Research reportLong-term effects of BIBN-99, a selective muscarinic M2 receptor antagonist, on improving spatial memory performance in aged cognitively impaired rats
Introduction
Alzheimer’s Disease (AD) is the most widely recognized age-related disease that results in cognitive impairment. A prominent neuropathological feature of AD is neuronal loss in the basal forebrain, resulting in reduced cholinergic input to the hippocampus and neocortex (for recent reviews, see [1], [2], [14], [37]). Several animal models have been adapted to study the cholinergic decline that characterizes AD [4], [10], [13]. Our laboratory has demonstrated that alterations in the cholinergic system occur simply as a function of age in male Long–Evans rats, and that these alterations correlate with cognitive ability [1], [28], [32]. Using the Morris water maze task to evaluate spatial learning and memory, 24–25-month-old rats can be identified as either aged cognitively impaired (AI) or aged cognitively unimpaired (AU) relative to 6-month-old (young adult) animals (for details, see [17], [36]). One of the major findings is that degeneration of spatial learning and memory is not an inevitable consequence of aging. Indeed, the preservation of cognitive function in aged rats, like that in aged humans, shows a great deal of individual variability. Behavioral screening of more than 800 animals over the last decade has revealed spatial memory impairments in 25–30% of aged (24-month-old) Long–Evans rats while 36–40% were found to be unimpaired [16], [17], [32], [35], [36], [46]. Other laboratories have also reported individual differences amongst populations of aged rats, with the percentages of impaired animals varying as a function of both the strain as well as the sex of the animals [12], [23], [38], [42], [51], [52].
While some muscarinic agonists, such as oxotremorine and carbamylcholine (carbachol), decrease endogenous or radiolabeled ACh release [2], [19], [33], non-selective antagonists (atropine and scopolamine) and purported muscarinic M2 antagonists (AFDX 116, AFDX 384, BIBN-99, and SCH 72788) stimulate the release of ACh from cortical, hippocampal, and striatal areas in the rat brain [18], [19], [31], [32], [41]. Lesions of the basal forebrain or destruction of cholinergic neurons result in decreased numbers of muscarinic M2 binding sites in cortex and hippocampus, suggesting that a proportion of these receptors may be located on pre-synaptic cholinergic terminals [6], [20], [25]. This observation is consistent with the finding that the muscarinic M2 gene and its receptor protein are expressed in the basal forebrain in a distribution pattern that is similar to that of cholinergic neurons [20], [30], [34], [47]. Accordingly, we and others have hypothesized that the selective blockade of pre-synaptically located M2 autoreceptors could increase ACh release into the synaptic cleft and thereby enhance cognitive performance [8], [10], [30], [31].
Although many studies have documented a decline in spatial memory performance in the rat with increased age, there have only been a handful of studies testing the ability of rats to retain a learned task over a long period of time. Two separate studies [7], [26] describe reference memory (i.e. the location of the platform) being retained for at least 14 weeks between testing and re-testing in the Morris water maze. In another study, Van Groen et al. [44] demonstrated that rats that were trained to find the platform in the Morris water maze at 12 months of age displayed significantly enhanced performance compared to age-matched controls (no pre-training), when re-tested at 24 months of age. Somehow, the pre-training experience in the repeated acquisition paradigm assisted the rat in remembering for long periods of time. What mechanism may underlie this long-term memory? Could long-term memory be induced pharmacologically? In the present studies, we examine a selected population of aged rats that display poor reference memory performance, the AI rat, and study long-term memory effects of therapeutic drug treatment. We administered the muscarinic M2 receptor antagonist, 5,11-dihydro-8-chloro-11-[[4-[3-[(2,2-dimethyl-1-oxopentyl)ethylamino]propyl]-1-piperidinyl]acetyl]-6H-pyrido[2,3-b][1,4]benzodiazepin-6-one (BIBN-99), a compound known to enhance reference memory performance in the AI rat to an optimal level [32]. In this experiment, however, we ceased drug application after 3 days and examined how long the enhanced memory performance could endure. Next, we delayed the onset of the BIBN-99 treatment for 8 weeks and assessed water maze performance. If the AI animal’s memory for the platform location was already compromised, as witnessed following initial screening, then allowing a long interval of time to pass before pharmacological intervention was to begin might allow for memory decay to occur and potentially decrease the likelihood that reference memory could still be enhanced in these animals.
Section snippets
Animals
Male Long–Evans rats (retired breeders) were obtained from Charles River (St. Constant, Que., Canada) at 12 months of age and were housed in our vivarium for 12–13 months before the experiments began. Young rats were obtained at 3–4 months of age and were housed for 2–3 months prior to behavioral testing. The animals were housed in groups of three ( cm polycarbonate cages) and maintained on a 12 h/12 h light/dark schedule, with ad libitum access to food (Purina Lab Chow) and water. At
Results
Aged (24–25 months old) male Long–Evans rats were screened in the spatial version of the Morris water maze task and were classified into groups based upon their performance as described in Section 2. A significant group by days interaction was observed for both latency to platform (F(8,472)=7.045, P<0.0001) and for distance traveled (F(8,472)=4.4, P<0.0001); post hoc comparisons indicated that the AI rats were significantly impaired at finding the hidden platform on days 2, 3, 4, and 5 of
Discussion
Administration of BIBN-99 improved spatial memory in AI rats. The effect appeared very rapidly, even when drug administration was delayed for 2 months following the initial behavioral screening. Additionally, the improvement induced by BIBN-99 was enduring, outlasting the treatment period by several weeks.
In the first study, improved water maze performance was noted on the first day (day 1) following BIBN-99 treatment (Fig. 2). In fact, performance was significantly enhanced on the first trial
Conclusion
The underlying mechanism(s) for BIBN-99 induced long-term memory enhancement remains largely unknown. It may be mediated by the cholinergic system as eluded to above. M2 receptor immunoreactivity has been found to be co-localized on acetylcholinesterase-rich neurons in hippocampal, cortical and basal forebrain regions [21], [22], [39]. However, other neurotransmitter systems may also be responsible for mediating these effects as M2 receptors have been described on gamma amino butyric acid
Acknowledgements
This work was supported by research grants from the Canadian Institute for Health Research (CIHR) to R.Q. and M.J.M. Both R. Quirion’s research scholarship and W.B. Rowe’s post-doctoral fellowship were supported by Le Fonds de la Recherche en Santé du Québec. J.-P. O’Donnell was supported by a studentship award from CIHR. D. Pearson was supported by an NSERC studentship award. The authors would also like to thank the animals care staff from McGill University and the Douglas Hospital Research
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