RG3487, a novel nicotinic α7 receptor partial agonist, improves cognition and sensorimotor gating in rodents.

Neuronal nicotinic α7 acetylcholine receptors (α7nAChRs) are expressed primarily in the brain and are implicated in modulating many cognitive functions (e.g., attention, working and episodic memory). Not surprisingly, much effort has been committed to the development of molecules acting at α7nAChRs as potential therapies for a variety of central nervous system diseases (e.g., Alzheimer's). N-[(3S)-1-azabicyclo[2.2.2]oct-3-yl]-1H-indazole-3-carboxamide hydrochloride (RG3487) binds potently to the human α7nAChR (K(i) = 6 nM), in which it acts as a partial agonist (63-69% of acetylcholine) as assessed by whole-cell patch-clamp recordings in both oocytes and QM7 cell lines. RG3487 activates human α7nAChRs with an EC(50) of 0.8 µM (oocytes) and 7.7 µM (QM7 cells). RG3487 also exhibits antagonist properties at the serotonin 3 receptor [IC(50) = 2.8 nM (oocytes), 32.7 nM (N1E-115 cells)]. In vivo, RG3487 improved object recognition memory in rats after acute [minimally effective dose (MED) 1.0 mg/kg p.o.] or repeated (10 day) administration at brain and plasma concentrations in the low-nanomolar range. Spatial learning deficits in age-impaired rats were reversed after RG3487 administration (MED: 0.03 mg/kg i.p.) as evaluated in the Morris water maze task. In the prepulse inhibition (PPI) of startle model of sensorimotor gating, RG3487 improved apomorphine-induced deficits in PPI performance (MED: 0.03 mg/kg i.p.) and reversed phencyclidine-induced impairments in an attentional set-shifting model of executive function (MED: ≤0.03 mg/kg i.p.). Cumulative evidence from these studies indicates RG3487 is a novel and potent α7nAChR partial agonist that improves cognitive performance and sensorimotor gating.

Hippocampal expression analyses reveal selective association of immediate-early, neuroenergetic, and myelinogenic pathways with cognitive impairment in aged rats.

Although expression of some genes is known to change during neuronal activity or plasticity, the overall relationship of gene expression changes to memory or memory disorders is not well understood. Here, we combined extensive statistical microarray analyses with behavioral testing to comprehensively identify genes and pathways associated with aging and cognitive dysfunction. Aged rats were separated into cognitively unimpaired (AU) or impaired (AI) groups based on their Morris water maze performance relative to young-adult (Y) animals. Hippocampal gene expression was assessed in Y, AU, and AI on the fifth (last) day of maze training (5T) or 21 d posttraining (21PT) and in nontrained animals (eight groups total, one array per animal; n = 78 arrays). ANOVA and linear contrasts identified genes that differed from Y generally with aging (differed in both AU and AI) or selectively, with cognitive status (differed only in AI or AU). Altered pathways/processes were identified by overrepresentation analyses of changed genes. With general aging, there was downregulation of axonal growth, cytoskeletal assembly/transport, signaling, and lipogenic/uptake pathways, concomitant with upregulation in immune/inflammatory, lysosomal, lipid/protein degradation, cholesterol transport, transforming growth factor, and cAMP signaling pathways, primarily independent of training condition. Selectively, in AI, there was downregulation at 5T of immediate-early gene, Wnt (wingless integration site), insulin, and G-protein signaling, lipogenesis, and glucose utilization pathways, whereas Notch2 (oligodendrocyte development) and myelination pathways were upregulated, particularly at 21PT. In AU, receptor/signal transduction genes were upregulated, perhaps as compensatory responses. Immunohistochemistry confirmed and extended selected microarray results. Together, the findings suggest a new model, in which deficient neuroenergetics leads to downregulated neuronal signaling and increased glial activation, resulting in aging-related cognitive dysfunction.

The slow afterhyperpolarization in hippocampal CA1 neurons covaries with spatial learning ability in aged Fisher 344 rats.

Rodents commonly exhibit age-related impairments in spatial learning tasks, deficits widely thought to reflect cellular or synaptic dysfunction in the hippocampus. Using whole-cell recordings, we examined the afterhyperpolarization (AHP) in CA1 pyramidal cells in hippocampal slices from young (4-6 months of age) and aged (24-26 months of age) Fisher 344 male rats that had been behaviorally characterized in the Morris water maze. The slow AHP (sAHP) recorded from learning-impaired aged rats (AI) was significantly larger than that seen in either age-matched unimpaired rats or young controls. Among aged rats, sAHP amplitude was inversely correlated with both acquisition and probe performance in the water maze. Action potential parameters among the three groups were similar, except for spike accommodation, which was more pronounced in the AI group. Intracellular application of the cAMP analog 8-CPT-cAMP suppressed the sAHP but failed to reveal any age- or performance-related differences in the medium AHP. 8-CPT-cAMP abolished the age-related difference in spike accommodation, whereas instantaneous firing frequency was unchanged. Calcium spikes were of similar amplitude in all three groups but were broader and had significantly larger tails in aged rats; these age-related changes could be mimicked in young neurons after exposure to BayK8644. The calcium spike among aged rats correlated with task acquisition in the maze but, unlike the sAHP, failed to correlate with probe performance. This is the first demonstration that sAHP amplitude covaries with spatial learning ability in aged rats, implying that CA1 excitability strongly influences certain aspects of cognitive function. Our findings also indicate that multiple processes, in addition to elevated calcium influx, conspire to induce cognitive decline during aging.

