Sprague-Dawley Rats Differ in Responses to Medial Perforant Path Paired Pulse and Tetanic Activation as a Function of Sex and Age.

Network plasticity in the medial perforant path (MPP) of adult (five to nine months) and aged (18-20 months) urethane-anesthetized male and female Sprague Dawley rats was characterized. Paired pulses probed recurrent networks before and after a moderate tetanic protocol. Adult females exhibited greater EPSP-spike coupling suggesting greater intrinsic excitability than adult males. Aged rats did not differ in EPSP-spike coupling but aged females had larger spikes at high currents than males. Paired pulses suggested lower GABA-B inhibition in females. Absolute population spike (PS) measures were larger post-tetani in female rats than male rats. Relative population spike increases were greatest in adult males relative to females and to aged males. EPSP slope potentiation was detected with normalization in some post-tetanic intervals for all groups except aged males. Tetani shortened spike latency across groups. Tetani-associated NMDA-mediated burst depolarizations were larger for the first two trains in each tetanus in adult males than other groups. EPSP slopes over 30 min post-tetani predicted spike size in female rats but not in males. Replicating newer evidence MPP plasticity in adult males was mediated by increased intrinsic excitability. Female MPP plasticity was related to synaptic drive increases, not excitability increases. Aged male rats were deficient in MPP plasticity.

Locus Coeruleus Optogenetic Modulation: Lessons Learned from Temporal Patterns.

After reviewing seminal studies using optogenetics to interrogate the functional role of the locus coeruleus in behavior, we conclude that differences in firing rates and firing patterns of locus coeruleus neurons contribute to locus coeruleus nucleus heterogeneity by recruiting different output circuitry, and differentially modifying behavior. The outcomes initiated by different optogenetic input activation patterns and frequencies can have opposite consequences for behavior, activate different neurons in the same target structure, be supported by distinct adrenoceptors and vary with behavioral state.

Locus Coeruleus Activation Patterns Differentially Modulate Odor Discrimination Learning and Odor Valence in Rats.

The locus coeruleus (LC) produces phasic and tonic firing patterns that are theorized to have distinct functional consequences. However, how different firing modes affect learning and valence encoding of sensory information are unknown. Here, we show bilateral optogenetic activation of rat LC neurons using 10-Hz phasic trains of either 300 ms or 10 s accelerated acquisition of a similar odor discrimination. Similar odor discrimination learning was impaired by noradrenergic blockade in the piriform cortex (PC). However, 10-Hz phasic light-mediated learning facilitation was prevented by a dopaminergic antagonist in the PC, or by ventral tegmental area (VTA) silencing with lidocaine, suggesting a LC-VTA-PC dopamine circuitry involvement. Ten-hertz tonic stimulation did not alter odor discrimination acquisition, and was ineffective in activating VTA DA neurons. For valence encoding, tonic stimulation at 25 Hz induced conditioned odor aversion, whereas 10-Hz phasic stimulations produced an odor preference. Both conditionings were prevented by noradrenergic blockade in the basolateral amygdala (BLA). Cholera Toxin B retro-labeling showed larger engagement of nucleus accumbens-projecting neurons in the BLA with 10-Hz phasic activation, and larger engagement of central amygdala projecting cells with 25-Hz tonic light. These outcomes argue that the LC activation patterns differentially influence both target networks and behavior.

The 'a, b, c's of pretangle tau and their relation to aging and the risk of Alzheimer's Disease.

Braak has described the beginnings of Alzheimer's Disease as occurring in the locus coeruleus. Here we review these pretangle stages and relate their expression to recently described normal features of tau biology. We suggest pretangle tau depends on characteristics of locus coeruleus operation that promote tau condensates. We examine the timeline of pretangle and tangle appearance in locus coeruleus. We find catastrophic loss of locus coeruleus neurons is a late event. The strong relationship between locus coeruleus neuron number and human cognition underscores the utility of a focus on locus coeruleus. Promoting locus coeruleus health will benefit normal aging as well as aid in the prevention of dementia. Two animal models offering experimental approaches to understanding the functional change initiated by pretangles in locus coeruleus neurons are discussed.

Novelty-like activation of locus coeruleus protects against deleterious human pretangle tau effects while stress-inducing activation worsens its effects.

