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Depression is a potentially life-threatening mental disorder affecting approximately 300 million people worldwide. Despite much effort, the molecular underpinnings of clinical depression remain poorly defined, and current treatments carry limited therapeutic efficacy and potentially burdensome side effects. Recently, small noncoding RNA molecules known as microRNA (miRNA) have gained prominence as a target for therapeutic intervention, given their capacity to regulate neuronal physiology. Further, mounting evidence suggests a prominent role for miRNA in depressive molecular signaling. Recent studies have demonstrated that dysregulation of miRNA expression occurs in animal models of depression, and in the post-mortem tissue of clinically depressed patients. Investigations into depression-associated miRNA disruption reveals dramatic effects on downstream targets, many of which are thought to contribute to depressive symptoms. Furthermore, selective serotonin reuptake inhibitors, as well as other antidepressant drugs, have the capacity to reverse aberrant depressive miRNA expression and their downstream targets. Given the powerful effects that miRNA have on the central nervous system transcriptome, and the aforementioned studies, there is a compelling rationale to begin to assess the potential contribution of miRNA to depressive etiology. Here, we review the molecular biology of miRNA, our current understanding of miRNA in relation to clinical depression, and the utility of targeting miRNA for antidepressant treatment [ABSTRACT FROM AUTHOR]

The CREB/CRE transcriptional pathway has been implicated in circadian clock timing and light-evoked clock resetting. To date, much of the work on CREB in circadian physiology has focused on how changes in the phosphorylation state of CREB regulate the timing processes. However, beyond changes in phosphorylation, CREB-dependent transcription can also be regulated by the CREB coactivator CRTC (CREB-regulated transcription coactivator), also known as TORC (transducer of regulated CREB). Here we profiled both the rhythmic and light-evoked regulation of CRTC1 and CRTC2 in the murine suprachiasmatic nucleus (SCN), the locus of the master mammalian clock. Immunohistochemical analysis revealed rhythmic expression of CRTC1 in the SCN. CRTC1 expression was detected throughout the dorsoventral extent of the SCN in the middle of the subjective day, with limited expression during early night, and late night expression levels intermediate between mid-day and early night levels. In contrast to CRTC1, robust expression of CRTC2 was detected during both the subjective day and night. During early and late subjective night, a brief light pulse induced strong nuclear accumulation of CRTC1 in the SCN. In contrast with CRTC 1, photic stimulation did not affect the subcellular localization of CRTC2 in the SCN. Additionally, reporter gene profiling and chromatin immunoprecipitation analysis indicated that CRTC1 was associated with CREB in the 5' regulatory region of the periodl gene, and that overexpression of CRTC1 leads to a marked upregulation in periodl transcription. Together, these data raise the prospect that CRTC1 plays a role in fundamental aspects of SCN clock timing and entrainment. [ABSTRACT FROM AUTHOR]

The transcriptional feedback circuit, which is at the core of the suprachiasmatic nucleus (SCN) circadian (i.e., 24 h) clock, is tightly coupled to both external entrainment cues, such as light, as well as rhythmic cues that arise on a system-wide level within the SCN. One potential signaling pathway by which these cues are conveyed to the molecular clock is the CREB/CRE transcriptional cascade. In this study, we employed a tetracycline-inducible CREB repressor mouse strain, in which approximately 60% of the SCN neurons express the transgene, to test CREB functionality in the clock and its effects on overt rhythmicity. We show that attenuated CREB signaling in the SCN led to a significant reduction in light-evoked clock entrainment. An examination of circadian timing revealed that CREB repressor mice exhibited normal free-running rhythms in the absence of external lighting cues. However, under conditions of constant light, which typically leads to a lengthening of the circadian period, CREB repressor mice exhibited a dramatic arrhythmic phenotype, which could be reversed with doxycycline. At a cellular level, the repression of CREB led to a significant reduction in both the expression of the circadian clock proteins PERIOD1 and PERIOD2 and the clock output hormones AVP and VIP. Together, these data support the idea that the CRE transcriptional pathway orchestrates transcriptional events that are essential for both the maintenance of SCN timing and light entrainment of the circadian clock. [ABSTRACT FROM PUBLISHER]

The development of epilepsy is often associated with marked changes in central nervous system cell structure and function. Along these lines, reactive gliosis and granule cell axonal sprouting within the dentate gyrus of the hippocampus are commonly observed in individuals with temporal lobe epilepsy (TLE). Here we used the pilocarpine model of TLE in mice to screen the proteome and phosphoproteome of the dentate gyrus to identify molecular events that are altered as part of the pathogenic process. Using a two-dimensional gel electrophoresis-based approach, followed by liquid chromatography-tandem mass spectrometry, 24 differentially expressed proteins, including 9 phosphoproteins, were identified. Functionally, these proteins were organized into several classes, including synaptic physiology, cell structure, cell stress, metabolism and energetics. The altered expression of three proteins involved in synaptic physiology, actin, profilin 1 and α-synuclein was validated by secondary methods. Interestingly, marked changes in protein expression were detected in the supragranular cell region, an area where robust mossy fibers sprouting occurs. Together, these data provide new molecular insights into the altered protein profile of the epileptogenic dentate gyrus and point to potential pathophysiologic mechanisms underlying epileptogenesis. [ABSTRACT FROM AUTHOR]

Cannabinoids, the primary active agent in drugs of abuse such as marijuana and hashish, tend to generate a distorted sense of time. Here we study the effect of cannabinoids on the brain's circadian clock, the suprachiasmatic nucleus (SCN), using patch clamp and cell-attached electrophysiological recordings, RT-PCR, immunocytochemistry, and behavioral analysis. The SCN showed strong expression of the cannabinoid receptor CB1R, as detected with RT-PCR. SCN neurons, including those using GABAas a transmitter, and axons within the SCN, expressed CB1R immunoreactivity. Behaviorally, cannabinoids did not alter the endogenous free-running circadian rhythm in the mouse brain, but did attenuate the ability of the circadian clock to entrain to light zeitgebers. In the absence of light, infusion of the CB1R antagonist AM251 caused a modest phase shift, suggesting endocannabinoid modulation of clock timing. Interestingly, cannabinoids had no effect on glutamate release from the retinohypothalamic projection, suggesting a direct action of cannabinoids on the retinohypothalamic tract was unlikely to explain the inhibition of the phase shift. Within the SCN, cannabinoids were excitatory by a mechanism based on presynaptic CB1R attenuation of axonal GABA release. These data raise the possibility that the time dissociation described by cannabinoid users may result in part from altered circadian clock function and/or entrainment to environmental time cues. [ABSTRACT FROM AUTHOR]

