RESUMO
Research in preclinical models indicates that estrogens are neuroprotective and positively impact cognitive aging. However, clinical data are equivocal as to the benefits of menopausal estrogen therapy to the brain and cognition. Pre-existing cardiometabolic disease may modulate mechanisms by which estrogens act, potentially reducing or reversing protections they provide against cognitive decline. In the current review we propose mechanisms by which cardiometabolic disease may alter estrogen effects, including both alterations in actions directly on brain memory systems and actions on cardiometabolic systems, which in turn impact brain memory systems. Consideration of mechanisms by which estrogen administration can exert differential effects dependent upon health phenotype is consistent with the move towards precision or personalized medicine, which aims to determine which treatment interventions will work for which individuals. Understanding effects of estrogens in both healthy and unhealthy models of aging is critical to optimizing the translational link between preclinical and clinical research.
Assuntos
Doenças Cardiovasculares , Estrogênios , Humanos , Encéfalo , Menopausa/psicologia , Cognição , Doenças Cardiovasculares/tratamento farmacológicoRESUMO
Thalamocortical neurons (TCNs) play a critical role in the maintenance of thalamocortical oscillations, dysregulation of which can result in certain types of seizures. Precise control over firing rates of TCNs is foundational to these oscillations, yet the transcriptional mechanisms that constrain these firing rates remain elusive. We hypothesized that Shox2 is a transcriptional regulator of ion channels important for TCN function and that loss of Shox2 alters firing frequency and activity, ultimately perturbing thalamocortical oscillations into an epilepsy-prone state. In this study, we used RNA sequencing and quantitative PCR of control and Shox2 knockout mice to determine Shox2-affected genes and revealed a network of ion channel genes important for neuronal firing properties. Protein regulation was confirmed by Western blotting, and electrophysiological recordings showed that Shox2 KO impacted the firing properties of a subpopulation of TCNs. Computational modeling showed that disruption of these conductances in a manner similar to Shox2's effects modulated frequency of oscillations and could convert sleep spindles to near spike and wave activity, which are a hallmark for absence epilepsy. Finally, Shox2 KO mice were more susceptible to pilocarpine-induced seizures. Overall, these results reveal Shox2 as a transcription factor important for TCN function in adult mouse thalamus.
Assuntos
Potenciais de Ação/fisiologia , Córtex Cerebral/metabolismo , Proteínas de Homeodomínio/biossíntese , Neurônios/metabolismo , Convulsões/metabolismo , Tálamo/metabolismo , Animais , Proteínas de Homeodomínio/genética , Canais Iônicos/biossíntese , Canais Iônicos/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Rede Nervosa/metabolismo , Convulsões/genética , Convulsões/prevenção & controle , Fatores de Transcrição/biossíntese , Fatores de Transcrição/genéticaRESUMO
Post-traumatic stress disorder (PTSD) is characterized by the development of paradoxical memory disturbances including intrusive memories and amnesia for specific details of the traumatic experience. Despite evidence that women are at higher risk to develop PTSD, most animal research has focused on the processes by which male rodents develop adaptive fear memory. As such, the mechanisms contributing to sex differences in the development of PTSD-like memory disturbances are poorly understood. In this investigation, we exposed adult male and female Wistar rats to the synthetic alarm odor 2,4,5-trimethylthiazole (TMT) to assess development of generalized fear behavior and rapid modulation of glutamate uptake and signaling cascades associated with hippocampus-dependent long-term memory. We report that female Wistar rats exposed to alarm odor exhibit context discrimination impairments relative to TMT-exposed male rats, suggesting the intriguing possibility that females are at greater risk in developing generalized fear memories. Mechanistically, alarm odor exposure rapidly modulated signaling cascades consistent with activation of the CREB shut-off cascade in the male, but not the female hippocampus. Moreover, TMT exposure dampened glutamate uptake and affected expression of the glutamate transporter, GLT-1 in the hippocampus. Taken together, these results provide evidence for rapid sex-dependent modulation of CREB signaling in the hippocampus by alarm odor exposure which may contribute to the development of generalized fear.
