Control of central auditory processing by a brain-generated oestrogen.

Recent discoveries show that behaviourally relevant sensory experience drives the production of oestradiol - the classic sex steroid oestrogen - in auditory neurons in the adult brain of both males and females. This brain-generated oestrogen markedly enhances the efficiency of the neural coding of acoustic cues and shapes auditory-based behaviours on a timescale that is relevant for sensory processing and congruent with the action of rapid neuromodulators. These findings are re-shaping our current understanding of the mechanistic framework that supports sensory processing and the functional roles of hormones in the brain, and have implications for multiple health issues.

Mechanistic basis and functional roles of long-term plasticity in auditory neurons induced by a brain-generated estrogen.

The classic estrogen 17ß-estradiol (E2) was recently identified as a novel modulator of hearing function. It is produced rapidly, in an experience-dependent fashion, by auditory cortical neurons of both males and females. This brain-generated E2 enhances the efficiency of auditory coding and improves the neural and behavioral discrimination of auditory cues. Remarkably, the effects of E2 are long-lasting and persist for hours after local rises in hormone levels have subsided. The mechanisms and functional consequences of this E2-induced plasticity of auditory responses are unknown. Here, we addressed these issues in the zebra finch model by combining intracerebral pharmacology, biochemical assays, in vivo neurophysiology in awake animals, and computational and information theoretical approaches. We show that auditory experience activates the MAPK pathway in an E2-dependent manner. This effect is mediated by estrogen receptor ß (ERß), which directly associates with MEKK1 to sequentially modulate MEK and ERK activation, where the latter is required for the engagement of downstream molecular targets. We further show that E2-mediated activation of the MAPK cascade is required for the long-lasting enhancement of auditory-evoked responses in the awake brain. Moreover, a functional consequence of this E2/MAPK activation is to sustain enhanced information handling and neural discrimination by auditory neurons for several hours following hormonal challenge. Our results demonstrate that brain-generated E2 engages, via a nongenomic interaction between an estrogen receptor and a kinase, a persistent form of experience-dependent plasticity that enhances the neural coding and discrimination of behaviorally relevant sensory signals in the adult vertebrate brain.

Brain-generated estradiol drives long-term optimization of auditory coding to enhance the discrimination of communication signals.

Auditory processing and hearing-related pathologies are heavily influenced by steroid hormones in a variety of vertebrate species, including humans. The hormone estradiol has been recently shown to directly modulate the gain of central auditory neurons, in real time, by controlling the strength of inhibitory transmission via a nongenomic mechanism. The functional relevance of this modulation, however, remains unknown. Here we show that estradiol generated in the songbird homolog of the mammalian auditory association cortex, rapidly enhances the effectiveness of the neural coding of complex, learned acoustic signals in awake zebra finches. Specifically, estradiol increases mutual information rates, coding efficiency, and the neural discrimination of songs. These effects are mediated by estradiol's modulation of both the rate and temporal coding of auditory signals. Interference with the local action or production of estradiol in the auditory forebrain of freely behaving animals disrupts behavioral responses to songs, but not to other behaviorally relevant communication signals. Our findings directly show that estradiol is a key regulator of auditory function in the adult vertebrate brain.

Estradiol-dependent modulation of auditory processing and selectivity in songbirds.

The steroid hormone estradiol plays an important role in reproductive development and behavior and modulates a wide array of physiological and cognitive processes. Recently, reports from several research groups have converged to show that estradiol also powerfully modulates sensory processing, specifically, the physiology of central auditory circuits in songbirds. These investigators have discovered that (1) behaviorally-relevant auditory experience rapidly increases estradiol levels in the auditory forebrain; (2) estradiol instantaneously enhances the responsiveness and coding efficiency of auditory neurons; (3) these changes are mediated by a non-genomic effect of brain-generated estradiol on the strength of inhibitory neurotransmission; and (4) estradiol regulates biochemical cascades that induce the expression of genes involved in synaptic plasticity. Together, these findings have established estradiol as a central regulator of auditory function and intensified the need to consider brain-based mechanisms, in addition to peripheral organ dysfunction, in hearing pathologies associated with estrogen deficiency.

Neurochemical organization and experience-dependent activation of estrogen-associated circuits in the songbird auditory forebrain.

