Cognitive abnormalities and hippocampal alterations in monoamine oxidase A and B knockout mice.

The monoamine oxidase isoenzymes (MAOs) A and B play important roles in the homeostasis of monoaminergic neurotransmitters. The combined deficiency of MAO A and B results in significantly elevated levels of serotonin (5-hydroxytryptamine), norepinephrine, dopamine, and ß-phenylethylamine; in humans and mice, these neurochemical changes are accompanied by neurodevelopmental perturbations as well as autistic-like responses. Ample evidence indicates that normal levels of monoamines in the hippocampus, amygdala, frontal cortex, and cerebellum are required for the integrity of learning and memory. Thus, in the present study, the cognitive status of MAO A/B knockout (KO) mice was examined with a wide array of behavioral tests. In comparison with male wild-type littermates, MAO A/B KO mice exhibited abnormally high and overgeneralized fear conditioning and enhanced eye-blink conditioning. These alterations were accompanied by significant increases in hippocampal long-term potentiation and alterations in the relative expression of NMDA glutamate receptor subunits. Our data suggest that chronic elevations of monoamines, because of the absence of MAO A and MAO B, cause functional alterations that are accompanied with changes in the cellular mechanisms underlying learning and memory. The characteristics exhibited by MAO A/B KO mice highlight the potential of these animals as a useful tool to provide further insight into the molecular bases of disorders associated with abnormal monoaminergic profiles.

Prolonging the postcomplex spike pause speeds eyeblink conditioning.

Climbing fiber input to the cerebellum is believed to serve as a teaching signal during associative, cerebellum-dependent forms of motor learning. However, it is not understood how this neural pathway coordinates changes in cerebellar circuitry during learning. Here, we use pharmacological manipulations to prolong the postcomplex spike pause, a component of the climbing fiber signal in Purkinje neurons, and show that these manipulations enhance the rate of learning in classical eyelid conditioning. Our findings elucidate an unappreciated aspect of the climbing fiber teaching signal, and are consistent with a model in which convergent postcomplex spike pauses drive learning-related plasticity in the deep cerebellar nucleus. They also suggest a physiological mechanism that could modulate motor learning rates.

Allopregnanolone reverses neurogenic and cognitive deficits in mouse model of Alzheimer's disease.

Our previous analyses showed that allopregnanolone (APalpha) significantly increased proliferation of rodent and human neural progenitor cells in vitro. In this study, we investigated the efficacy of APalpha to promote neurogenesis in the hippocampal subgranular zone (SGZ), to reverse learning and memory deficits in 3-month-old male triple transgenic mouse model of Alzheimer's (3xTgAD) and the correlation between APalpha-induced neural progenitor cell survival and memory function in 3xTgAD mice. Neural progenitor cell proliferation was determined by unbiased stereological analysis of BrdU incorporation and survival determined by FACS for BrdU+ cells. Learning and memory function was assessed using the hippocampal-dependent trace eye-blink conditioning paradigm. At 3 months, basal level of BrdU+ cells in the SGZ of 3xTgAD mice was significantly lower relative to non-Tg mice, despite the lack of evident AD pathology. APalpha significantly increased, in a dose-dependent manner, BrdU+ cells in SGZ in 3xTgAD mice and restored SGZ proliferation to normal magnitude. As with the deficit in proliferation, 3xTgAD mice exhibited deficits in learning and memory. APalpha reversed the cognitive deficits to restore learning and memory performance to the level of normal non-Tg mice. In 3xTgAD mice, APalpha-induced survival of neural progenitors was significantly correlated with APalpha-induced memory performance. These findings suggest that early neurogenic deficits, which were evident before immunodetectable Abeta, may contribute to the cognitive phenotype of AD, and that APalpha could serve as a regenerative therapeutic to prevent or delay neurogenic and cognitive deficits associated with mild cognitive impairment and Alzheimer's disease.

Differential effects and rates of normal aging in cerebellum and hippocampus.