Inducible gene expression appears to be an essential event that couples light to entrainment of the master mammalian circadian clock located in the suprachiasmatic nucleus (SCN) of the hypothalamus. Recently, we reported that light triggers phase-dependent activation of the mammalian target of rapamycin (mTOR) signaling pathway, a major regulator of protein synthesis, in the SCN, thus raising the possibility that mTOR-evoked mRNA translation contributes to clock entrainment. Here, we used a combination of cellular, molecular, and behavioral assays to address this question. To this end, we show that the in vivo infusion of the mTOR inhibitor rapamycin led to a significant attenuation of the phase-delaying effect of early-night light. Conversely, disruption of mTOR during the late night augmented the phase-advancing effect of light. To assess the role of mTOR signaling within the context of molecular entrainment, the effects of rapamycin on light-induced expression of PERIOD1 and PERIOD2 were examined. At both the early- and late-night time points, abrogation of mTOR signaling led to a significant attenuation of light-evoked PERIOD protein expression. Our results also reveal that light-induced mTOR activation leads to the translation of mRNAs with a 5'-terminal oligopyrimidine tract such as eukaryotic elongation factor 1A and the immediate early gene JunB. Together, these data indicate that the mTOR pathway functions as potent and selective regulator of light-evoked protein translation and SCN clock entrainment. [ABSTRACT FROM AUTHOR]

Formation of reactive oxygen and nitrogen species is a precipitating event in an array of neuropathological conditions. In response to excessive reactive oxygen species (ROS) levels, transcriptionally dependent mechanisms drive the up-regulation of ROS scavenging proteins which, in turn, limit the extent of brain damage. Here, we employed a transgenic approach in which cAMP-response element binding protein (CREB)-mediated transcription is repressed (via A-CREB) to examine the contribution of the CREB/cAMP response element pathway to neuroprotection and its potential role in limiting ROS toxicity. Using the pilocarpine-evoked repetitive seizure model, we detected a marked enhancement of cell death in A-CREB transgenic mice. Paralleling this, there was a dramatic increase in tyrosine nitration (a marker of reactive species formation) in A-CREB transgenic mice. In addition, inducible expression of peroxisome proliferator-activated receptor gamma coactivator-1α was diminished in A-CREB transgenic mice, as was activity of complex I of the mitochondrial electron transport chain. Finally, the neuroprotective effect of brain-derived neurotrophic factor (BDNF) against ROS-mediated cell death was abrogated by disruption of CREB-mediated transcription. Together, these data both extend our understanding of CREB functionality and provide in vivo validation for a model in which CREB functions as a pivotal upstream integrator of neuroprotective signaling against ROS-mediated cell death. [ABSTRACT FROM AUTHOR]

Cannabinoids modulate energy homeostasis and decrease cognitive arousal, possibly by acting on hypothalamic neurons including those that synthesize melanin-concentrating hormone(MCH)or hypocretin/orexin. Using patch-clamp recordings, we compared the actions of cannabinoid agonists and antagonists on identified MCH or hypocretin neurons in green fluorescent protein-expressing transgenic mice. The cannabinoid type-1 receptor (CB1R) agonist R-(+)-[2,3-dihydro-5-methyl-3-(4-morpho linylmethyl)pyrrolo[1,2,3-de]-1,4-benzoxazin-6-yl]-1-naphthalenylmethanone mesylate (WIN55,212,2) depolarized MCH cells and increased spike frequency; in contrast, WIN55,212,2 hyperpolarized and reduced spontaneous firing of the neighboring hypocretin cells, both results consistent with reduced activity seen with intracerebral cannabinoid infusions. These effects were prevented by AM251 [N-(piperidin-1-yl)-5-(4-iodophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide], a CB1R antagonist, and by tetrodotoxin, suggesting no postsynaptic effect on either neuron type. In MCH cells, depolarizing WIN55,212,2 actions were abolished by the GABAA receptor antagonist bicuculline, suggesting that the CB1R-mediated depolarization was attributable to reduced synaptic GABA release. WIN55,212,2 decreased spontaneous IPSCs, reduced the frequency but not amplitude of miniature IPSCs, and reduced electrically evoked synaptic currents in MCHcells. Glutamate microdrop experiments suggest that WIN55,212,2 acted on axons arising from lateral hypothalamus local inhibitory cells that innervate MCH neurons. In hypocretin neurons, the reduced spike frequency induced by WIN55,212,2 was attributable to presynaptic attenuation of glutamate release; CB1R agonists depressed spontaneous and evoked glutamatergic currents and reduced the frequency of miniature EPSCs. Cannabinoid actions on hypocretin neurons were abolished by ionotropic glutamate receptor antagonists. Together, these results show that cannabinoids have opposite effects on MCHand hypocretin neurons. These opposing actions could help explain the increase in feeding and reduction in arousal induced by cannabinoids. [ABSTRACT FROM AUTHOR]

