Abstract

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Ciproxifan, an H~3~ Receptor Antagonist, Improves Learning and Memory in the APP Mouse Model of Alzheimer's Disease