Catherine C. Kaczorowski

Memory deficits are associated with impaired ability to modulate neuronal excitability in middle-aged mice

Normal aging disrupts hippocampal neuroplasticity and learning and memory. Aging deficits were exposed in a subset (30%) of middle-aged mice that performed below criterion on a hippocampal-dependent contextual fear conditioning task. Basal neuronal excitability was comparable in middle-aged and young mice, but learning-related modulation of the post-burst afterhyperpolarization (AHP)--a general mechanism engaged during learning--was impaired in CA1 neurons from middle-aged weak learners. Thus, modulation of neuronal excitability is critical for retention of context fear in middle-aged mice. Disruption of AHP plasticity may contribute to contextual fear deficits in middle-aged mice--a model of age-associated cognitive decline (AACD).

Stability and plasticity of intrinsic membrane properties in hippocampal CA1 pyramidal neurons: effects of internal anions

CA1 pyramidal neurons from animals that have acquired hippocampal tasks show increased neuronal excitability, as evidenced by a reduction in the postburst afterhyperpolarization (AHP). Studies of AHP plasticity require stable long‐term recordings, which are affected by the intracellular solutions potassium methylsulphate (KMeth) or potassium gluconate (KGluc). Here we show immediate and gradual effects of these intracellular solutions on measurement of the AHP and basic membrane properties, and on the induction of AHP plasticity in CA1 pyramidal neurons from rat hippocampal slices. The AHP measured immediately after establishing whole‐cell recordings was larger with KMeth than with KGluc. In general, the AHP in KMeth was comparable to the AHP measured in the perforated‐patch configuration. However, KMeth induced time‐dependent changes in the intrinsic membrane properties of CA1 pyramidal neurons. Specifically, input resistance progressively increased by 70% after 50 min; correspondingly, the current required to trigger an action potential and the fast afterdepolarization following action potentials gradually decreased by about 50%. Conversely, these measures were stable in KGluc. We also demonstrate that activity‐dependent plasticity of the AHP occurs with physiologically relevant stimuli in KGluc. AHPs triggered with theta‐burst firing every 30 s were progressively reduced, whereas AHPs elicited every 150 s were stable. Blockade of the apamin‐sensitive AHP current (IAHP) was insufficient to block AHP plasticity, suggesting that plasticity is manifested through changes in the apamin‐insensitive slow AHP current (sIAHP). These changes were observed in the presence of synaptic blockers, and therefore reflect changes in the intrinsic properties of the neurons. However, no AHP plasticity was observed using KMeth. In summary, these data show that KMeth produces time‐dependent changes in basic membrane properties and prevents or obscures activity‐dependent reduction of the AHP. In whole‐cell recordings using KGluc, repetitive theta‐burst firing induced AHP plasticity that mimics learning‐related reduction in the AHP.

Mechanisms underlying basal and learning-related intrinsic excitability in a mouse model of Alzheimer's disease.

Accumulations of β-amyloid (Aβ) contribute to neurological deficits associated with Alzheimers disease (AD). The effects of Aβ on basal neuronal excitability and learning-related AHP plasticity were examined using whole-cell recordings from hippocampal neurons in the 5XFAD mouse model of AD. A robust increase in Aβ42 (and elevated levels of Aβ38-40) in naïve 5XFAD mice was associated with decreased basal neuronal excitability, evidenced by a select increase in Ca(2+)-sensitive afterhyperpolarization (AHP). Moreover, trace fear deficits observed in a subset of 5XFAD weak-learner mice were associated with a greater enhancement of the AHP in neurons, as compared to age-matched 5XFAD learner and 5XFAD naïve mice. Importantly, learning-related plasticity of the AHP remained intact in a subset of 5XFAD mice that learned trace fear conditioning to a set criterion. We show that APP-PS1 mutations enhance Aβ and disrupt basal excitability via a Ca(2+)-dependent enhancement of the AHP, and suggest disruption to learning-related modulation of intrinsic excitability resulted, in part, from altered cholinergic modulation of the AHP in the 5XFAD mouse model of AD (170 of 170).