The classic steroid hormone estradiol is rapidly produced by central auditory neurons in the songbird brain and instantaneously modulates auditory coding to enhance the neural and behavioral discrimination of acoustic signals. Although recent advances highlight novel roles for estradiol in the regulation of central auditory processing, current knowledge on the functional and neurochemical organization of estrogen-associated circuits, as well as the impact of sensory experience in these auditory forebrain networks, remains very limited. Here we show that both estrogen-producing and -sensitive neurons are highly expressed in the caudomedial nidopallium (NCM), the zebra finch analog of the mammalian auditory association cortex, but not other auditory forebrain areas. We further demonstrate that auditory experience primarily engages estrogen-producing, and to a lesser extent, estrogen-responsive neurons in NCM, that these neuronal populations moderately overlap and that acute episodes of sensory experience do not quantitatively affect these circuits. Finally, we show that whereas estrogen-producing cells are neurochemically heterogeneous, estrogen-sensitive neurons are primarily glutamatergic. These findings reveal the neurochemical and functional organization of estrogen-associated circuits in the auditory forebrain, demonstrate their activation and stability in response to sensory experience in behaving animals, and highlight estrogenic circuits as fundamental components of central networks supporting sensory processing.

Estradiol shapes auditory processing in the adult brain by regulating inhibitory transmission and plasticity-associated gene expression.

Estradiol impacts a wide variety of brain processes, including sex differentiation, mood, and learning. Here we show that estradiol regulates auditory processing of acoustic signals in the vertebrate brain, more specifically in the caudomedial nidopallium (NCM), the songbird analog of the mammalian auditory association cortex. Multielectrode recordings coupled with local pharmacological manipulations in awake animals reveal that both exogenous and locally generated estradiol increase auditory-evoked activity in NCM. This enhancement in neuronal responses is mediated by suppression of local inhibitory transmission. Surprisingly, we also found that estradiol is both necessary and sufficient for the induction of multiple mitogen-activated protein kinase (MAPK)-dependent genes thought to be required for synaptic plasticity and memorization of birdsong. Specifically, we show that local blockade of estrogen receptors or aromatase activity in awake birds decrease song-induced MAPK-dependent gene expression. Infusions of estradiol in acoustically isolated birds induce transcriptional activation of these genes to levels comparable with song-stimulated animals. Our results reveal acute and rapid nongenomic functions for estradiol in central auditory physiology and suggest that such roles may be ubiquitously expressed across sensory systems.

Profiling of experience-regulated proteins in the songbird auditory forebrain using quantitative proteomics.

Auditory and perceptual processing of songs are required for a number of behaviors in songbirds such as vocal learning, territorial defense, mate selection and individual recognition. These neural processes are accompanied by increased expression of a few transcription factors, particularly in the caudomedial nidopallium (NCM), an auditory forebrain area believed to play a key role in auditory learning and song discrimination. However, these molecular changes are presumably part of a larger, yet uncharacterized, protein regulatory network. In order to gain further insight into this network, we performed two-dimensional differential in-gel expression (2D-DIGE) experiments, extensive protein quantification analyses, and tandem mass spectrometry in the NCM of adult songbirds hearing novel songs. A subset of proteins was selected for immunocytochemistry in NCM sections to confirm the 2D-DIGE findings and to provide additional quantitative and anatomical information. Using these methodologies, we found that stimulation of freely behaving birds with conspecific songs did not significantly impact the NCM proteome 5 min after stimulus onset. However, following 1 and 3 h of stimulation, a significant number of proteins were consistently regulated in NCM. These proteins spanned a range of functional categories that included metabolic enzymes, cytoskeletal molecules, and proteins involved in neurotransmitter secretion and calcium binding. Our findings suggest that auditory processing of vocal communication signals in freely behaving songbirds triggers a cascade of protein regulatory events that are dynamically regulated through activity-dependent changes in calcium levels.

Postinhibitory rebound spikes are modulated by the history of membrane hyperpolarization in the SCN.

The suprachiasmatic nucleus (SCN) of the hypothalamus regulates biological circadian time thereby directly impacting numerous physiological processes. The SCN is composed almost exclusively of gamma-aminobutyric acid (GABA)ergic neurons, many of which synapse with other GABAergic cells in the SCN to exert an inhibitory influence on their postsynaptic targets for most, if not all, phases of the circadian cycle. The overwhelmingly GABAergic nature of the SCN, along with its internal connectivity properties, provide a strong model to examine how inhibitory neurotransmission generates output signals. In the present work we show that hyperpolarizations that range from 5 to 1000 ms elicit rebound spikes in 63% of all SCN neurons tested in voltage-clamp in the SCN of adult rats and hamsters. In current-clamp recordings, hyperpolarizations led to rebound spike formation in all cells; however, low-amplitude or short-duration current injections failed to consistently activate rebound spikes. Increasing the duration of hyperpolarization from 5 to 1000 ms is strongly and positively correlated with enhanced spike probability. Additionally, the magnitude of hyperpolarization exerts a strong influence on both the amplitude of the spike, as revealed by voltage-clamp recordings, and the latency to peak current obtained in either voltage- or current-clamp mode. Our results suggest that SCN neurons may use rebound spikes as one means of producing output signals from a largely interconnected network of GABAergic neurons.

