Progressive Excitability Changes in the Medial Entorhinal Cortex in the 3xTg Mouse Model of Alzheimer's Disease Pathology.

Alzheimer's disease (AD) is a chronic neurodegenerative disorder characterized by memory loss and progressive cognitive impairments. In mouse models of AD pathology, studies have found neuronal and synaptic deficits in hippocampus, but less is known about changes in medial entorhinal cortex (MEC), which is the primary spatial input to the hippocampus and an early site of AD pathology. Here, we measured neuronal intrinsic excitability and synaptic activity in MEC layer II (MECII) stellate cells, MECII pyramidal cells, and MEC layer III (MECIII) excitatory neurons at 3 and 10 months of age in the 3xTg mouse model of AD pathology, using male and female mice. At 3 months of age, before the onset of memory impairments, we found early hyperexcitability in intrinsic properties of MECII stellate and pyramidal cells, but this was balanced by a relative reduction in synaptic excitation (E) compared with inhibition (I; E/I ratio), suggesting intact homeostatic mechanisms regulating MECII activity. Conversely, MECIII neurons had reduced intrinsic excitability at this early time point with no change in synaptic E/I ratio. By 10 months of age, after the onset of memory deficits, neuronal excitability of MECII pyramidal cells and MECIII excitatory neurons was largely normalized in 3xTg mice. However, MECII stellate cells remained hyperexcitable, and this was further exacerbated by an increased synaptic E/I ratio. This observed combination of increased intrinsic and synaptic hyperexcitability suggests a breakdown in homeostatic mechanisms specifically in MECII stellate cells at this postsymptomatic time point, which may contribute to the emergence of memory deficits in AD.SIGNIFICANCE STATEMENT AD causes cognitive deficits, but the specific neural circuits that are damaged to drive changes in memory remain unknown. Using a mouse model of AD pathology that expresses both amyloid and tau transgenes, we found that neurons in the MEC have altered excitability. Before the onset of memory impairments, neurons in layer 2 of MEC had increased intrinsic excitability, but this was balanced by reduced inputs onto the cell. However, after the onset of memory impairments, stellate cells in MEC became further hyperexcitable, with increased excitability exacerbated by increased synaptic inputs. Thus, it appears that MEC stellate cells are uniquely disrupted during the progression of memory deficits and may contribute to cognitive deficits in AD.

Distinct changes to hippocampal and medial entorhinal circuits emerge across the progression of cognitive deficits in epilepsy.

Temporal lobe epilepsy (TLE) causes pervasive and progressive memory impairments, yet the specific circuit changes that drive these deficits remain unclear. To investigate how hippocampal-entorhinal dysfunction contributes to progressive memory deficits in epilepsy, we performed simultaneous in vivo electrophysiology in hippocampus (HPC) and medial entorhinal cortex (MEC) of control and epileptic mice 3 or 8 weeks after pilocarpine-induced status epilepticus (Pilo-SE). We found that HPC synchronization deficits (including reduced theta power, coherence, and altered interneuron spike timing) emerged within 3 weeks of Pilo-SE, aligning with early-onset, relatively subtle memory deficits. In contrast, abnormal synchronization within MEC and between HPC-MEC emerged later, by 8 weeks after Pilo-SE, when spatial memory impairment was more severe. Furthermore, a distinct subpopulation of MEC layer 3 excitatory neurons (active at theta troughs) was specifically impaired in epileptic mice. Together, these findings suggest that hippocampal-entorhinal circuit dysfunction accumulates and shifts as cognitive impairment progresses in TLE.

Progressive excitability changes in the medial entorhinal cortex in the 3xTg mouse model of Alzheimer's disease pathology.

Alzheimer's disease (AD) is a chronic neurodegenerative disorder that is characterized by memory loss and progressive cognitive impairments. In mouse models of AD pathology, studies have found neuronal and synaptic deficits in the hippocampus, but less is known about what happens in the medial entorhinal cortex (MEC), which is the primary spatial input to the hippocampus and an early site of AD pathology. Here, we measured the neuronal intrinsic excitability and synaptic activity in MEC layer II (MECII) stellate cells, MECII pyramidal cells, and MEC layer III (MECIII) excitatory neurons at early (3 months) and late (10 months) time points in the 3xTg mouse model of AD pathology. At 3 months of age, prior to the onset of memory impairments, we found early hyperexcitability in MECII stellate and pyramidal cells' intrinsic properties, but this was balanced by a relative reduction in synaptic excitation (E) compared to inhibition (I), suggesting intact homeostatic mechanisms regulating activity in MECII. Conversely, MECIII neurons had reduced intrinsic excitability at this early time point with no change in the synaptic E/I ratio. By 10 months of age, after the onset of memory deficits, neuronal excitability of MECII pyramidal cells and MECIII excitatory neurons was largely normalized in 3xTg mice. However, MECII stellate cells remained hyperexcitable and this was further exacerbated by an increased synaptic E/I ratio. This observed combination of increased intrinsically and synaptically generated excitability suggests a breakdown in homeostatic mechanisms specifically in MECII stellate cells at this post-symptomatic time point. Together, these data suggest that the breakdown in homeostatic excitability mechanisms in MECII stellate cells may contribute to the emergence of memory deficits in AD.

Chronotate: An open-source tool for manual timestamping and quantification of animal behavior.

A core necessity to behavioral neuroscience research is the ability to accurately measure performance on behavioral assays, such as the novel object location and novel object recognition tasks. These tasks are widely used in neuroscience research and measure a rodent's instinct for investigating novel features as a proxy to test their memory of a previous experience. Automated tools for scoring behavioral videos can be cost prohibitive and often have difficulty distinguishing between active investigation of an object and simply being in close proximity to an object. As such, many experimenters continue to rely on hand scoring interactions using stopwatches, which makes it difficult to review scoring after-the-fact and results in the loss of temporal information. Here, we introduce Chronotate, a free, open-source tool to aid in manually scoring novel object behavior videos. The software consists of an interactive video player with keyboard integration for marking timestamps of behavioral events during video playback, making it simple to quickly score and review bouts of rodent-object interaction. In addition, Chronotate outputs detailed interaction bout data, allowing for nuanced behavioral performance analyses. Using this detailed temporal information, we demonstrate that novel object location performance peaks within the first 3 s of interaction time and preference for the novel location becomes reduced across the test session. Thus, Chronotate can be used to determine the temporal structure of interactions on this task and can provide new insight into the memory processes that drive this behavior. Chronotate is available for download at: https://github.com/ShumanLab/Chronotate.

ezTrack-A Step-by-Step Guide to Behavior Tracking.

Tracking animal behavior by video is one of the most common tasks in neuroscience. Previously, we have validated ezTrack, a free, flexible, and easy-to-use software for the analysis of animal behavior. ezTrack's Location Tracking Module can be used for the positional analysis of an individual animal and is applicable to a wide range of behavioral tasks. Separately, ezTrack's Freeze Analysis Module is designed for the analysis of defensive freezing behavior. ezTrack supports a range of desirable tools, including options for cropping and masking portions of the field of view, defining regions of interest, producing summary data for specified portions of time, algorithms to remove the influence of electrophysiology cables and other tethers, batch processing of multiple videos, and video down-sampling. Moreover, ezTrack produces a range of interactive plots and visualizations to promote users' confidence in their results. In this protocols paper, we provide step-by-step instructions for the use of ezTrack, from tips for recording behavior to instructions for using the software for video analysis. © 2021 Wiley Periodicals LLC. Basic Protocol 1: Software environment installation Basic Protocol 2: Using the Location Tracking Module Basic Protocol 3: Using the Freeze Analysis Module.