Investigation into the efficacy of the acetylcholinesterase inhibitor, donepezil, and novel procognitive agents to induce gamma oscillations in rat hippocampal slices
Introduction
Alzheimer’s Disease (AD) is a devastating disorder characterised by progressive cognitive decline and a host of other behavioural symptoms (Cummings, 2004). Due to the increasing age of the population the prevalence of AD is estimated to quadruple by 2050 to approximately 107 million people worldwide (Brookmeyer et al., 2002). The major neuropathological hallmark in AD is the deposition of extracellular plaques composed of beta-amyloid peptide, believed to be responsible for neurodegeneration and subsequent cognitive decline at least in the early stages of the disease (Tiraboschi et al., 2004). A major area of neuronal cell loss is the cholinergic neuronal population of the nucleus basalis of Meynert (NbM) (Whitehouse et al., 1982), which sends its projections to areas such as the hippocampus and cortex; areas highly associated with memory formation and storage. It is this loss of cholinergic drive which has formed the basis of the ‘cholinergic’ hypothesis of AD (Bartus et al., 1982) and to the subsequent development of acetylcholinesterase (AChE) inhibitors for the treatment of AD. AChE inhibitors, which increase the concentration of acetylcholine at the synapse by preventing its breakdown, are the current gold standards for symptomatic treatment of AD. The most widely prescribed AChE inhibitor is donepezil (Aricept), which has been approved for use in mild-moderate AD and more recently for moderate-severe AD in the US. Clinical data have indicated that donepezil can provide meaningful benefit in AD (Birks and Harvey, 2003, Whitehead et al., 2004), but other analyses suggest little benefit is provided by donepezil compared to placebo (Kaduszkiewicz et al., 2005).
Surprisingly there have been few studies exploring the physiological mechanisms by which donepezil mediates a cognitive enhancing effect. One in vivo study in aged rats examined long-term potentiation (LTP); a phenomenon believed to underlie learning and memory (Bliss and Collingridge, 1993). The authors showed that there was less decay of LTP over 5 days at synapses between the perforant path and dentate gyrus in aged rats treated with donepezil compared with control aged animals (Barnes et al., 2000).
To explore the physiological effects of donepezil further we have assessed the ability of donepezil to induce gamma oscillations in a hippocampal slice preparation. Gamma oscillations represent synchronous network activity which can be recorded in the hippocampus and cortex and which is believed to play a role in higher cognitive function (Bragin et al., 1995, Fell et al., 2001, Herrmann et al., 2009, Singer, 1993). Furthermore, gamma oscillations could be important in regulating synaptic plasticity, such as LTP (Traub et al., 1998) and have been shown to be disrupted in a variety of neuropsychiatric disorders including schizophrenia, Alzheimer’s disease and epilepsy (Herrmann and Demiralp, 2005). Gamma oscillations can be induced in hippocampal or cortical slice preparations by various methods including high-frequency stimulation (Traub et al., 1996), metabotropic glutamate receptor activation (Whittington et al., 1995), nanomolar concentrations of kainate (Cunningham et al., 2003, Hajos et al., 2000), cholinergic receptor activation with carbachol (Buhl et al., 1998, Fisahn et al., 1998) or with the AChE inhibitor physostigmine (Engel et al., 2002). Furthermore, it has been shown that nicotine and the AChE inhibitor, neostigmine, can induce gamma oscillations in vivo (Joosen and van Helden, 2007, Phillips et al., 2007).
Given the ability of cholinergic agonists and AChEs to induce gamma oscillations we investigated the activity of donepezil in a brain slice preparation and compared this to that of physostigmine.
Section snippets
Methods
Brain slices were prepared from CD rats weighing 150–250 g (Charles River, Margate, UK). Animals were humanely killed by overdose of isoflurane via inhalation in accordance with the UK Animal (scientific procedures) act 1986. All efforts were made to minimise the number of animals used. The brain was removed into ice-cold artificial cerebrospinal fluid (aCSF) comprising (in mM): 189 sucrose, 2.5 KCl, 1.2 NaH2PO4, 26 NaHCO3, 10 glucose, 0.1 CaCl2 and 5 MgCl2 pH 7.4. Horizontal brain slices
Data analysis
All data were captured using Spike 2 software (Cambridge Electronic Design). Power spectra were generated from 60 s epochs of data by means of fast Fourier transforms. The power and area under curve (AUC) in the gamma frequency range (20–80 Hz) were used to quantify the strength of gamma frequency oscillations. Data were normalised to a stable baseline obtained prior to compound application. Unless otherwise stated data are presented as means ± standard errors of the means (SEM). Statistical
Drugs
Drugs were bath applied to the hippocampal slice by addition to the perfusion medium. Donepezil, prucalopride, SL65.0155 and 77-LH-28-1 were synthesised at GlaxoSmithKline. WAY-208466 was synthesised by Syngene International Ltd. Atropine and bicuculline methiodide were obtained from Sigma-Aldrich Company Ltd. Physostigmine hemisulphate was obtained from Tocris-Cookson Ltd, UK.
Results
Gamma oscillatory activity can be induced in hippocampal slices by various mechanisms, including activation of muscarinic acetylcholine receptors using the cholinergic agonist, carbachol (Fisahn et al., 1998) and the AChE inhibitor physostigmine (Engel et al., 2002). For this reason we assessed whether the AChE inhibitor and gold standard treatment for AD, donepezil, was also capable of inducing gamma frequency oscillations in region CA3 of rat hippocampal slices. Bath application of donepezil
Discussion
We report here that donepezil, an AChE inhibitor approved for symptomatic treatment of patients with mild-moderate AD, is capable of inducing gamma oscillatory activity in rat hippocampal slices. This supports previous work showing that the cholinergic agents including carbachol, physostigmine and nicotine can induce gamma oscillations (Engel et al., 2002, Fisahn et al., 1998, Phillips et al., 2007). Given the proposed role of gamma oscillations in cognitive function (Herrmann et al., 2009), we
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