Aberrant Cortical Event-Related Potentials During Associative Learning in Rat Models for Presymptomatic Stages of Alzheimer's Disease.

Trace eyeblink conditioning is a hippocampus-dependent associative learning paradigm which is impaired in patients with Alzheimer's disease (AD) and animal AD models. Learning in this paradigm accompanies changes in oscillatory activity in forebrain regions, some of which are loci of pathogenic changes in prodromal AD stages. These observations motivated us to examine how cortical event-related potentials (ERPs) during this paradigm are affected by two features of the asymptomatic, AD-related brain abnormality, entorhinal tau accumulation and mild cholinergic deficit. Adult rats received viral overexpression of P301L mutant human tau in the entorhinal cortex, low-dose scopolamine treatment, or both. Electroencephalograms were recorded with epidural electrodes on the surface of the frontal, parietal, and temporal cortices during differential and reversal trace eyeblink conditioning. All rats developed conditioned responses to one of two stimuli (auditory or visual) paired with mild eyelid shock (CS+), but not to the other stimulus presented alone (CS-). They were also able to adjust the response when the stimulus contingency was reversed. With learning, the amplitude of several ERP components in the frontal and temporal cortices came to differentiate the CS+ from CS-. Scopolamine affected the learning-related change in temporal P2 and other learning-unrelated components in three regions. Entorhinal tau overexpression primary affected the amplitude of temporal visual ERPs and learning-unrelated frontal and temporal auditory ERP components. The double manipulation only affected two components of temporal auditory ERPs. Thus, cortical ERPs during differential associative learning are sensitive to asymptomatic brain abnormality associated with AD.

Entorhinal tau pathology disrupts hippocampal-prefrontal oscillatory coupling during associative learning.

A neural signature of asymptomatic preclinical Alzheimer's disease (AD) is disrupted connectivity between brain regions; however, its underlying mechanisms remain unknown. Here, we tested whether a preclinical pathologic feature, tau aggregation in the entorhinal cortex (EC) is sufficient to disrupt the coordination of local field potentials (LFPs) between its efferent regions. P301L-mutant human tau or green fluorescent protein (GFP) was virally overexpressed in the EC of adult rats. LFPs were recorded from the dorsal hippocampus and prelimbic medial prefrontal cortex while the rats underwent trace eyeblink conditioning where they learned to associate 2 stimuli separated by a short time interval. In GFP-expressing rats, the 2 regions strengthened phase-phase and amplitude-amplitude couplings of theta and gamma oscillations during the interval separating the paired stimuli. Despite normal memory acquisition, this learning-related, inter-region oscillatory coupling was attenuated in the tau-expressing rats while prefrontal phase-amplitude theta-gamma cross-frequency coupling was elevated. Thus, EC tau aggregation caused aberrant long-range circuit activity during associative learning, identifying a culprit for the neural signature of preclinical AD stages.

Enhancing Prefrontal Neuron Activity Enables Associative Learning of Temporally Disparate Events.

The ability to link events that are separated in time is important for extracting meaning from experiences and guiding behavior in the future. This ability likely requires the brain to continue representing events even after they have passed, a process that may involve the prefrontal cortex and takes the form of sustained, event-specific neuron activity. Here, we show that experimentally increasing the activity of excitatory neurons in the medial prefrontal cortex (mPFC) enables rats to associate two stimuli separated by a 750-ms long temporal gap. Learning is accompanied by ramping increases in prefrontal theta and beta rhythms during the interval between stimuli. This ramping activity predicts memory-related behavioral responses on a trial-by-trial basis but is not correlated with the same muscular activity during non-memory conditions. Thus, the enhancement of prefrontal neuron excitability extends the time course of evoked prefrontal network activation and facilitates the formation of associations of temporally disparate, but correlated, events.

Cholinergic, but not NMDA, receptors in the lateral entorhinal cortex mediate acquisition in trace eyeblink conditioning.

Anatomical and electrophysiological studies collectively suggest that the entorhinal cortex consists of several subregions, each of which is involved in the processing of different types of information. Consistent with this idea, we previously reported that the dorsolateral portion of the entorhinal cortex (DLE), but not the caudomedial portion, is necessary for the expression of a memory association between temporally discontiguous stimuli in trace eyeblink conditioning (Morrissey et al. (2012) J Neurosci 32:5356-5361). The present study examined whether memory acquisition depends on the DLE and what types of local neurotransmitter mechanisms are involved in memory acquisition and expression. Male Long-Evans rats experienced trace eyeblink conditioning, in which an auditory conditioned stimulus (CS) was paired with a mildly aversive electric shock to the eyelid (US) with a stimulus-free interval of 500 ms. Immediately before the conditioning, the rats received a microinfusion of neuroreactive substances into the DLE. We found that reversible inactivation of the DLE with GABAA receptor agonist, muscimol impaired memory acquisition. Furthermore, blockade of local muscarinic acetylcholine receptors (mACh) with scopolamine retarded memory acquisition while blockade of local NMDA receptors with APV had no effect. Memory expression was not impaired by either type of receptor blocker. These results suggest that the DLE is necessary for memory acquisition, and that acquisition depends on the integrity of local mACh receptor-dependent firing modulation, but not NMDA receptor-dependent synaptic plasticity.

Unilateral lateral entorhinal inactivation impairs memory expression in trace eyeblink conditioning.

Memory in trace eyeblink conditioning is mediated by an inter-connected network that involves the hippocampus (HPC), several neocortical regions, and the cerebellum. This network reorganizes after learning as the center of the network shifts from the HPC to the medial prefrontal cortex (mPFC). Despite the network reorganization, the lateral entorhinal cortex (LEC) plays a stable role in expressing recently acquired HPC-dependent memory as well as remotely acquired mPFC-dependent memory. Entorhinal involvement in recent memory expression may be attributed to its previously proposed interactions with the HPC. In contrast, it remains unknown how the LEC participates in memory expression after the network disengages from the HPC. The present study tested the possibility that the LEC and mPFC functionally interact during remote memory expression by examining the impact of pharmacological inactivation of the LEC in one hemisphere and the mPFC in the contralateral hemisphere on memory expression in rats. Memory expression one day and one month after learning was significantly impaired after LEC-mPFC inactivation; however, the degree of impairment was comparable to that after unilateral LEC inactivation. Unilateral mPFC inactivation had no effect on recent or remote memory expression. These results suggest that the integrity of the LEC in both hemispheres is necessary for memory expression. Functional interactions between the LEC and mPFC should therefore be tested with an alternative design.