The earliest abnormality associated with Alzheimer's disease (AD) is the presence of persistently phosphorylated pretangle tau in locus coeruleus (LC) neurons. LC neuron numbers and fiber density are positive predictors of cognition prior to death. Using an animal model of LC pretangle tau, we ask if LC activity patterns influence the sequelae of pretangle tau. We seeded LC neurons with a pretangle human tau gene. We provided daily novelty- or stress-associated optogenetic activation patterns to LC neurons for 6 weeks in mid-adulthood and, subsequently, probed cognitive and anatomical changes. Prior LC phasic stimulation prevented spatial and olfactory discrimination deficits and preserved LC axonal density. A stress-associated activation pattern increased indices of anxiety and depression, did not improve cognition, and worsened LC neuronal health. These results argue that variations in environmental experiences associated with differing LC activity patterns may account for individual susceptibility to development of AD in humans.

Locus coeruleus: a new look at the blue spot.

The locus coeruleus (LC), or 'blue spot', is a small nucleus located deep in the brainstem that provides the far-reaching noradrenergic neurotransmitter system of the brain. This phylogenetically conserved nucleus has proved relatively intractable to full characterization, despite more than 60 years of concerted efforts by investigators. Recently, an array of powerful new neuroscience tools have provided unprecedented access to this elusive nucleus, revealing new levels of organization and function. We are currently at the threshold of major discoveries regarding how this tiny brainstem structure exerts such varied and significant influences over brain function and behaviour. All LC neurons receive inputs related to autonomic arousal, but distinct subpopulations of those neurons can encode specific cognitive processes, presumably through more specific inputs from the forebrain areas. This ability, combined with specific patterns of innervation of target areas and heterogeneity in receptor distributions, suggests that activation of the LC has more specific influences on target networks than had initially been imagined.

Learning-induced mRNA alterations in olfactory bulb mitral cells in neonatal rats.

In the olfactory bulb, a cAMP/PKA/CREB-dependent form of learning occurs in the first week of life that provides a unique mammalian model for defining the epigenetic role of this evolutionarily ancient plasticity cascade. Odor preference learning in the week-old rat pup is rapidly induced by a 10-min pairing of odor and stroking. Memory is demonstrable at 24 h, but not 48 h, posttraining. Using this paradigm, pups that showed peppermint preference 30 min posttraining were sacrificed 20 min later for laser microdissection of odor-encoding mitral cells. Controls were given odor only. Microarray analysis revealed that 13 nonprotein-coding mRNAs linked to mRNA translation and splicing and 11 protein-coding mRNAs linked to transcription differed with odor preference training. MicroRNA23b, a translation inhibitor of multiple plasticity-related mRNAs, was down-regulated. Protein-coding transcription was up-regulated for Sec23b, Clic2, Rpp14, Dcbld1, Magee2, Mstn, Fam229b, RGD1566265, and Mgst2. Gng12 and Srcg1 mRNAs were down-regulated. Increases in Sec23b, Clic2, and Dcbld1 proteins were confirmed in mitral cells in situ at the same time point following training. The protein-coding changes are consistent with extracellular matrix remodeling and ryanodine receptor involvement in odor preference learning. A role for CREB and AP1 as triggers of memory-related mRNA regulation is supported. The small number of gene changes identified in the mitral cell input/output link for 24 h memory will facilitate investigation of the nature, and reversibility, of changes supporting temporally restricted long-term memory.

An experimental model of Braak's pretangle proposal for the origin of Alzheimer's disease: the role of locus coeruleus in early symptom development.