Leptin Modulates the Intrinsic Excitability of AgRP/NPY Neurons in the Arcuate Nucleus of the Hypothalamus

The hypothalamic arcuate nucleus (ARH) is a brain region critical for regulation of food intake and a primary area for the action of leptin in the CNS. In lean mice, the adipokine leptin inhibits neuropeptide Y (NPY) and agouti-related peptide (AgRP) neuronal activity, resulting in decreased food intake. Here we show that diet-induced obesity in mice is associated with persistent activation of NPY neurons and a failure of leptin to reduce the firing rate or hyperpolarize the resting membrane potential. However, the molecular mechanism whereby diet uncouples leptins effect on neuronal excitability remains to be fully elucidated. In NPY neurons from lean mice, the Kv channel blocker 4-aminopyridine inhibited leptin-induced changes in input resistance and spike rate. Consistent with this, we found that ARH NPY neurons have a large, leptin-sensitive delayed rectifier K+ current and that leptin sensitivity of this current is blunted in neurons from diet-induced obese mice. This current is primarily carried by Kv2-containing channels, as the Kv2 channel inhibitor stromatoxin-1 significantly increased the spontaneous firing rate in NPY neurons from lean mice. In HEK cells, leptin induced a significant hyperpolarizing shift in the voltage dependence of Kv2.1 but had no effect on the function of the closely related channel Kv2.2 when these channels were coexpressed with the long isoform of the leptin receptor LepRb. Our results suggest that dynamic modulation of somatic Kv2.1 channels regulates the intrinsic excitability of NPY neurons to modulate the spontaneous activity and the integration of synaptic input onto these neurons in the ARH.

Aging redistributes medial prefrontal neuronal excitability and impedes extinction of trace fear conditioning

Cognitive flexibility is critical for survival and reflects the malleability of the central nervous system (CNS) in response to changing environmental demands. Normal aging results in difficulties modifying established behaviors, which may involve medial prefrontal cortex (mPFC) dysfunction. Using extinction of conditioned fear in rats to assay cognitive flexibility, we demonstrate that extinction deficits reminiscent of mPFC dysfunction first appear during middle age, in the absence of hippocampus-dependent context deficits. Emergence of aging-related extinction deficits paralleled a redistribution of neuronal excitability across two critical mPFC regions via two distinct mechanisms. First, excitability decreased in regular spiking neurons of infralimbic-mPFC (IL), a region whose activity is required for extinction. Second, excitability increased in burst spiking neurons of prelimbic-mPFC (PL), a region whose activity hinders extinction. Experiments using synaptic blockers revealed that these aging-related differences were intrinsic. Thus, changes in IL and PL intrinsic excitability may contribute to cognitive flexibility impairments observed during normal aging.

TRPC3 channels critically regulate hippocampal excitability and contextual fear memory

Memory formation requires de novo protein synthesis, and memory disorders may result from misregulated synthesis of critical proteins that remain largely unidentified. Plasma membrane ion channels and receptors are likely candidates given their role in regulating neuron excitability, a candidate memory mechanism. Here we conduct targeted molecular monitoring and quantitation of hippocampal plasma membrane proteins from mice with intact or impaired contextual fear memory to identify putative candidates. Here we report contextual fear memory deficits correspond to increased Trpc3 gene and protein expression, and demonstrate TRPC3 regulates hippocampal neuron excitability associated with memory function. These data provide a mechanistic explanation for enhanced contextual fear memory reported herein following knockdown of TRPC3 in hippocampus. Collectively, TRPC3 modulates memory and may be a feasible target to enhance memory and treat memory disorders.

Hippocampal proteomics defines pathways associated with memory decline and resilience in normal aging and Alzheimer's disease mouse models

GRAPHICAL ABSTRACT Figure. No caption available. HIGHLIGHTSProteomics detects 36 hippocampal proteins associated with AD and normal aging memory deficits.Pathway analysis highlights HDAC4 as global regulator of memory deficits.103 proteins differ specifically in AD mice with intact vs impaired memory.Pathway analysis indicates disease‐specific involvement of REST and Gi signaling.Publically available proteomics resource for hypothesis generation and testing. ABSTRACT Alzheimers disease (AD), the most common form of dementia in the elderly, has no cure. Thus, the identification of key molecular mediators of cognitive decline in AD remains a top priority. As aging is the most significant risk factor for AD, the goal of this study was to identify altered proteins and pathways associated with the development of normal aging and AD memory deficits, and identify unique proteins and pathways that may contribute to AD‐specific symptoms. We used contextual fear conditioning to diagnose 8‐month‐old 5XFAD and non‐transgenic (Ntg) mice as having either intact or impaired memory, followed by liquid chromatography‐tandem mass spectrometry (LC–MS/MS) to quantify hippocampal membrane proteins across groups. Subsequent analysis detected 113 proteins differentially expressed relative to memory status (intact vs impaired) in Ntg mice and 103 proteins in 5XFAD mice. Thirty‐six proteins, including several involved in neuronal excitability and synaptic plasticity (e.g., GRIA1, GRM3, and SYN1), were altered in both normal aging and AD. Pathway analysis highlighted HDAC4 as a regulator of observed protein changes in both genotypes and identified the REST epigenetic regulatory pathway and Gi intracellular signaling as AD‐specific pathways involved in regulating the onset of memory deficits. Comparing the hippocampal membrane proteome of Ntg versus AD, regardless of cognitive status, identified 138 differentially expressed proteins, including confirmatory proteins APOE and CLU. Overall, we provide a novel list of putative targets and pathways with therapeutic potential, including a set of proteins associated with cognitive status in normal aging mice or gene mutations that cause AD.