Cognitive functions show many alternative outcomes and great individual variation during normal aging. We examined learning over the adult life span in CBA mice, along with morphological and electrophysiological substrates. Our aim was to compare cerebellum-dependent delay eyeblink classical conditioning and hippocampus-dependent contextual fear conditioning in the same animals using the same conditioned and unconditioned stimuli for eyeblink and fear conditioning. In a subset of the behaviorally tested mice, we used unbiased stereology to estimate the total number of Purkinje neurons in cerebellar cortex and pyramidal neurons in the hippocampus. Several forms of synaptic plasticity were assessed at different ages in CBA mice: long-term depression (LTD) in both cerebellum and hippocampus and NMDA-mediated long-term potentiation (LTP) and voltage-dependent calcium channel LTP in hippocampus. Forty-four CBA mice tested at one of five ages (4, 8, 12, 18, or 24 months) demonstrated statistically significant age differences in cerebellum-dependent delay eyeblink conditioning, with 24-month mice showing impairment in comparison with younger mice. These same CBA mice showed no significant differences in contextual or cued fear conditioning. Stereology indicated significant loss of Purkinje neurons in the 18- and 24-month groups, whereas pyramidal neuron numbers were stable across age. Slice electrophysiology recorded from an additional 48 CBA mice indicated significant deficits in LTD appearing in cerebellum between 4 and 8 months, whereas 4- to 12-month mice demonstrated similar hippocampal LTD and LTP values. Our results demonstrate that processes of aging impact brain structures and associated behaviors differentially, with cerebellum showing earlier senescence than hippocampus.

Multiple memory mechanisms in the cerebellum?

Long-term potentiation (LTP) and long-term depression (LTD) are arguably two of the most widely discussed cellular plasticity mechanisms for learning and memory. However, the extent to which they are required for behavioral plasticity and learning is not clear. In this issue of Neuron, Boyden et al. use mice lacking CaMKIV and Hansel et al. use mice lacking alphaCaMKII to assess the contribution of LTD to cerebellar learning.

Extinction of a classically conditioned response: red nucleus and interpositus.

It is well established that the cerebellum and its associated circuitry are essential for classical conditioning of the eyeblink response and other discrete motor responses (e.g., limb flexion, head turn, etc.) learned with an aversive unconditioned stimulus. However, brain mechanisms underlying extinction of these responses are still relatively unclear. Behavioral studies have demonstrated extinction to be an active learning process distinct from acquisition. Accordingly, this current understanding of extinction has guided neural studies that have tried to identify possible brain structures that could support this new learning. However, whether extinction engages the same brain sites necessary for acquisition is not yet clear. This poses an overriding problem for understanding brain mechanisms necessary for extinction because such analysis cannot be done without first identifying brain sites and pathways involved in this phenomenon. Equally elusive is the validity of a behavioral theory of extinction that can account for the properties of extinction. In this study, we looked at the involvement of the interpositus and the red nucleus in extinction. Results show that, although inactivation of both nuclei blocks response expression, only inactivation of the interpositus has a detrimental effect on extinction. Moreover, this detrimental effect was completely removed when inactivation of the interpositus was paired with electrical stimulation of the red nucleus. These findings speak to the important role of cerebellar structures in the extinction of discrete motor responses and provide important insight as to the validity of a particular theory of extinction.

Progesterone receptors: form and function in brain.

Emerging data indicate that progesterone has multiple non-reproductive functions in the central nervous system to regulate cognition, mood, inflammation, mitochondrial function, neurogenesis and regeneration, myelination and recovery from traumatic brain injury. Progesterone-regulated neural responses are mediated by an array of progesterone receptors (PR) that include the classic nuclear PRA and PRB receptors and splice variants of each, the seven transmembrane domain 7TMPRbeta and the membrane-associated 25-Dx PR (PGRMC1). These PRs induce classic regulation of gene expression while also transducing signaling cascades that originate at the cell membrane and ultimately activate transcription factors. Remarkably, PRs are broadly expressed throughout the brain and can be detected in every neural cell type. The distribution of PRs beyond hypothalamic borders, suggests a much broader role of progesterone in regulating neural function. Despite the large body of evidence regarding progesterone regulation of reproductive behaviors and estrogen-inducible responses as well as effects of progesterone metabolite neurosteroids, much remains to be discovered regarding the functional outcomes resulting from activation of the complex array of PRs in brain by gonadally and/or glial derived progesterone. Moreover, the impact of clinically used progestogens and developing selective PR modulators for targeted outcomes in brain is a critical avenue of investigation as the non-reproductive functions of PRs have far-reaching implications for hormone therapy to maintain neurological health and function throughout menopausal aging.

Habituation revisited: an updated and revised description of the behavioral characteristics of habituation.