BACKGROUND: The earliest brain pathology related to Alzheimer's disease (AD) is hyperphosphorylated soluble tau in the noradrenergic locus coeruleus (LC) neurons. Braak characterizes five pretangle tau stages preceding AD tangles. Pretangles begin in young humans and persist in the LC while spreading from there to other neuromodulatory neurons and, later, to the cortex. While LC pretangles appear in all by age 40, they do not necessarily result in AD prior to death. However, with age and pretangle spread, more individuals progress to AD stages. LC neurons are lost late, at Braak stages III-IV, when memory deficits appear. It is not clear if LC hyperphosphorylated tau generates the pathology and cognitive changes associated with preclinical AD. We use a rat model expressing pseudohyperphosphorylated human tau in LC to investigate the hypothesis that LC pretangles generate preclinical Alzheimer pathology. METHODS: We infused an adeno-associated viral vector carrying a human tau gene pseudophosphorylated at 14 sites common in LC pretangles into 2-3- or 14-16-month TH-Cre rats. We used odor discrimination to probe LC dysfunction, and we evaluated LC cell and fiber loss. RESULTS: Abnormal human tau was expressed in LC and exhibited somatodendritic mislocalization. In rats infused at 2-3 months old, 4 months post-infusion abnormal LC tau had transferred to the serotonergic raphe neurons. After 7 months, difficult similar odor discrimination learning was impaired. Impairment was associated with reduced LC axonal density in the olfactory cortex and upregulated ß1-adrenoceptors. LC infusions in 14-16-month-old rats resulted in more severe outcomes. By 5-6 months post-infusion, rats were impaired even in simple odor discrimination learning. LC neuron number was reduced. Human tau appeared in the microglia and cortical neurons. CONCLUSIONS: Our animal model suggests, for the first time, that Braak's hypothesis that human AD originates with pretangle stages is plausible. LC pretangle progression here generates both preclinical AD pathological changes and cognitive decline. The odor discrimination deficits are similar to human odor identification deficits seen with aging and preclinical AD. When initiated in aged rats, pretangle stages progress rapidly and cause LC cell loss. These age-related outcomes are associated with a severe learning impairment consistent with memory decline in Braak stages III-IV.

Emotional arousal amplifies competitions across goal-relevant representation: A neurocomputational framework.

Emotional arousal often facilitates memory for some aspects of an event while impairing memory for other aspects of the same event. Across three experiments, we found that emotional arousal amplifies competition among goal-relevant representations, such that arousal impairs memory for multiple goal-relevant representations while enhancing memory for solo goal-relevant information. We also present a computational model to explain the mechanisms by which emotional arousal can modulate memory in opposite ways via the local/synaptic-level noradrenergic system. The model is based on neurophysiological observations that norepinephrine (NE) released under emotional arousal is locally controlled by glutamate levels, resulting in different NE effects across regions, gating either long-term potentiation or long-term depression by activating different adrenergic receptors depending on NE concentration levels. This model successfully replicated behavioral findings from the three experiments. These findings suggest that the NE's local effects are key in determining the effects of emotion on memory.

Locus Coeruleus Phasic, But Not Tonic, Activation Initiates Global Remapping in a Familiar Environment.

Locus coeruleus (LC) neurons, the source of hippocampal norepinephrine (NE), are activated by novelty and changes in environmental contingencies. Based on the role of monoamines in reconfiguring invertebrate networks, and data from mammalian systems, a network reset hypothesis for the effects of LC activation has been proposed. We used the cellular compartmental analysis of temporal FISH technique based on the cellular distribution of immediate early genes to examine the effect of LC activation and inactivation, on regional hippocampal maps in male rats, when LC activity was manipulated just before placement in a second familiar (A/A) and/or novel environment (A/B). We found that bilateral phasic, but not tonic, activation of LC reset hippocampal maps in the A/A condition, whereas silencing the LC with clonidine before placement in the A/B condition blocked map reset and a familiar map emerged in the dentate gyrus, proximal and distal CA1, and CA3c. However, CA3a and CA3b encoded the novel environment. These results support a role for phasic LC responses in generating novel hippocampal sequences during memory encoding and, potentially, memory updating. The silencing experiments suggest that novel environments may not be recognized as different by dentate gyrus and CA1 without LC input. The functional distinction between phasic and tonic LC activity argues that these parameters are critical for determining network changes. These data are consistent with the hippocampus activating internal network representations to encode novel experiential episodes and suggest LC input is critical for this role.SIGNIFICANCE STATEMENT Burst activation of the broadly projecting novelty signaling system of the locus coeruleus initiates new network representations throughout the hippocampus despite unchanged external environments. Tonic activation does not alter network representations in the same condition. This suggests differences in the temporal parameters of neuromodulator network activation are critical for neuromodulator function. Silencing this novelty signaling system prevented the appearance of new network representations in a novel environment. Instead, familiar representations were expressed in a subset of hippocampal areas, with another subset encoding the novel environment. This "being in two places at once" argues for independent functional regions within the hippocampus. These experiments strengthen the view that internal states are major determinants of the brain's construction of environmental representations.