Excitotoxic cell death is one of the precipitating events in the development of temporal lobe epilepsy. Of particular prominence is the loss of GABAergic hilar neurons. Although the molecular mechanisms responsible for the selective vulnerability of these cells are not well understood, activation of the extracellular signal-regulated kinase/mitogen-activated protein kinase (ERK/MAPK) pathway has been implicated in neuroprotective responses to excitotoxicity in other neuronal populations. Here, we report that high levels of the striatal-enriched protein tyrosine phosphatase (STEP), a key regulator of ERK/MAPK signaling, are found in vulnerable somatostatin-immunoreactive hilar interneurons. Under both control conditions and after pilocarpine-induced status epilepticus (SE), ERK/MAPK activation was repressed in STEP-immunoreactive hilar neurons. This contrasts with robust SE-induced ERK/MAPK activation in the granule cell layer of the dentate gyrus, a cell region that does not express STEP. During pilocarpine-induced SE, in vivo disruption of STEP activity allowed activation of the MAPK pathway, leading to immediate-early gene expression and significant rescue from cell death. Thus, STEP increases the sensitivity of neurons to SE-induced excitotoxicity by specifically blocking a latent neuroprotective response initiated by the MAPK pathway. These findings identify a key set of signaling events that render somatostatinergic hilar interneurons vulnerable to SE-induced cell death. [ABSTRACT FROM AUTHOR]

The mammalian master clock, located in the suprachiasmatic nucleus (SCN), is exquisitely sensitive to photic timing cues, but the key molecular events that sculpt both the phasing and magnitude of responsiveness are not understood. Here, we show that the Ras-like G-protein Dexras1 is a critical factor in these processes. Dexras1-deficient mice (dexras1-/-) exhibit a restructured nighttime phase response curve and a loss of gating to photic resetting during the day. Dexras1 affects the photic sensitivity by repressing or activating time-of-day-specific signaling pathways that regulate extracellular signal-regulated kinase (ERK)/mitogen-activated protein kinase (MAPK). During the late night, Dexras1 limits the capacity of pituitary adenylate cyclase (PAC) activating peptide (PACAP)/PAC1 to affect ERK/MAPK, and in the early night, light-induced phase delays, which are mediated predominantly by NMDA receptors, are reduced as reported previously. Daytime photic phase advances are mediated by a novel signaling pathway that does not affect the SCN core but rather stimulates ERK/MAPK in the SCN shell and triggers downregulation of clock protein expression. [ABSTRACT FROM AUTHOR]

Signaling via the p42/44 mitogen-activated protein kinase (MAPK) pathway has been shown to be a key intracellular signaling event that couples light to entrainment of the mammalian circadian clock located in the suprachiasmatic nucleus (SCN). Because many of the physiological effects of the MAPK pathway are mediated by extracellular signal-regulated kinase (ERK)-regulated kinases, it was of interest to identify kinase targets of ERK in the SCN. In this study, we examined whether mitogen- and stress-activated protein kinase 1 (MSK1) is a downstream target of ERK in the SCN and whether it couples to clock gene expression. Here we show that photic stimulation during the subjective night stimulates MSK1 phosphorylation at serine 360, an event required for robust kinase activation. Activated ERK and MSK1 were colocalized in SEN cell nuclei after photic stimulation. The in vivo administration of the MAP kinase kinase 1/2 inhibitor U0126 [1,4-diamino-2,3-dicyano-1,4-bis(o-aminophenylmercapto)butadiene] attenuated MSK1 phosphorylation. MSK1 phosphorylation was more responsive to late-night than early-night photic stimulation, indicating that MSK1 may differentially contribute to light-induced phase advancing and phase delaying of the clock. The potential connection between pituitary adenylate cyclase-activating polypeptide (PACAP) (a regulator of clock entrainment) and MSK1 phosphorylation was examined. PACAP infusion stimulated MSK1 phosphorylation, whereas PACAP receptor antagonist infusion attenuated light-induced MSK1 phosphorylation in the SCN. In reporter gene assays, MSK1 was shown to couple to mPeriod1 via a cAMP response element-binding protein-dependent mechanism. Together, these data identify MSK1 as both a downstream target of the MAPK cascade within the SCN and a regulator of clock gene expression. [ABSTRACT FROM AUTHOR]

Activation of the transcription factor cAMP response element-binding protein (CREB) by neurotrophins is believed to regulate the survival, differentiation, and maturation of neurons in the CNS and PNS. Although phosphorylation of Ser133 is critical for the expression of CREB-regulated genes, the identity of neurotrophin-regulated Ser133 kinases has remained controversial. We show here that neurotrophin-induced CREB phosphorylation in CNS neurons depends exclusively on the extracellular signal-regulated kinase ½-activated kinase mitogen- and stress-activated protein kinase 1 (MSK1). Small interfering RNA directed against ribosomal S6 (RSK1) and RSK2 reduced phosphorylation of a RSK substrate but did not effect CREB-dependent transcription. However, expression of a selective inhibitory MSK1 mutant markedly attenuated BDNF-stimulated CREB phosphorylation and CREB-mediated transcription. Moreover, the ability of neurotrophins to stimulate CREB phosphorylation was abolished in CNS neurons from MSK1 knock-out mice. Consistent with a role for MSK1 in Ser133 phosphorylation, neurotrophin-induced expression of CREB-regulated genes was attenuated in MSK-deficient neurons. These results indicate that MSK1 is the major neurotrophin-activated Ser133 kinase in CNS neurons. [ABSTRACT FROM AUTHOR]

Disruption of cAMP response element (CRE)-dependent transcription has been hypothesized to contribute to neuronal death and dysfunction in Huntington's disease (HD) and other polyglutamine repeat disorders. Whether dysregulation of CRE-dependent transcription actually occurs in vivo in response to expression of expanded polyglutamine repeats has not been tested. We directly tested whether CRE-dependent transcription is affected in vivo by cross breeding a transgenic mouse model of HD (line R6/2) with a transgenic mouse that expresses a CRE-regulated reporter gene. Instead of compromised CRE-dependent transcription in HD mice, we found a robust upregulation of CRE-dependent transcription in several brain regions (striatum, hippocampus, cortex). CRE-mediated transcription was also evoked by striatal forskolin infusion and by photic stimulation in HD animals. Increased cAMP response element-binding protein (CREB) phosphorylation and elevated levels of the CREB-regulated gene product, CCAAT/enhancer binding protein β, were also found in HD mice. Significant alterations in CREB binding protein expression and localization were not observed in symptomatic R6/2 mice. Thus, rather than repressing CRE-mediated transcription, mutant huntingtin appears to facilitate transcription via a CRE-dependent mechanism in vivo. [ABSTRACT FROM AUTHOR]