The most commonly cited descriptions of the behavioral characteristics of habituation come from two papers published almost 40 years ago [Groves, P. M., & Thompson, R. F. (1970). Habituation: A dual-process theory. Psychological Review, 77, 419-450; Thompson, R. F., & Spencer, W. A. (1966). Habituation: A model phenomenon for the study of neuronal substrates of behavior. Psychological Review, 73, 16-43]. In August 2007, the authors of this review, who study habituation in a wide range of species and paradigms, met to discuss their work on habituation and to revisit and refine the characteristics of habituation. This review offers a re-evaluation of the characteristics of habituation in light of these discussions. We made substantial changes to only a few of the characteristics, usually to add new information and expand upon the description rather than to substantially alter the original point. One additional characteristic, relating to long-term habituation, was added. This article thus provides a modern summary of the characteristics defining habituation, and can serve as a convenient primer for those whose research involves stimulus repetition.

The role of the cerebellar interpositus nucleus in short and long term memory for trace eyeblink conditioning.

In previous studies the cerebellar interpositus (IP) nucleus, but not the hippocampus, was shown to be necessary both for initial learning and retention and for long-term retention of the standard delay eyeblink conditioned response (CR). However, in the trace eyeblink CR procedure, the hippocampus is also necessary for initial learning and retention, but not for long-term retention. Here the authors evaluate the role of the IP nucleus in both initial learning and retention, and in long-term retention of the trace eyeblink CR, using muscimol infusion to reversibly inactivate the IP nucleus. For the short-term study, there were two subgroups, the first sequentially passed through acquisition, inactivation, and reacquisition phases, whereas the second subgroup went through inactivation, acquisition, and inactivation phases. For the long-term study, the rabbits acquired the CR and then rested for a month. Next, they were distributed into two subgroups: with or without retention training, and finally went through inactivation and reacquisition phases. The results showed that the prelearning IP nucleus inactivation prevented the acquisition of the trace CR, whereas the postlearning inactivation reversibly abolished the expression of both the short- and long-term CR.

Disruption of cerebellar cortical inhibition in the absence of learning promotes sensory-evoked eyeblink responses.

Theories of cerebellar learning propose that alterations in synaptic plasticity resulting in decreases in cerebellar cortical inhibition and increases in sensory activation of interpositus nuclei underlie the development of adaptively timed conditioned motor responses. The authors found that with concurrent pharmacological disconnection of the cerebellar cortex and intense sensory stimulation in the untrained rabbit, eyeblink responses were generated. Neither sensory stimulation nor disconnection alone generated significant eyeblink responses. These results are consistent with dual plasticity models of cerebellar learning and strongly support the general hypothesis that conditioned responses are the result of strengthening of preexisting connections in the nervous system.

Impaired memory of eyeblink conditioning in CaMKIV KO mice.

The calcium/calmodulin-dependent protein kinase type IV (CaMKIV) is highly expressed in cerebellar cortical granule cells and deep nuclear neurons in the cerebellum. It mediates the phosphorylation and activation of the cAMP-dependent response element binding protein (CREB). In several paradigms CREB-dependent transcription is required for cellular events underlying long-term memory processes. Also, CaMKIV deficiency results in impaired long-term depression (LTD) induction in cerebellar cortex. To investigate the function of CaMKIV in the cerebellum, Wild-type (WT) and CaMKIV KO mice were tested with delay eyeblink conditioning. KO and WT mice did not differ in acquisition, but the KO mice showed a significantly lower conditioned response (CR) percentage than the WT mice in the retention testing and retraining period. The CR peak latencies for the two groups did not differ in acquisition but were shorter for the KO mice in the testing period. No significant differences were found between KO and WT mice in spontaneous eyeblink activity, auditory brainstem response (ABR) amplitudes, and tail-flick latency. The results suggest an important role for CaMKIV in long-term memory in the cerebellum.

Progesterone regulation of synaptic transmission and plasticity in rodent hippocampus.

Ovarian hormones influence memory formation by eliciting changes in neural activity. The effects of various concentrations of progesterone (P4) on synaptic transmission and plasticity associated with long-term potentiation (LTP) and long-term depression (LTD) were studied using in vitro hippocampal slices. Extracellular studies show that the highest concentration of P4 tested (10(-6) M) decreased the baseline synaptic transmission and magnitude of LTP, but did not affect LTD. Intracellular studies suggest the P4 effect to be mediated, at least in part, by GABA(A) activity. These results establish a general effect of P4 on synaptic transmission, multiple forms of synaptic plasticity, and a possible mechanism of P4 action in hippocampus.