A songbird forebrain area potentially involved in auditory discrimination and memory formation.

Songbirds rely on auditory processing of natural communication signals for a number of social behaviors,including mate selection,individual recognition and the rare behavior of vocal learning - the ability to learn vocalizations through imitation of an adult model,rather than by instinct. Like mammals,songbirds possess a set of interconnected ascending and descending auditory brain pathways that process acoustic information and that are presumably involved in the perceptual processing of vocal communication signals. Most auditory areas studied to date are located in the caudomedial forebrain of the songbird and include the thalamo-recipient field L (sub fields L1,L2 and L3),the caudomedial and caudolateral mesopallium (CMM and CLM,respectively) and the caudomedial nidopallium (NCM). This review focuses on NCM,an auditory area previously proposed to be analogous to parts of the primary auditory cortex in mammals. Stimulation of songbirds with auditory stimuli drives vigorous electrophysiological responses and the expression of several activity-regulated genes in NCM.Interestingly,NCM neurons are tuned to species-specific songs and undergo some forms of experience-dependent plasticity in-vivo . These activity-dependent changes may underlie long-term modifications in the functional performance of NCM and constitute a potential neural substrate for auditory discrimination. We end this review by discussing evidence that suggests that NCM may be a site of auditory memory formation and/or storage.

Alteration of cingulate long-term plasticity and behavioral sensitization to inflammation by environmental enrichment.

Exposure to an enriched environment (EE) has been shown to induce cortical plasticity. Considerable amount of research is focused on the effects of EE in the hippocampus; however, effects of EE on other brain regions and the mechanisms involved are not well known. To investigate this, we induced cortical plasticity by placing mice in an EE for one month and measured the effects of EE in the anterior cingulate cortex (ACC). Here, we show that EE enhanced the expression of the plasticity gene, egr-1, in the ACC of EE animals accompanied by enhanced cingulate long-term potentiation (LTP) and decreased cingulate long-term depression (LTD). The increased NMDA receptor NR2B/NR2A subunits current ratio is associated with the plasticity seen in the ACC while total protein levels remain unchanged. Furthermore, behavioral experiments show that these mice exposed to EE demonstrate enhanced responses to acute and long-term inflammation. Our findings suggest that exposure to EE alters physiological properties within the ACC which results in enhanced responses to inflammation.

The excitatory thalamo-"cortical" projection within the song control system of zebra finches is formed by calbindin-expressing neurons.

The learning and production of vocalizations in songbirds are controlled by a system of interconnected brain nuclei organized into a direct vocal motor pathway and an anterior forebrain (pallium-basal ganglia-thalamo-pallial) loop. Here we show that the thalamo-pallial ("thalamo-cortical") projection (from the medial part of the dorsolateral thalamic nucleus to the lateral magnocellular nucleus of the anterior nidopallium--DLM to LMAN) within the anterior forebrain loop is composed of cells positive for the calcium-binding protein calbindin. We show that the vast majority of cells within DLM express calbindin, based both on immunocytochemistry (ICC) for calbindin protein and in situ hybridization for calb mRNA. Using a combination of tract-tracing and ICC we show that the neurons that participate in the DLM-to-LMAN projection are calbindin-positive. We also demonstrate that DLM is devoid of cells expressing mRNA for the GABAergic marker zGAD65. This observation confirms that the calbindin-expressing cells in DLM are not GABAergic, in accordance with previous electrophysiological data indicating that the DLM-to-LMAN projection is excitatory. Furthermore, we use ICC to determine the trajectory of the fibers within the DLM-to-LMAN projection, and to demonstrate a sex difference in calbindin expression levels in the fibers of the DLM-to-LMAN projection. Our findings provide a clear-cut neurochemical signature for a critical projection in the songbird vocal control pathways that enable song learning.

GABA immunoreactivity in auditory and song control brain areas of zebra finches.