The hippocampus is critical for some forms of memory and spatial navigation, but previous research has mostly neglected the CA2, a unique region situated between CA3 and CA1. Here, we show that CA2 pyramidal neurons have distinctive physiological characteristics that include an unprecedented synaptic stability. Although basal synaptic currents in CA1 and CA2 are quite similar, synaptic plasticity including long-term potentiation and long-term depression is absent or less likely to be induced with conventional methods of stimulation in CA2. We also find that CA2 neurons have larger leak currents and more negative resting membrane potentials than CA1 neurons, and consequently, more current is required for action potential generation in CA2 neurons. These data suggest that the molecular "conspiracy against plasticity" in CA2 makes it functionally distinct from the other hippocampal CA regions. This work provides critical insight into hippocampal function and may lead to an understanding of the resistance of CA2 to damage from disease, trauma, and hypoxia. [ABSTRACT FROM AUTHOR]

A large body of literature has shown that the disruption of circadian clock timing has profound effects on mood, memory and complex thinking. Central to this time keeping process is the master circadian pacemaker located within the suprachiasmatic nucleus (SCN). Of note, within the central nervous system, clock timing is not exclusive to the SCN, but rather, ancillary oscillatory capacity has been detected in a wide range of cell types and brain regions, including forebrain circuits that underlie complex cognitive processes. These observations raise questions about the hierarchical and functional relationship between the SCN and forebrain oscillators, and, relatedly, about the underlying clock-gated synaptic circuitry that modulates cognition. Here, we utilized a clock knockout strategy in which the essential circadian timing gene Bmal1 was selectively deleted from excitatory forebrain neurons, whilst the SCN clock remained intact, to test the role of forebrain clock timing in learning, memory, anxiety, and behavioral despair. With this model system, we observed numerous effects on hippocampus-dependent measures of cognition. Mice lacking forebrain Bmal1 exhibited deficits in both acquisition and recall on the Barnes maze. Notably, loss of forebrain Bmal1 abrogated time-of-day dependent novel object location memory. However, the loss of Bmal1 did not alter performance on the elevated plus maze, open field assay, and tail suspension test, indicating that this phenotype specifically impairs cognition but not affect. Together, these data suggest that forebrain clock timing plays a critical role in shaping the efficiency of learning and memory retrieval over the circadian day. [ABSTRACT FROM AUTHOR]

Major depressive disorders are common and disabling conditions associated with significant psychosocial impairment and suicide risk. At least 3-4 % of all depressive individuals die by suicide. Evidence suggests that small non-coding RNAs, in particular microRNAs (miRNAs), play a critical role in major affective disorders as well as suicide. We performed a detailed review of the current literature on miRNAs and their targets in major depression and related disorders as well as suicidal behavior, with a specific focus on miR-185 and miR-491-3p, which have been suggested to participate in the pathogenesis of major depression and/or suicide. miRNAs play a fundamental role in the development of the brain. Several miRNAs are reported to influence neuronal and circuit formation by negatively regulating gene expression. Global miRNA reduced expression was found in the prefrontal cortex of depressed suicide completers when compared to that of nonpsychiatric controls who died of other causes. One particular miRNA, miR-185, was reported to regulate TrkB-T1, which has been associated with suicidal behavior upon truncation. Furthermore, cAMP response element-binding protein-brain-derived neurotrophic factor pathways may regulate, through miRNAs, the homeostasis of neural and synaptic pathways playing a crucial role in major depression. miRNAs have gained attention as key players involved in nervous system development, physiology, and disease. Further evidence is needed to clarify the exact role that miRNAs play in major depression and related disorders and suicidal behavior. [ABSTRACT FROM AUTHOR]

Within the central nervous system, microRNAs have emerged as important effectors of an array of developmental, physiological, and cognitive processes. Along these lines, the CREB-regulated microRNA miR-132 has been shown to influence neuronal maturation via its effects on dendritic arborization and spinogenesis. In the mature nervous system, dysregulation of miR-132 has been suggested to play a role in a number of neurocognitive disorders characterized by aberrant synaptogenesis. However, little is known about the inducible expression and function of miR-132 under normal physiological conditions in vivo. Here, we begin to explore this question within the context of learning and memory. Using in situ hybridization, we show that the presentation of a spatial memory task induced a significant ~1.5-fold increase in miR-132 expression within the CA1, CA3, and GCL excitatory cell layers of the hippocampus. To examine the role of miR-132 in hippocampal-dependent learning and memory, we employ a doxycycline-regulated miR-132 transgenic mouse strain to drive varying levels of transgenic miR-132 expression. These studies revealed that relatively low levels of transgenic miR-132 expression, paralleling the level of expression in the hippocampus following a spatial memory task, significantly enhanced cognitive capacity. In contrast, higher (supra-physiological) levels of miR-132 (>3-fold) inhibited learning. Interestingly, both the impaired cognition and elevated levels of dendritic spines resulting from supra-physiological levels of transgenic miR-132 were reversed by doxycycline suppression of transgene expression. Together, these data indicate that miR-132 functions as a key activity-dependent regulator of cognition, and that miR-132 expression must be maintained within a limited range to ensure normal learning and memory formation. [ABSTRACT FROM AUTHOR]