Pheromone-Induced Odor Associative Fear Learning in Rats.

Alarm pheromones alert conspecifics to the presence of danger. Can pheromone communication aid in learning specific cues? Such facilitation has an evident evolutionary advantage. We use two associative learning paradigms to test this hypothesis. The first is stressed cage mate-induced conditioning. One pair-housed adult rat received 4 pairings of terpinene + shock over 30 min. Ten minutes after return to the home cage, its companion rat was removed and exposed to terpinene. Single-housed controls were exposed to either terpinene or shock only. Companion rats showed terpinene-specific freezing, which was prevented by ß-adrenoceptor blockade. Using Arc to index neuronal activation in response to terpinene re-exposure, stressed cage-mate induced associative learning was measured. Companion rats showed increased neuronal activity in the accessory olfactory bulb, while terpinene + shock-conditioned rats showed increased activity in the main olfactory bulb. Both groups had enhanced activity in the anterior basolateral amygdala and central amygdala. To test involvement of pheromone mediation, in the 2nd paradigm, we paired terpinene with soiled bedding from odor + shock rats or a rat alarm pheromone. Both conditioning increased rats' freezing to terpinene. Blocking NMDA receptors in the basolateral amygdala prevented odor-specific learning suggesting shock and pheromone-paired pathways converge in the amygdala. An alarm pheromone thus enables cue-specific learning as well as signalling danger.

Revisiting metaplasticity: The roles of calcineurin and histone deacetylation in unlearning odor preference memory in rat pups.

Previous work has shown that 24 h duration odor preference learning, induced by one-trial training, generates a down-regulation of the GluN1 receptor in anterior piriform cortex at 3 h, and results in metaplastic unlearning if a second training trial is given at 3 h. The GluN1 receptor upregulates at 24 h so 24 h spaced training is highly effective in extending memory duration. The present study replicates the piriform cortex unlearning result in the olfactory bulb circuit and further studies the relationship between the initial training strength and its associated metaplastic effect. Intrabulbar infusions that block calcineurin or inhibit histone deacetylation normally produce extended days-long memory. If given during training, they are not associated with GluN1 downregulation at 3 h and do not recruit an unlearning process at that time. The two memory strengthening protocols do not appear to interact, but are also not synergistic. These outcomes argue that it is critical to understand the metaplastic effects of training in order to optimize training protocols in the service of either memory strengthening or of memory weakening.

Locus Coeruleus Optogenetic Light Activation Induces Long-Term Potentiation of Perforant Path Population Spike Amplitude in Rat Dentate Gyrus.

Norepinephrine (NE) in dentate gyrus (DG) produces NE-dependent long-term potentiation (NE-LTP) of the perforant path-evoked potential population spike both in vitro and in vivo. Chemical activators infused near locus coeruleus (LC), the source of DG NE, produce a NE-LTP that is associative, i.e., requires concurrent pairing with perforant path (PP) input. Here, we ask if LC optogenetic stimulation that allows us to activate only LC neurons can induce NE-LTP in DG. We use an adeno-associated viral vector containing a depolarizing channel (AAV8-Ef1a-DIO-eChR2(h134r)-EYFP-WPRE) infused stereotaxically into the LC of TH:Cre rats to produce light-sensitive LC neurons. A co-localization of ~62% in LC neurons was observed for these channels. Under urethane anesthesia, we demonstrated that 5-10 s 10 Hz trains of 30 ms light pulses in LC reliably activated neurons near an LC optoprobe. Ten minutes of the same train paired with 0.1 Hz PP electrical stimulation produced a delayed NE-LTP of population spike amplitude, but not EPSP slope. A leftward shift in the population spike input/output curve at the end of the experiment was also consistent with long-term population spike potentiation. LC neuron activity during the 10 min light train was unexpectedly transient. Increased LC neuronal firing was seen only for the first 2 min of the light train. NE-LTP was more delayed and less robust than reported with LC chemo-activation. Previous estimates of LC axonal conduction times suggest acute release of NE occurs 40-70 ms after an LC neuron action potential. We used single LC light pulses to examine acute effects of NE release and found potentiated population spike amplitude when a light pulse in LC occurred 40-50 ms, but not 20-30 ms, prior to a PP pulse, consistent with conduction estimates. These effects of LC optogenetic activation reinforce evidence for a continuum of NE potentiation effects in DG. The single pulse effects mirror an earlier report using LC electrical stimulation. These acute effects support an attentional role of LC activation. The LTP of PP responses induced by optogenetic LC activation is consistent with the role of LC in long-term learning and memory.