Estrogen and hippocampal plasticity in rodent models.

Accumulating evidence indicates that ovarian hormones regulate a wide variety of non-reproductive functions in the central nervous system by interacting with several molecular and cellular processes. A growing animal literature using both adult and aged rodent models indicates that 17beta-estradiol, the most potent of the biologically relevant estrogens, facilitates some forms of learning and memory, in particular those that involve hippocampal-dependent tasks. A recently developed triple-transgenic mouse (3xTg-AD) has been widely used as an animal model of Alzheimer's disease, as this mouse exhibits an age-related and progressive neuropathological phenotype that includes both plaque and tangle pathology mainly restricted to hippocampus, amygdala and cerebral cortex. In this report, we examine recent studies that compare the effects of ovarian hormones on synaptic transmission and synaptic plasticity in adult and aged rodents. A better understanding of the non-reproductive functions of ovarian hormones has far-reaching implications for hormone therapy to maintain health and function within the nervous system throughout aging.

17beta-estradiol modifies stress-induced and age-related changes in hippocampal synaptic plasticity.

The female steroid hormone 17beta-estradiol enhances synaptic transmission and the magnitude of longterm potentiation (LTP) in adult rodent hippocampal slices. Long-term depression (LTD), another form of synaptic plasticity, occurs more prominently in hippocampal slices from aged rodents. A decrease in LTP has been recorded in hippocampal slices from adult rodents behaviorally stressed just before tissue preparation and electrophysiological recording. Here, the authors test the hypothesis that estrogen modifies synaptic plasticity in both adult and aged rodents, whether behaviorally stressed or not. Our results indicate that estrogen enhances LTP and attenuates LTD, thus producing a protective effect against both aging and stress. These results also provide new approaches that can be used to reverse age and stress-related learning and memory dysfunction.

Eye-blink conditioning is associated with changes in synaptic ultrastructure in the rabbit interpositus nuclei.

Eye-blink conditioning involves the pairing of a conditioned stimulus (usually a tone) to an unconditioned stimulus (air puff), and it is well established that an intact cerebellum and interpositus nucleus, in particular, are required for this form of classical conditioning. Changes in synaptic number or structure have long been proposed as a mechanism that may underlie learning and memory, but localizing these changes has been difficult. Thus, the current experiment took advantage of the large amount of research conducted on the neural circuitry that supports eye-blink conditioning by examining synaptic changes in the rabbit interpositus nucleus. Synaptic quantifications included total number of synapses per neuron, numbers of excitatory versus inhibitory synapses, synaptic curvature, synaptic perforations, and the maximum length of the synapses. No overall changes in synaptic number, shape, or perforations were observed. There was, however, a significant increase in the length of excitatory synapses in the conditioned animals. This increase in synaptic length was particularly evident in the concave-shaped synapses. These results, together with previous findings, begin to describe a sequence of synaptic change in the interpositus nuclei following eye-blink conditioning that would appear to begin with structural change and end with an increase in synaptic number.

Inhibiting the expression of a classically conditioned behavior prevents its extinction.

The underlying neuronal substrates and behavioral properties that might mediate extinction of the classically conditioned eye-blink response (CR) were examined. Four groups of rabbits were trained to perform the CR. Two of the groups then received either three or six sessions of tone-alone extinction training while the motor nuclei that mediate expression of the CR (facial nucleus and accessory abducens) were reversibly inactivated with microinjections of the GABA agonist muscimol. After these inactivation extinction sessions, rabbits received four more extinction sessions without inactivation. Two groups of controls received either three or six extinction sessions while saline vehicle was infused into the motor nuclei, followed by four sessions with no infusions. Saline infusions had no effect on extinction, and controls extinguished the CR normally over the first three to four sessions. In contrast, muscimol inactivation of the motor nuclei completely prevented any performance of CRs during the three or six inactivation extinction sessions. At the start of the four extinction sessions without inactivation, rabbits performed CRs at the same rate and amplitude as controls on their first extinction sessions. The muscimol rabbits then extinguished the CR normally over the four sessions without inactivation. In short, inactivation of the motor nuclei completely prevented any extinction of the eye-blink CR with no effect on subsequent extinction without inactivation. These results are discussed in terms of possible neuroanatomical loci that might mediate the extinction process as well as how effects of manipulating CR performance during extinction may affect the extinction process.

Brain-derived neurotrophic factor plays a critical role in contextual fear conditioning.