The master circadian clock in mammals, the suprachiasmatic nucleus ( SCN), is under the entraining influence of the external light cycle. At a mechanistic level, intracellular signaling via the p42/44 mitogen-activated protein kinase pathway appears to play a central role in light-evoked clock entrainment; however, the precise downstream mechanisms by which this pathway influences clock timing are not known. Within this context, we have previously reported that light stimulates activation of the mitogen-activated protein kinase effector mitogen-stress-activated kinase 1 ( MSK1) in the SCN. In this study, we utilised MSK1−/− mice to further investigate the potential role of MSK1 in circadian clock timing and entrainment. Locomotor activity analysis revealed that MSK1 null mice entrained to a 12 h light/dark cycle and exhibited circadian free-running rhythms in constant darkness. Interestingly, the free-running period in MSK1 null mice was significantly longer than in wild-type control animals, and MSK1 null mice exhibited a significantly greater variance in activity onset. Further, MSK1 null mice exhibited a significant reduction in the phase-delaying response to an early night light pulse (100 lux, 15 min), and, using an 8 h phase-advancing 'jet-lag' experimental paradigm, MSK1 knockout animals exhibited a significantly delayed rate of re-entrainment. At the molecular level, early night light-evoked cAMP response element-binding protein (CREB) phosphorylation, histone phosphorylation and Period1 gene expression were markedly attenuated in MSK1−/− animals relative to wild-type mice. Together, these data provide key new insights into the molecular mechanisms by which MSK1 affects the SCN clock. [ABSTRACT FROM AUTHOR]

The neurogenic niche within the subgranular zone ( SGZ) of the dentate gyrus is a source of new neurons throughout life. Interestingly, SGZ proliferative capacity is regulated by both physiological and pathophysiological conditions. One outstanding question involves the molecular mechanisms that regulate both basal and inducible adult neurogenesis. Here, we examined the role of the MAPK-regulated kinases, mitogen- and stress-activated kinase ( MSK)1 and MSK2. as regulators of dentate gyrus SGZ progenitor cell proliferation and neurogenesis. Under basal conditions, MSK1/2 null mice exhibited significantly reduced progenitor cell proliferation capacity and a corollary reduction in the number of doublecortin ( DCX)-positive immature neurons. Strikingly, seizure-induced progenitor proliferation was totally blocked in MSK1/2 null mice. This blunting of cell proliferation in MSK1/2 null mice was partially reversed by forskolin infusion, indicating that the inducible proliferative capacity of the progenitor cell population was intact. Furthermore, in MSK1/2 null mice, DCX-positive immature neurons exhibited reduced neurite arborization. Together, these data reveal a critical role for MSK1/2 as regulators of both basal and activity-dependent progenitor cell proliferation and morphological maturation in the SGZ. [ABSTRACT FROM AUTHOR]

Murine epidermal stem cells undergo alternate cycles of dormancy and activation, fuelling tissue renewal. However, only a subset of stem cells becomes active during each round of morphogenesis, indicating that stem cells coexist in heterogeneous responsive states. Using a circadian-clock reporter-mouse model, here we show that the dormant hair-follicle stem cell niche contains coexisting populations of cells at opposite phases of the clock, which are differentially predisposed to respond to homeostatic cues. The core clock protein Bmal1 modulates the expression of stem cell regulatory genes in an oscillatory manner, to create populations that are either predisposed, or less prone, to activation. Disrupting this clock equilibrium, through deletion of Bmal1 (also known as Arntl) or Per1/2, resulted in a progressive accumulation or depletion of dormant stem cells, respectively. Stem cell arrhythmia also led to premature epidermal ageing, and a reduction in the development of squamous tumours. Our results indicate that the circadian clock fine-tunes the temporal behaviour of epidermal stem cells, and that its perturbation affects homeostasis and the predisposition to tumorigenesis. [ABSTRACT FROM AUTHOR]

Activity-regulated gene expression is believed to play a key rote in the development and refinement of neuronal circuitry. Nevertheless, the transcriptional networks that regulate synapse growth and plasticity remain largely uncharacterized. Here, we show that microRNA 132 (miR132) is an activity-dependent rapid response gene regulated by the CAMP response element-binding (CREB) protein pathway. Introduction of miR132 into hippocampal neurons enhanced dendrite morphogenesis whereas inhibition of miR132 by 2'O-methyl RNA antagonists blocked these effects. Furthermore, neuronal activity inhibited translation of p250GAP, a miR132 target, and siRNA-mediated knockdown of p250GAP mimicked miR132-induced dendrite growth. Experiments using dominant-interfering mutants suggested that Rac signaling is downstream of miR132 and p250GAP. We propose that the miR132-p250GAP pathway plays a key role in activity-dependent structural and functional plasticity. [ABSTRACT FROM AUTHOR]

Abstract: Status epilepticus (SE) triggers neuronal death, reactive gliosis and remodeling of synaptic circuitry, thus leading to profound pathological alterations in CNS physiology. These processes are, in part, regulated by the rapid upregulation of both cytotoxic and cytoprotective genes. One pathway that may couple SE to transcriptionally dependent alterations in CNS physiology is the CREB (cAMP response element-binding protein)/CRE (cAMP response element) cascade. Here, we utilized the pilocarpine model of SE on a mouse strain transgenic for a CRE-reporter construct (β-galactosidase) to begin to characterize how seizure activity regulates the activation state of the CREB/CRE pathway in both glia and neurons of the hippocampus. SE triggered a rapid (4–8 h post-SE) but transient increase in CRE-mediated gene expression in the neuronal sublayers. In contrast to neurons, SE induced a lasting increase (up to 20 days) in CRE-mediated transcription in both reactive astrocytes and microglia. CRE-mediated gene expression correlated with expression of the pro-inflammatory enzyme cyclooxygenase-2 (COX-2). To examine the role of CREB in SE-induced COX-2 expression, we generated a transgenic mouse strain that expresses A-CREB, a potent repressor of CREB-dependent transcription. In these animals, the capacity of SE to stimulate COX-2 expression was markedly attenuated, indicating that CREB is a key intermediate in SE-induced COX-2 expression. Collectively these data show that SE triggers two waves of CREB-mediated gene expression, a transient wave in neurons and a long-lasting wave in reactive glial cells, and that CREB couples SE to COX-2 expression. [Copyright &y& Elsevier]