Inhibitory transmission is critical to sensory and motor processing and is believed to play a role in experience-dependent plasticity. The main inhibitory neurotransmitter in vertebrates, GABA, has been implicated in both sensory and motor aspects of vocalizations in songbirds. To understand the role of GABAergic mechanisms in vocal communication, GABAergic elements must be characterized fully. Hence, we investigated GABA immunohistochemistry in the zebra finch brain, emphasizing auditory areas and song control nuclei. Several nuclei of the ascending auditory pathway showed a moderate to high density of GABAergic neurons including the cochlear nuclei, nucleus laminaris, superior olivary nucleus, mesencephalic nucleus lateralis pars dorsalis, and nucleus ovoidalis. Telencephalic auditory areas, including field L subfields L1, L2a and L3, as well as the caudomedial nidopallium (NCM) and mesopallium (CMM), contained GABAergic cells at particularly high densities. Considerable GABA labeling was also seen in the shelf area of caudodorsal nidopallium, and the cup area in the arcopallium, as well as in area X, the lateral magnocellular nucleus of the anterior nidopallium, the robust nucleus of the arcopallium and nidopallial nucleus HVC. GABAergic cells were typically small, most likely local inhibitory interneurons, although large GABA-positive cells that were sparsely distributed were also identified. GABA-positive neurites and puncta were identified in most nuclei of the ascending auditory pathway and in song control nuclei. Our data are in accordance with a prominent role of GABAergic mechanisms in regulating the neural circuits involved in song perceptual processing, motor production, and vocal learning in songbirds.

Presynaptic Na+ channels: locus, development, and recovery from inactivation at a high-fidelity synapse.

Na+ channel recovery from inactivation limits the maximal rate of neuronal firing. However, the properties of presynaptic Na+ channels are not well established because of the small size of most CNS boutons. Here we study the Na+ currents of the rat calyx of Held terminal and compare them with those of postsynaptic cells. We find that presynaptic Na+ currents recover from inactivation with a fast, single-exponential time constant (24 degrees C, tau of 1.4-1.8 ms; 35 degrees C, tau of 0.5 ms), and their inactivation rate accelerates twofold during development, which may contribute to the shortening of the action potential as the terminal matures. In contrast, recordings from postsynaptic cells in brainstem slices, and acutely dissociated, reveal that their Na+ currents recover from inactivation with a double-exponential time course (tau(fast) of 1.2-1.6 ms; tau(slow) of 80-125 ms; 24 degrees C). Surprisingly, confocal immunofluorescence revealed that Na+ channels are mostly absent from the calyx terminal but are instead highly concentrated in an unusually long (approximately 20-40 microm) unmyelinated axonal heminode. Outside-out patch recordings confirmed this segregation. Expression of Na(v)1.6 alpha-subunit increased during development, whereas the Na(v)1.2alpha-subunit was not present. Serial EM reconstructions also revealed a long pre-calyx heminode, and biophysical modeling showed that exclusion of Na+ channels from the calyx terminal produces an action potential waveform with a shorter half-width. We propose that the high density and polarized locus of Na+ channels on a long heminode are critical design features that allow the mature calyx of Held terminal to fire reliably at frequencies near 1 kHz.

Presynaptic regulation of the inhibitory transmission by GluR5-containing kainate receptors in spinal substantia gelatinosa.

GluR5-containing kainate receptors (KARs) are known to be involved in nociceptive transmission. Our previous work has shown that the activation of presynaptic KARs regulates GABAergic and glycinergic synaptic transmission in cultured dorsal horn neurons. However, the role of GluR5-containing KARs in the modulation of inhibitory transmission in the spinal substantia gelatinosa (SG) in slices remains unknown. In the present study, pharmacological, electrophysiological and genetic methods were used to show that presynaptic GluR5 KARs are involved in the modulation of inhibitory transmission in the SG of spinal slices in vitro. The GluR5 selective agonist, ATPA, facilitated the frequency but not amplitude of spontaneous inhibitory postsynaptic currents (sIPSCs) in SG neurons. ATPA increased sIPSC frequency in all neurons with different firing patterns as delayed, tonic, initial and single spike patterns. The frequency of either GABAergic or glycinergic sIPSCs was significantly increased by ATPA. ATPA could also induce inward currents in all SG neurons recorded. The frequency, but not amplitude, of action potential-independent miniature IPSCs (mIPSCs) was also facilitated by ATPA in a concentration-dependent manner. However, the effect of ATPA on the frequency of either sIPSCs or mIPSCs was abolished in GluR5-/- mice. Deletion of the GluR5 subunit gene had no effect on the frequency or amplitude of mIPSCs in SG neurons. However, GluR5 antagonist LY293558 reversibly inhibited sIPSC and mIPSC frequencies in spinal SG neurons. Taken together, these results suggest that GluR5 KARs, which may be located at presynaptic terminals, contribute to the modulation of inhibitory transmission in the SG. GluR5-containing KARs are thus important for spinal sensory transmission/modulation in the spinal cord.

Plasticity-driven gene expression in the rat retina.