Brain derived neurotrophic factor differentially modulates excitability of two classes of hippocampal output neurons

We show that BDNF mediates acute and long-lasting changes in intrinsic neuronal excitability in two classes of subicular pyramidal neurons (EB and LB neurons). Although BDNF plays similar roles in the induction of synaptic plasticity in these two cell types, it differentially and bidirectionally affects intrinsic excitability and burst plasticity in EB and LB neurons. These cell type-specific effects represent new avenues by which BDNF can influence the well-established information storage function of the hippocampus.

Rhythmic intrinsic bursting neurons in human neocortex obtained from pediatric patients with epilepsy

Neocortical oscillations result from synchronized activity of a synaptically coupled network and can be strongly influenced by the intrinsic firing properties of individual neurons. As such, the intrinsic electroresponsive properties of individual neurons may have important implications for overall network function. Rhythmic intrinsic bursting (rIB) neurons are of particular interest, as they are poised to initiate and/or strongly influence network oscillations. Although neocortical rIB neurons have been recognized in multiple species, the current study is the first to identify and characterize rIB neurons in the human neocortex. Using whole‐cell current‐clamp recordings, rIB neurons (n = 12) are identified in human neocortical tissue resected from pediatric patients with intractable epilepsy. In contrast to human regular spiking neurons (n = 12), human rIB neurons exhibit rhythmic bursts of action potentials at frequencies of 0.1–4 Hz. These bursts persist after blockade of fast excitatory neurotransmission and voltage‐gated calcium channels. However, bursting is eliminated by subsequent application of the persistent sodium current (INaP) blocker, riluzole. In the presence of riluzole (either 10 or 20 μm), human rIB neurons no longer burst, but fire tonically like regular spiking neurons. These data demonstrate that INaP plays a critical role in intrinsic oscillatory activity observed in rIB neurons in the human neocortex. It is hypothesized that aberrant changes in INaP expression and/or function may ultimately contribute to neurological diseases that are linked to abnormal network activity, such as epilepsy.

Bidirectional pattern-specific plasticity of the slow afterhyperpolarization in rats: role for high-voltage activated Ca2+ channels and I h.

A burst of action potentials in hippocampal neurons is followed by a slow afterhyperpolarization (sAHP) that serves to limit subsequent firing. A reduction in the sAHP accompanies acquisition of several types of learning, whereas increases in the sAHP are correlated with cognitive impairment. The present study demonstrates in vitro that activity‐dependent bidirectional plasticity of the sAHP does not require synaptic activation, and depends on the pattern of action potential firing. Whole‐cell current‐clamp recordings from CA1 pyramidal neurons in hippocampal slices from young rats (postnatal days14–24) were performed in blockers of synaptic transmission. The sAHP was evoked by action potential firing at gamma‐related (50 Hz, gamma‐AHP) or theta frequencies (5 Hz, theta‐AHP), two firing frequencies implicated in attention and memory. Interestingly, when the gamma‐AHP and theta‐AHP were evoked in the same cell, a gradual potentiation of the gamma‐AHP (186 ± 31%) was observed that was blocked using Ca2+ channel blockers nimodipine (10 μm) or ω‐conotoxin MVIIC (1 μm). In experiments that exclusively evoked the sAHP with 50 Hz firing, the gamma‐AHP was similarly potentiated (198 ± 44%). However, theta‐burst firing pattern alone resulted in a decrease (65 ± 19%) of the sAHP. In these experiments, application of the h‐channel blocker ZD7288 (25 μm) selectively prevented enhancement of the gamma‐AHP. These data demonstrate that induction requirements for bidirectional AHP plasticity depend on the pattern of action potential firing, and result from distinct mechanisms. The identification of novel mechanisms underlying AHP plasticity in vitro provides additional insight into the dynamic processes that may regulate neuronal excitability during learning in vivo.