The Ribosomal S6 Kinase (RSK) family of serine/threonine kinases function as key downstream effectors of the MAPK signaling cascade. In the nervous system, RSK signaling plays crucial roles in neuronal development and contributes to activity-dependent neuronal plasticity. This study examined the role of RSK signaling in cell viability during neuronal development and in neuroprotection in the mature nervous system. Using neuronal cell-culture-based profiling, we found that suppressing RSK signaling led to significant cell death in developing primary neuronal cultures. To this end, treatment with the RSK inhibitors BiD1870 or SL0101 on the first day of culturing resulted in over 80% cell death. In contrast, more mature cultures showed attenuated cell death upon RSK inhibition. Inhibition of RSK signaling during early neuronal development also disrupted neurite outgrowth and cell growth. In maturing hippocampal explant cultures, treatment with BiD1870 had minimal effects on cell viability, but led to a striking augmentation of NMDA-induced cell death. Finally, we used the endothelin 1 (ET-1) model of ischemia to examine the neuroprotective effects of RSK signaling in the mature hippocampus in vivo. Notably, in the absence of RSK inhibition, the granule cell layer (GCL) was resistant to the effects of ET-1; However, disruption of RSK signaling (via the microinjection of BiD1870) prior to ET-1 injection triggered cell death within the GCL, thus indicating a neuroprotective role for RSK signaling in the mature nervous system. Together these data reveal distinct, developmentally-defined, roles for RSK signaling in the nervous system., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 International Brain Research Organization (IBRO). Published by Elsevier Inc. All rights reserved.)

Microglia undergo two-stage activation in neurodegenerative diseases, known as disease-associated microglia (DAM). TREM2 mediates the DAM2 stage transition, but what regulates the first DAM1 stage transition is unknown. We report that glucose dyshomeostasis inhibits DAM1 activation and PKM2 plays a role. As in tumors, PKM2 was aberrantly elevated in both male and female human AD brains, but unlike in tumors, it is expressed as active tetramers, as well as among TREM2 + microglia surrounding plaques in 5XFAD male and female mice. snRNAseq analyses of microglia without Pkm2 in 5XFAD mice revealed significant increases in DAM1 markers in a distinct metabolic cluster, which is enriched in genes for glucose metabolism, DAM1, and AD risk. 5XFAD mice incidentally exhibited a significant reduction in amyloid pathology without microglial Pkm2 Surprisingly, microglia in 5XFAD without Pkm2 exhibited increases in glycolysis and spare respiratory capacity, which correlated with restoration of mitochondrial cristae alterations. In addition, in situ spatial metabolomics of plaque-bearing microglia revealed an increase in respiratory activity. These results together suggest that it is not only glycolytic but also respiratory inputs that are critical to the development of DAM signatures in 5XFAD mice., Competing Interests: The authors declare no competing financial interests., (Copyright © 2024 the authors.)

Previous work in the suprachiasmatic nucleus (SCN), the locus of the principal circadian clock, has shown that the activation state of the ERK/MAPK effector p90 ribosomal S6 kinase (RSK) is responsive to photic stimulation and is modulated across the circadian cycle. These data raise the prospect that RSK signaling contributes to both SCN clock timing and entrainment. Here, we found marked expression of the three main RSK isoforms (RSK1/2/3) within the SCN of C57/Bl6 mice. Further, using a combination of immunolabeling and proximity ligation assays, we show that photic stimulation led to the dissociation of RSK from ERK and the translocation of RSK from the cytoplasm to the nucleus. To test for RSK functionality following light treatment, animals received an intraventricular infusion of the selective RSK inhibitor, SL0101, 30 min prior to light (100 lux) exposure during the early circadian night (circadian time 15). Notably, the disruption of RSK signaling led to a significant reduction (∼45 min) in the phase delaying effects of light, relative to vehicle-infused mice. To test the potential contribution of RSK signaling to SCN pacemaker activity, slice cultures from a per1-Venus circadian reporter mouse line were chronically treated with SL0101. Suppression of RSK signaling led to a significant lengthening of the circadian period (∼40 min), relative to vehicle-treated slices. Together, these data reveal that RSK functions as a signaling intermediate that regulates light-evoked clock entrainment and the inherent time keeping properties of the SCN., (Copyright © 2023 IBRO. Published by Elsevier Ltd. All rights reserved.)

MicroRNAs (miRNA, miR-) play important roles in disease development. In this study, we identified an anti-proliferative miRNA, miR-212-5p, that is induced in pulmonary artery smooth muscle cells (PASMCs) and lungs of pulmonary hypertension (PH) patients and rodents with experimental PH. We found that smooth muscle cell (SMC)-specific knockout of miR-212-5p exacerbated hypoxia-induced pulmonary vascular remodeling and PH in mice, suggesting that miR-212-5p may be upregulated in PASMCs to act as an endogenous inhibitor of PH, possibly by suppressing PASMC proliferation. Extracellular vesicles (EVs) have been shown recently to be promising drug delivery tools for disease treatment. We generated endothelium-derived EVs with an enriched miR-212-5p load, 212-eEVs, and found that they significantly attenuated hypoxia-induced PH in mice and Sugen/hypoxia-induced severe PH in rats, providing proof of concept that engineered endothelium-derived EVs can be used to deliver miRNA into lungs for treatment of severe PH., Competing Interests: The authors declare no competing interests., (© 2022 The Authors.)