Histone deacetylase inhibition induces odor preference memory extension and maintains enhanced AMPA receptor expression in the rat pup model.

Histone deacetylase (HDAC) plays a role in synaptic plasticity and long-term memory formation. We hypothesized that trichostatin-A (TSA), an HDAC inhibitor, would promote long-term odor preference memory and maintain enhanced GluA1 receptor levels that have been hypothesized to support memory. We used an early odor preference learning model in neonate rat pups that normally produces only 24-h memory to test behavior and examine receptor protein expression. Our behavioral studies showed that intrabulbar infusion of TSA, prior to pairing of the conditioned stimulus (peppermint odor) with the unconditioned stimulus (tactile stimulation), prolonged 24-h odor preference memory for at least 9 d. The prolonged odor preference memory was selective for the paired odor and was also observed using a specific HDAC6 inhibitor, tubacin, supporting a role for histone acetylation in associative memory. Immunoblot analysis showed that GluA1 receptor membrane expression in the olfactory bulbs of the TSA-treated group was significantly increased at 48 h unlike control rats without TSA. Immunohistochemistry revealed significant increase of GluA1 expression in olfactory bulb glomeruli 5 d after training. These results extend previous evidence for a close relationship between enhanced GluA1 receptor membrane expression and memory expression. Together, these findings provide a new single-trial appetitive model for understanding the support and maintenance of memories of varying duration.

Learning-Induced Metaplasticity? Associative Training for Early Odor Preference Learning Down-Regulates Synapse-Specific NMDA Receptors via mGluR and Calcineurin Activation.

Rat pups readily form a 24-h associative odor preference after a single trial of odor paired with intermittent stroking. Recent evidence shows that this training trial, which normally increases AMPA receptor responses in the anterior piriform cortex both 3 and 24 h following training, induces a down-regulation of NMDA receptors 3 h later followed by NMDA receptor up-regulation at 24 h. When retrained with the same odor at 3 h, rat pups unlearn the original odor preference. Unlearning can be prevented by blocking NMDA receptors during retraining. Here, the mechanisms that initiate NMDA receptor down-regulation are assessed. Blocking mGluR receptors or calcineurin during training prevents down-regulation of NMDA receptors 3 h following training. Blocking NMDA receptors during training does not affect NMDA receptor down-regulation. Thus, down-regulation can be engaged separately from associative learning. When unlearning occurs, AMPA and NMDA receptor levels at 24 h are reset to control levels. Calcineurin blockade during retraining prevents unlearning consistent with the role of NMDA receptor down-regulation. The relationship of these events to the metaplasticity and plasticity mechanisms of long-term depression and depotentiation is discussed. We suggest a possible functional role of NMDA receptor down-regulation in offline stabilization of learned odor representations.

CaMKII mediates stimulus specificity in early odor preference learning in rats.

After naturalistic odor preference training, Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) was rapidly phosphorylated in the olfactory bulb, specifically in the odor encoding regions of the glomerular layer and external plexiform layer. Intrabulbar CaMKII antagonist experiments revealed that CaMKII supports short- and long-term preference memory formation. With bulbar PKA activation as the unconditioned stimulus odor preferences could be induced despite CaMKII blockade, but now odor specificity was lost, with odor preference generalizing to an untrained odor. Odor-specific learning was associated with increased membrane-associated AMPA receptors, while nonspecific odor preference was not. Thus CaMKII activation provides a tag to confer stimulus specificity as well as supporting natural odor preference learning.

The Locus Coeruleus: Essential for Maintaining Cognitive Function and the Aging Brain.