In this study, brain-derived neurotrophic factor (BDNF) heterozygous knock-outs were tested on fear conditioning, and their wild-type littermates were used as controls. Results showed that BDNF(+/-) mice are impaired in contextual learning, whereas tone learning remains intact. Because BDNF is involved in synaptic transmission and contextual learning is hippocampal dependent, we hypothesized that this deficit is attributable to abnormal BDNF-modulated synaptic plasticity in the hippocampus. A "gain-of-function" experiment was performed next by infusing recombinant BDNF protein into the hippocampal formation to investigate whether this deficit can be rescued. Infusion of BDNF protein into the hippocampus appeared to partially restore contextual fear learning of BDNF(+/-) mice. In conclusion, the present study suggests that BDNF plays a critical role in fear conditioning. Loss of one copy of the BDNF gene leads to impairment of contextual fear learning in BDNF(+/-). This deficit can be partially rescued by infusing BDNF protein into the hippocampus. Other brain regions interacting with the hippocampus in the context conditioned stimulus pathway, for example, the amygdala, may also require normal BDNF expression levels to fully rescue this impairment.

Long-term storage of an associative memory trace in the cerebellum.

Distinct neural regions may be engaged during acquisition and maintenance of some memories. In delay classical conditioning of the eyeblink response, the cerebellum is necessary for acquisition and expression of the conditioned response (CR), but loci of long-term memory storage are not known. Rabbits (Oryctolagus cuniculus) were trained, overtrained, and given either 30 additional days of training or 30 days of rest. Half the subjects in the rest group were given a reminder training session. Subjects then received either reversible inactivation of the cerebellar interpositus nucleus (muscimol) or permanent electrolytic lesions. In all cases, inactivation and lesions of the interpositus completely abolished the CR. The site of memory formation in the interpositus nucleus also appears to be the site of long-term memory storage.

Localization and characterization of an essential associative memory trace in the mammalian brain.

We argue here that we have succeeded in localizing an essential memory trace for a basic form of associative learning and memory - classical conditioning of discrete responses learned with an aversive stimulus - to the anterior interpositus nucleus of the cerebellum. We first identified the entire essential circuit, using eyelid conditioning as the model system, and used reversible inactivation, during training, of critical structures and activation of pathways to localize definitively the essential memory trace. This discovery and the associated studies have: 1) shown that the essential cerebellar circuit applies equally to all mammals studied, including humans; 2) shown that this cerebellar circuit holds for the learning of any discrete behavioral response elicited by an aversive US, not just eyelid closure; 3) identified the essential circuit and process for reinforcement for this form of learning; 4) shown that this form of learning and its essential cerebellar circuitry is phylogenetically very old; 5) solved the long-standing puzzle of where memory traces are formed in the brain when the CS is electrical stimulation of the cerebral cortex in conditioning; 6) shown that this cerebellar circuitry forms the essential neural substrate for the behavioral phenomenon of "blocking", and hence, 7) provides the first clear neural instantiation of the Rescorla-Wagner learning algorithm; 8) shown that the fundamental neural process underlying this form of learning is a strengthening of preexisting pathways, and 9) shown that the basic mechanism underlying this strengthening is the formation of new excitatory synapses. This article is part of a Special Issue entitled SI: Brain and Memory.

Effects of estrogen, age, and calpain on MAP kinase and NMDA receptors in female rat brain.

17-beta-Estradiol (E2), by activating Src and ERK/MAP kinases, enhances NMDA receptor phosphorylation and function. NR2 subunits of NMDA receptors are truncated by calpain, an effect prevented by tyrosine phosphorylation of the subunits. The present study investigated whether E2-mediated activation of ERK and NR2 subunits phosphorylation were altered in 24-month-old female rats. Ovariectomy reduced ERK2 phosphorylation in brains from 3- but not 24-month-old female rats. In ovariectomized rats, restoration of estrogen levels increased ERK2 and NR2 phosphorylation in young but not aged animals. Calcium treatment of frozen-thawed brain sections decreased NR2 levels in both young and aged female rats. This effect was absent in E2-treated young ovariectomized female rats, but was not modified in aged ovariectomized female rats. These results indicate that E2 activation of ERK2 and NR2 phosphorylation is markedly reduced in aged female rats, whereas calpain-mediated truncation of NR2 subunits is not different in young and aged rats. They suggest that several key elements of the mechanisms involved in estrogen-mediated regulation of synaptic plasticity are altered in aged animals.