Modulation of basic biochemical and physiological processes by the circadian timing system is now recognized as a fundamental feature of all mammalian organ systems. Within the central nervous system, these clock-modulating effects are reflected in some of the most complex behavioral states including learning, memory, and mood. How the clock shapes these behavioral processes is only now beginning to be realized. In this review we describe recent findings regarding the complex set of cellular signaling events, including kinase pathways, gene networks, and synaptic circuits that are under the influence of the clock timing system and how this, in turn, shapes cognitive capacity over the circadian cycle. Further, we discuss the functional roles of the master circadian clock located in the suprachiasmatic nucleus, and peripheral oscillator populations within cortical and limbic circuits, in the gating of synaptic plasticity and memory over the circadian cycle. These findings are then used as the basis to discuss the connection between clock dysregulation and cognitive impairments resulting from Alzheimer's disease (AD). In addition, we discuss the conceptually novel idea that in AD, there is a selective disruption of circadian timing within cortical and limbic circuits, and that it is the disruption/desynchronization of these regions from the phase-entraining effects of the SCN that underlies aspects of the early- and mid-stage cognitive deficits in AD. Further, we discuss the prospect that the disruption of circadian timing in AD could produce a self-reinforcing feedback loop, where disruption of timing accelerates AD pathogenesis (e.g., amyloid deposition, oxidative stress and cell death) that in turn leads to a further disruption of the circadian timing system. Lastly, we address potential therapeutic approaches that could be used to strengthen cellular timing networks and, in turn, how these approaches could be used to improve cognitive capacity in Alzheimer's patients., (© 2022. The Author(s).)

Cancer patients experience circadian rhythm disruptions in activity cycles and cortisol release that correlate with poor quality of life and decreased long-term survival rates. However, the extent to which chemotherapy contributes to altered circadian rhythms is poorly understood. In the present study, we examined the extent to which paclitaxel, a common chemotherapy drug, altered entrained and free-running circadian rhythms in wheel running behavior, circulating corticosterone, and circadian clock gene expression in the brain and adrenal glands of tumor-free mice. Paclitaxel injections delayed voluntary wheel running activity onset in a light-dark cycle (LD) and lengthened the free-running period of locomotion in constant darkness (DD), indicating an effect on inherent suprachiasmatic nucleus (SCN) pacemaker activity. Paclitaxel attenuated clock gene rhythms in multiple brain regions in LD and DD. Furthermore, paclitaxel disrupted circulating corticosterone rhythms in DD by elevating its levels across a 24-hour cycle, which correlated with blunted amplitudes of Arntl, Nr1d1, Per1, and Star rhythms in the adrenal glands. Paclitaxel also shortened SCN slice rhythms, increased the amplitude of adrenal gland oscillations in PER2::luciferase cultures, and increased the concentration of pro-inflammatory cytokines and chemokines released from the SCN. These findings indicate that paclitaxel disrupts clock genes and behavior driven by the SCN, other brain regions, and adrenal glands, which were associated with chemotherapy-induced inflammation. Together, this preclinical work demonstrates that chemotherapy disrupts both central and peripheral circadian rhythms and supports the possibility that targeted circadian realignment therapies may be a novel and non-invasive way to improve patient outcomes after chemotherapy., (Copyright © 2021 Elsevier Inc. All rights reserved.)

The mammalian master circadian pacemaker within the suprachiasmatic nucleus (SCN) maintains tight entrainment to the 24 hr light/dark cycle via a sophisticated clock-gated rhythm in the responsiveness of the oscillator to light. A central event in this light entrainment process appears to be the rapid induction of gene expression via the ERK/MAPK pathway. Here, we used RNA array-based profiling in combination with pharmacological disruption methods to examine the contribution of ERK/MAPK signaling to light-evoked gene expression. Transient photic stimulation during the circadian night, but not during the circadian day, triggered marked changes in gene expression, with early-night light predominately leading to increased gene expression and late-night light predominately leading to gene downregulation. Functional analysis revealed that light-regulated genes are involved in a diversity of physiological processes, including DNA transcription, RNA translation, mRNA processing, synaptic plasticity and circadian timing. The disruption of MAPK signaling led to a marked reduction in light-evoked gene regulation during the early night (32/52 genes) and late night (190/191 genes); further, MAPK signaling was found to gate gene expression across the circadian cycle. Together, these experiments reveal potentially important insights into the transcriptional-based mechanisms by which the ERK/MAPK pathway regulates circadian clock timing and light-evoked clock entrainment., Competing Interests: The authors have declared that no competing interests exist.

Newborns exposed to prenatal opioids often experience intense postnatal withdrawal after cessation of the opioid, called neonatal opioid withdrawal syndrome (NOWS), with limited pre- and postnatal therapeutic options available. In a prior study in pregnant mice we demonstrated that the peripherally selective opioid antagonist, 6β-naltrexol (6BN), is a promising drug candidate for preventive prenatal treatment of NOWS, and a therapeutic mechanism was proposed based on preferential delivery of 6BN to fetal brain with relative exclusion from maternal brain. Here, we have developed methadone (MTD) treated pregnant guinea pigs as a physiologically more suitable model, enabling detection of robust spontaneous neonatal withdrawal. Prenatal MTD significantly aggravates two classic maternal separation stress behaviors in newborn guinea pigs: calling (vocalizing) and searching (locomotion) - natural attachment behaviors thought to be controlled by the endogenous opioid system. In addition, prenatal MTD significantly increases the levels of plasma cortisol in newborns, showing that cessation of MTD at birth engages the hypothalamic-pituitary-adrenal (HPA) axis. We find that co-administration of 6BN with MTD prevents these withdrawal symptoms in newborn pups with extreme potency (ID50 ∼0.02 mg/kg), at doses unlikely to induce maternal or fetal withdrawal or to interfere with opioid antinociception based on many prior studies in rodents and non-human primates. Furthermore, we demonstrate a similarly high potency of 6BN in preventing opioid withdrawal in adult guinea pigs (ID50 = 0.01 mg/kg). This high potency appears to run counter to our pharmacokinetic studies showing slow 6BN transit of both the placenta and maternal blood brain barrier in guinea pigs, and calls into question the preferential delivery mechanism. Rather, it suggests a novel receptor mechanism to account for the selectively high potency of 6BN to suppress opioid dependence at all developmental stages, even in adults, as compared to its well-established low potency as a classical opioid antagonist. In conclusion, 6BN is an attractive compound for development of a preventive therapy for NOWS., Competing Interests: WS is Chief Scientific Officer of Aether Therapeutics and holds share in Aether. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Safa, Lau, Aten, Schilling, Bales, Miller, Fitzgerald, Chen, Hill, Dzwigalski, Obrietan, Phelps, Sadee and Oberdick.)