Research on cognitive aging has focused on how decline in various cortical and hippocampal regions influence cognition. However, brainstem regions play essential modulatory roles, and new evidence suggests that, among these, the integrity of the locus coeruleus (LC)-norepinephrine (NE) system plays a key role in determining late-life cognitive abilities. The LC is especially vulnerable to toxins and infection and is often the first place Alzheimer's-related pathology appears, with most people showing at least some tau pathology by their mid-20s. On the other hand, NE released from the LC during arousing, mentally challenging, or novel situations helps to protect neurons from damage, which may help to explain how education and engaging careers prevent cognitive decline in later years.

Norepinephrine ignites local hotspots of neuronal excitation: How arousal amplifies selectivity in perception and memory.

Emotional arousal enhances perception and memory of high-priority information but impairs processing of other information. Here, we propose that, under arousal, local glutamate levels signal the current strength of a representation and interact with norepinephrine (NE) to enhance high priority representations and out-compete or suppress lower priority representations. In our "glutamate amplifies noradrenergic effects" (GANE) model, high glutamate at the site of prioritized representations increases local NE release from the locus coeruleus (LC) to generate "NE hotspots." At these NE hotspots, local glutamate and NE release are mutually enhancing and amplify activation of prioritized representations. In contrast, arousal-induced LC activity inhibits less active representations via two mechanisms: 1) Where there are hotspots, lateral inhibition is amplified; 2) Where no hotspots emerge, NE levels are only high enough to activate low-threshold inhibitory adrenoreceptors. Thus, LC activation promotes a few hotspots of excitation in the context of widespread suppression, enhancing high priority representations while suppressing the rest. Hotspots also help synchronize oscillations across neural ensembles transmitting high-priority information. Furthermore, brain structures that detect stimulus priority interact with phasic NE release to preferentially route such information through large-scale functional brain networks. A surge of NE before, during, or after encoding enhances synaptic plasticity at NE hotspots, triggering local protein synthesis processes that enhance selective memory consolidation. Together, these noradrenergic mechanisms promote selective attention and memory under arousal. GANE not only reconciles apparently contradictory findings in the emotion-cognition literature but also extends previous influential theories of LC neuromodulation by proposing specific mechanisms for how LC-NE activity increases neural gain.

GANEing traction: The broad applicability of NE hotspots to diverse cognitive and arousal phenomena.

The GANE (glutamate amplifies noradrenergic effects) model proposes that local glutamate-norepinephrine interactions enable "winner-take-more" effects in perception and memory under arousal. A diverse range of commentaries addressed both the nature of this "hotspot" feedback mechanism and its implications in a variety of psychological domains, inspiring exciting avenues for future research.

Arc-Expressing Neuronal Ensembles Supporting Pattern Separation Require Adrenergic Activity in Anterior Piriform Cortex: An Exploration of Neural Constraints on Learning.

Arc ensembles in adult rat olfactory bulb (OB) and anterior piriform cortex (PC) were assessed after discrimination training on highly similar odor pairs. Nonselective α- and ß-adrenergic antagonists or saline were infused in the OB or anterior PC during training. OB adrenergic blockade slowed, but did not prevent, odor discrimination learning. After criterion performance, Arc ensembles in anterior piriform showed enhanced stability for the rewarded odor and pattern separation for the discriminated odors as described previously. Anterior piriform adrenergic blockade prevented acquisition of similar odor discrimination and of OB ensemble changes, even with extended overtraining. Mitral and granule cell Arc ensembles in OB showed enhanced stability for rewarded odor only in the saline group. Pattern separation was not seen in the OB. Similar odor discrimination co-occurs with increased stability in rewarded odor representations and pattern separation to reduce encoding overlap. The difficulty of similar discriminations may relate to the necessity to both strengthen rewarded representations and weaken overlap across similar representations. SIGNIFICANCE STATEMENT: We show for the first time that adrenoceptors in anterior piriform cortex (aPC) must be engaged for adult rats to learn to discriminate highly similar odors. Loss of adrenergic activation in olfactory bulb (OB) slows, but does not prevent, discrimination learning. Both increased stability of the rewarded odor representation and increased pattern separation of the rewarded and unrewarded odors in aPC accompany successful discrimination. In the OB, rewarded odors increase in ensemble stability, but there is no evidence of pattern separation. We suggest that the slow acquisition of similar odor discriminations is related to the differing plasticity requirements for increased stability and pattern separation.