Theta-frequency synaptic potentiation in CA1 in vitro distinguishes cognitively impaired from unimpaired aged Fischer 344 rats.

Hippocampal-dependent learning and memory deficits have been well documented in aging rodents. The results of several recent studies have suggested that these deficits arise from weakened synaptic plasticity within the hippocampus. In the present study, we examined the relationship between hippocampal long-term potentiation (LTP) in vitro and spatial learning in aged (24-26 months) Fischer 344 rats. We found that LTP induced in the CA1 region using theta-frequency stimulation (5 Hz) is selectively impaired in slices from a subpopulation of aged rats that had shown poor spatial learning in the Morris water maze. LTP at 5 Hz in aged rats that did not show learning deficits was similar to that seen in young (4-6 months) controls. We also found that 5 Hz LTP amplitude strongly correlated with individual learning performance among aged rats. The difference in 5 Hz LTP magnitude among aged rats was not attributable to an altered response to 5 Hz stimulation or to differences in the NMDA receptor-mediated field EPSP. In addition, no performance-related differences in LTP were seen when LTP was induced with 30 or 70 Hz stimulation protocols. Finally, both 5 Hz LTP and spatial learning in learning-impaired rats were enhanced with the selective muscarinic M2 antagonist BIBN-99 (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). These findings reinforce the idea that distinct types of hippocampal LTP offer mechanistic insight into age-associated cognitive decline.

Chronic treatment with the antidepressant amitriptyline prevents impairments in water maze learning in aging rats.

Increasing evidence links chronically elevated glucocorticoid levels and cognitive impairments in a subpopulation of aged rodents and humans. Antidepressant drugs improve hypothalamic-pituitary-adrenal axis feedback regulation and reduce plasma glucocorticoid levels. Decreasing the cumulative lifetime exposure to glucocorticoid excess by long-term exposure to antidepressants may prevent the emergence of cognitive impairments in aged rats. To test this hypothesis, we treated middle-aged male Lister hooded rats (16 months) with amitriptyline until they were 24 months of age, and their cognitive function was assessed in the water maze. Performance in the spatial learning task declined significantly with aging (p < 0.01), with 33% of aged controls showing poorer (<2.5 SD) probe test performance than young controls. Amitriptyline treatment from midlife preserved water maze performance with aging (p < 0.01 compared with aged controls) and significantly (p < 0.01) reduced the proportion of poor performers (7%). Measures of anxiety-related behaviors in the elevated plus-maze were significantly (p < 0.05) decreased in the aged rats after amitriptyline. Furthermore, evening plasma corticosterone levels were reduced (30% decrease; p < 0.01 compared with aged controls) after 6 months of amitriptyline. These data suggest that long-term treatment with amitriptyline decreases the prevalence of cognitive impairment in aged rats and that this may, in part, be a consequence of reduced plasma corticosterone levels and reduced anxiety.

Water maze testing to identify compounds for cognitive enhancement.

The water maze task is widely used to evaluate spatial learning and memory in rodents. The basic paradigm requires an animal to swim in a pool until it finds a hidden escape platform. The animals learn to find the platform using extra-maze cues and, after several training trials, are able to swim directly to it from any starting location. Memory for the platform location is assessed by examining swimming behavior with the platform removed from the maze, while sensory, motor and motivational aspects of the task can be examined by making the platform visible to the animals. Described in this unit is the use of the water maze to identify rats with age-related spatial learning and memory impairments. The efficacy of potential pharmacological treatments for alleviating these deficits is then evaluated. This assay provides a means for studying the neurobiology of spatial learning and memory, and to identify potential pharmacotherapies for treating memory-impaired humans. While the use of aged rats is described in this unit, the protocol can also be employed for compound screening with other rodent models that have spatial learning and memory impairments, such as transgenic mouse models of Alzheimer's disease.

Genomic and proteomic strategies to identify novel targets potentially involved in learning and memory.

The hippocampus is a crucial player across several learning and memory domains, and is highly vulnerable to alterations during aging. Several products of neurotransmitter genes and neuromodulator genes (which play important parts in mediating and maintaining cognitive ability as a function of age) are expressed in hippocampal formation. However, they represent only a small fraction of genes known to be expressed in this region. We review here recent studies on the use of cDNA microarray and proteomic approaches to uncover novel genes and pathways that might be involved in cognitive processes in the aged brain. We and other authors have demonstrated major individual differences in cognitive ability in rats of a similar age, thereby making it possible to directly compare gene products expressed as a function of age and cognitive status. Examples of the possible functional role of some of these genes (e.g. transthyretin, quinone reductase 2) and gene products are discussed.