Circadian rhythms influence various aspects of biology, including hormonal, immunological, and behavioral processes. These 24-hour oscillations are necessary to optimize cellular functions and to synchronize these processes with the environment. Breast cancer patients and survivors frequently report disruptions in circadian oscillations that adversely affect quality-of-life, including fragmented sleep-wake cycles and flattened cortisol rhythms, which are associated with negative behavioral comorbidities (e.g., fatigue). However, the potential causal role of tumor biology in circadian dysregulation has not been investigated. Here, we examined the extent to which sham surgery, non-metastatic mammary tumors, or mammary tumor removal in mice disrupts circadian rhythms in brain clock gene expression, locomotor behavior (free-running and entrained), and physiological rhythms that have been associated with cancer behavioral comorbidities. Tumors and tumor resection altered time-of-day differences in hypothalamic expression of eight circadian-regulated genes. The onset of activity in entrained running behavior was advanced in tumor-bearing mice, and the amplitude of free-running rhythms was increased in tumor-resected mice. Tumors flattened rhythms in circulating corticosterone and Ly6c Hi monocytes which were largely restored by surgical tumor resection. This work implies that tumors alone may directly impact central and/or peripheral circadian rhythmicity in breast cancer patients, and that these effects may persist in cancer survivors, potentially contributing to behavioral comorbidities., (Copyright © 2019 Elsevier Inc. All rights reserved.)

The microRNA miR-132 serves as a key regulator of a wide range of plasticity-associated processes in the central nervous system. Interestingly, miR-132 expression has also been shown to be under the control of the circadian timing system. This finding, coupled with work showing that miR-132 is expressed in the hippocampus, where it influences neuronal morphology and memory, led us to test the idea that daily rhythms in miR-132 within the forebrain modulate cognition as a function of circadian time. Here, we show that hippocampal miR-132 expression is gated by the time-of-day, with peak levels occurring during the circadian night. Further, in miR-132 knockout mice and in transgenic mice, where miR-132 is constitutively expressed under the control of the tetracycline regulator system, we found that time-of-day dependent memory recall (as assessed via novel object location and contextual fear conditioning paradigms) was suppressed. Given that miRNAs exert their functional effects via the suppression of target gene expression, we examined the effects that transgenic miR-132 manipulations have on MeCP2 and Sirt1-two miR-132 targets that are associated with neuronal plasticity and cognition. In mice where miR-132 was either knocked out, or transgenically expressed, rhythmic expression of MeCP2 and Sirt1 was suppressed. Taken together, these results raise the prospect that miR-132 serves as a key route through which the circadian timing system imparts a daily rhythm on cognitive capacity., (© 2018 Aten et al.; Published by Cold Spring Harbor Laboratory Press.)

Daily rhythms in behavior and physiology are coordinated by an endogenous clock located in the suprachiasmatic nucleus (SCN) of the hypothalamus. This central pacemaker also relays day length information to allow for seasonal adaptation, a process for which melatonin signaling is essential. How the SCN encodes day length is not fully understood. MicroRNAs (miRNAs) are small, non-coding RNAs that regulate gene expression by directing target mRNAs for degradation or translational repression. The miR-132/212 cluster plays a key role in facilitating neuronal plasticity, and miR-132 has been shown previously to modulate resetting of the central clock. A recent study from our group showed that miR-132/212 in mice is required for optimal adaptation to seasons and non-24-hour light/dark cycles through regulation of its target gene, methyl CpG-binding protein (MeCP2), in the SCN and dendritic spine density of SCN neurons. Furthermore, in the seasonal rodent Mesocricetus auratus (Syrian hamster), adaptation to short photoperiods is accompanied by structural plasticity in the SCN independently of melatonin signaling, thus further supporting a key role for SCN structural and, in turn, functional plasticity in the coding of day length. In this commentary, we discuss our recent findings in context of what is known about day length encoding by the SCN, and propose future directions.

The microRNA (miRNA) class of small (typically 22-24 nt) non-coding RNA affects a wide range of physiological processes in the mammalian central nervous system (CNS). By acting as potent regulators of mRNA translation and stability, miRNAs fine-tune the expression of a multitude of genes that play critical roles in complex cognitive processes, including learning and memory. Of note, within the CNS, miRNAs can be expressed in an inducible, and cell-type specific manner. Here, we provide a brief overview of the expression and functional effects of the miR-132/212 gene locus in forebrain circuits of the CNS, and then discuss a recent publication that explored the contributions of miR-132 and miR-212 to cognition and to transcriptome regulation. We also discuss mechanisms by which synaptic activity regulates miR-132/212 expression, how miR-132 and miR-212 affect neuronal plasticity, and how the dysregulation of these two miRNAs could contribute to the development of cognitive impairments., Competing Interests: Conflicting interests The authors have declared that no conflict of interests exist.

Ischemia-induced progenitor cell proliferation is a prominent example of the adult mammalian brain's ability to regenerate injured tissue resulting from pathophysiological processes. In order to better understand and exploit the cell signaling mechanisms that regulate ischemia-induced proliferation, we examined the role of the p42/44 mitogen-activated protein kinase (MAPK) cascade effector ribosomal S6 kinase (RSK) in this process. Here, using the endothelin-1 ischemia model in wild type mice, we show that the activated form of RSK is expressed in the progenitor cells of the subgranular zone (SGZ) after intrahippocampal cerebral ischemia. Further, RSK inhibition significantly reduces ischemia-induced SGZ progenitor cell proliferation. Using the neurosphere assay, we also show that both SGZ- and subventricular zone (SVZ)-derived adult neural stem cells (NSC) exhibit a significant reduction in proliferation in the presence of RSK and MAPK inhibitors. Taken together, these data reveal RSK as a regulator of ischemia-induced progenitor cell proliferation, and as such, suggest potential therapeutic value may be gained by specifically targeting the regulation of RSK in the progenitor cell population of the SGZ., (Copyright © 2013 Elsevier Inc. All rights reserved.)