Animals exposed to an enriched environment display features of neural plasticity such as an increased brain volume, enhanced number of dendritic spines, as well as enlarged synapses. Here we report the first description of molecular plasticity in the mammalian retina, as revealed by gene expression. A marked upregulation of both NGFI-A and Arc, two candidate-plasticity genes, was observed in adult rats that had been exposed to an enriched environment for 3 weeks. This increase was paralleled by an increase in the expression of the late genes GAP-43 and Synapsin I, which also indicated changes in retinal connectivity. Our results suggest that both NGFI-A and Arc may regulate mechanisms of plasticity that had been invoked by heightened complexity of the visual environment.

Song-induced gene expression: a window on song auditory processing and perception.

We review here evidence that a large portion of the caudomedial telencephalon of songbirds, distinct from the song control circuit, is involved in the perceptual processing of birdsong. When songbirds hear song, a number of caudomedial pallial areas are activated, as revealed by expression of the activity-dependent gene zenk. These areas, which include field L subfields L1 and L3, as well as the adjacent caudomedial nidopallium (NCM) and caudomedial mesopallium (CMM), are part of the central auditory pathway and constitute a lobule in the caudomedial aspect of the telencephalon. Several lines of evidence indicate that the neural circuits integrating this lobule are capable of performing the auditory processing of song based on fine acoustic features. Thus, this lobule is well positioned to mediate song perceptual processing and discrimination, which are required for vocal communication and vocal learning. Importantly, the zenk gene encodes a transcription factor linked to synaptic plasticity, and it regulates the expression of target genes associated with specific neuronal cell functions. The induction of zenk likely represents a key regulatory event in a gene cascade triggered by song and leading to neuronal plasticity. Thus, zenk may be linked to molecular and cellular mechanisms underlying experience-dependent modification of song-responsive circuits. In summary, songbirds possess an elaborate system for song perceptual processing and discrimination that potentially also subserves song-induced neuronal plasticity and song memory formation. The continued use of a multidisciplinary approach that integrates molecular, anatomical, physiological and behavioral methodologies has the potential to provide further significant insights into the underlying neurobiology of the perceptual aspects of vocal communication and learning.

Age-related distribution of c-fos expression in the striatum of CD-1 mice after acute methylphenidate administration.

Ritalin (methylphenidate hydrochloride, MPH) is the drug of choice for the treatment of attention deficit hyperactivity disorder. Previous research has shown that MPH administration affects the adult brain in a manner different from the young brain. In the current study, we set out to determine the target brain regions of acutely administered MPH at different stages of development. On postnatal days 3, 7, 11, 24, and 45, mice were treated with a single injection (s.c.) of saline, 5 or 20 mg/kg of MPH, and sacrificed 1 h later. Localization of c-fos expression was determined by immunocytochemistry. Compared to saline treated controls, mice treated with the high dose of MPH (20 mg/kg) showed dense Fos-immunoreactivity (Fos-IR) in the striatum. In most cases the low dose of MPH (5 mg/kg) produced only weak c-fos expression that was nearly indistinguishable from saline-treated controls. At PND 3 and 7, Fos-IR was localized in patches in the striatum. This patchy distribution of c-fos positive cells began to decline by PND 11 and was absent in PND 45 mice, with Fos-IR showing a scattered distribution throughout the striatum. The results of this study indicate that MPH induces the expression of c-fos in the same brain regions as cocaine and amphetamine, and that this expression is distributed differentially according to the age of the mouse.

Light-induced Egr-1 expression in the striate cortex of the opossum.

In the present study, immunocytochemistry was used to assess the expression of Egr-1 nuclear protein across selected regions of the opossum visual system. In light-deprived (LD) animals, only a few scattered cell nuclei were found throughout the striate cortex (V1). Exposure to light promoted a significant increase in the density of Egr-1 labeled nuclei in V1. Laminar distribution of immunoreactive nuclei in light-stimulated animals (LS) tended to vary with topography: the lateral region, which corresponds to the central representation of the visual field, appeared to have higher density of cells expressing protein in the supragranular layers, as compared to the medial region, which corresponds to the representation of the peripheral field of vision. Finally, LS animals displayed a narrow band of labeled cell nuclei in the intergeniculate leaflet (IGL) and throughout the anteroposterior extent of the superior colliculus (SC). In contrast, almost no Egr-1 immunolabeling was found in the IGL and SC of LD animals. Our report is the first demonstration of light-regulated expression of the Egr-1 gene in the opossum visual system and provides evidence that the expression of an activity-dependent gene related to neural plasticity is evolutionarily conserved in the visual cortex of the mammalian lineage.