Assuntos
Medo/fisiologia , Ácido Glutâmico/metabolismo , Hipocampo/metabolismo , Memória/fisiologia , Odorantes , Animais , Feminino , Masculino , Ratos , Ratos Wistar , Fatores Sexuais , Transtornos de Estresse Pós-Traumáticos/metabolismoRESUMO
Post-traumatic stress disorder (PTSD) is characterized by memory disturbances following trauma. Acute predator threat has emerged as an ethological model of PTSD, yet the effects of predator odor on signaling cascades associated with long-term memory remain poorly understood. In this study, we exposed male and female Wistar rats to the synthetic predator odor 2,5-dihydro-2,4,5-trimethylthiazoline (TMT) to assess behavioral and physiological responses as well as rapid modulation of signal transduction cascades associated with learning and memory in the male and female hippocampus. During exposure to TMT in the homecage, both male and female animals displayed robust immobility, avoidance, and altered activity as a function of time. Physiologically, TMT exposure increased circulating corticosterone and blood glucose in both male and female rodents, suggesting that TMT evokes sex-independent behavioral and physiological responses. With respect to signal transduction, TMT exposure rapidly reduced phosphorylation of cyclic-adenosine monophosphate response element binding protein (CREB) in the male, but not the female hippocampus. Furthermore, TMT exposure reduced phosphorylation of extracellular signal-regulated kinase 1/2 and increased nuclear expression of the synapto-nuclear messenger protein Jacob in the male hippocampus, consistent with activation of the CREB shut-off pathway. In a follow-up behavioral experiment, post-training exposure to TMT did not affect spatial water maze performance of male rats. However, male rats re-introduced to the context in which TMT had previously been presented displayed avoidance and hyperactivity, but not freezing behavior or elevated corticosterone responses, suggesting that TMT exposure supports a form of contextual conditioning which is not characterized by immobility. Taken together, our findings suggest that TMT evokes similar behavioral and physiological responses in male and female Wistar rats, but affects distinct signaling cascades in the male and female hippocampus which may contribute to behavioral disruptions associated with predator exposure.
Assuntos
Proteína de Ligação a CREB/metabolismo , Medo/psicologia , Hipocampo/metabolismo , Odorantes , Transtornos de Estresse Pós-Traumáticos/metabolismo , Animais , Glicemia/efeitos dos fármacos , Corticosterona/sangue , AMP Cíclico/metabolismo , Modelos Animais de Doenças , Feminino , Resposta de Imobilidade Tônica/efeitos dos fármacos , Resposta de Imobilidade Tônica/fisiologia , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Sistema de Sinalização das MAP Quinases/fisiologia , Masculino , Aprendizagem em Labirinto/efeitos dos fármacos , Aprendizagem em Labirinto/fisiologia , Fosforilação/fisiologia , Ratos , Ratos Wistar , Fatores Sexuais , Transtornos de Estresse Pós-Traumáticos/induzido quimicamente , Tiazóis/administração & dosagemRESUMO
KEY POINTS: Synaptic excitation and inhibition must be properly balanced in individual neurons and neuronal networks to allow proper brain function. Disrupting this balance may lead to autism spectral disorders and epilepsy. We show the basic helix-loop-helix transcription factor NeuroD2 promotes inhibitory synaptic drive but also decreases cell-intrinsic neuronal excitability of cortical pyramidal neurons both in vitro and in vivo. We identify two genes potentially downstream of NeuroD2-mediated transcription that regulate these parameters: gastrin-releasing peptide and the small conductance, calcium-activated potassium channel, SK2. Our results reveal an important function for NeuroD2 in balancing synaptic neurotransmission and intrinsic excitability. Our results offer insight into how synaptic innervation and intrinsic excitability are coordinated during cortical development. ABSTRACT: Synaptic excitation and inhibition must be properly balanced in individual neurons and neuronal networks for proper brain function. Disruption of this balance during development may lead to autism spectral disorders and epilepsy. Synaptic excitation is counterbalanced by synaptic inhibition but also by attenuation of cell-intrinsic neuronal excitability. To maintain proper excitation levels during development, neurons must sense activity over time and regulate the expression of genes that control these parameters. While this is a critical process, little is known about the transcription factors involved in coordinating gene expression to control excitatory/inhibitory synaptic balance. We show here that the basic helix-loop-helix transcription factor NeuroD2 promotes inhibitory synaptic drive but also decreases cell-intrinsic neuronal excitability of cortical pyramidal neurons both in vitro and in vivo as shown by ex vivo analysis of a NeuroD2 knockout mouse. Using microarray analysis and comparing wild-type and NeuroD2 knockout cortical networks, we identified two potential gene targets of NeuroD2 that contribute to these processes: gastrin-releasing peptide (GRP) and the small conductance, calcium-activated potassium channel, SK2. We found that the GRP receptor antagonist RC-3059 and the SK2 specific blocker apamin partially reversed the effects of increased NeuroD2 expression on inhibitory synaptic drive and action potential repolarization, respectively. Our results reveal an important function for NeuroD2 in balancing synaptic neurotransmission and intrinsic excitability and offer insight into how these processes are coordinated during cortical development.
Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/fisiologia , Neuropeptídeos/fisiologia , Células Piramidais/fisiologia , Córtex Somatossensorial/fisiologia , Sinapses/fisiologia , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Células Cultivadas , Peptídeo Liberador de Gastrina/genética , Potenciais Pós-Sinápticos Inibidores , Camundongos Knockout , Neuropeptídeos/genética , Ratos , Canais de Potássio Ativados por Cálcio de Condutância Baixa/genéticaRESUMO
Females experience depression, posttraumatic stress disorder (PTSD), and anxiety disorders at approximately twice the rate of males, but the mechanisms underlying this difference remain undefined. The effect of sex hormones on neural substrates presents a possible mechanism. We investigated the effect of ovariectomy at two ages, before puberty and in adulthood, and 17ß-estradiol (E2) replacement administered chronically in drinking water on anxiety level, fear memory formation, and extinction. Based on previous studies, we hypothesized that estradiol replacement would impair fear memory formation and enhance extinction rate. Females, age 4 weeks and 10 weeks, were divided randomly into 4 groups; sham surgery, OVX, OVX+low E2 (200nM), and OVX+high E2 (1000nM). Chronic treatment with high levels of E2 significantly increased anxiety levels measured in the elevated plus maze. In both age groups, high levels of E2 significantly increased contextual fear memory but had no effect on cued fear memory. In addition, high E2 decreased the rate of extinction in both ages. Finally, catechol-O-methyltransferase (COMT) is important for regulation of catecholamine levels, which play a role in fear memory formation and extinction. COMT expression in the hippocampus was significantly reduced by high E2 replacement, implying increased catecholamine levels in the hippocampus of high E2 mice. These results suggest that estradiol enhanced fear memory formation, and inhibited fear memory extinction, possibly stabilizing the fear memory in female mice. This study has implications for a neurobiological mechanism for PTSD and anxiety disorders.
Assuntos
Ansiedade/fisiopatologia , Catecol O-Metiltransferase/metabolismo , Estradiol/fisiologia , Extinção Psicológica/fisiologia , Medo/fisiologia , Memória/fisiologia , Animais , Estradiol/administração & dosagem , Extinção Psicológica/efeitos dos fármacos , Medo/efeitos dos fármacos , Feminino , Expressão Gênica , Hipocampo/efeitos dos fármacos , Hipocampo/metabolismo , Memória/efeitos dos fármacos , Camundongos , Ovariectomia , Córtex Pré-Frontal/efeitos dos fármacos , Córtex Pré-Frontal/metabolismoRESUMO
The dentate gyrus of the hippocampus is thought to control information flow into the rest of the hippocampus. Under pathological conditions, such as epilepsy, this protective feature is circumvented and uninhibited activity flows throughout the hippocampus. Many factors can modulate excitability of the dentate gyrus and ultimately, the hippocampus. It is therefore of critical importance to understand the mechanisms involved in regulating excitability in the dentate gyrus. Dynorphin, the endogenous ligand for the kappa (κ) opioid receptor (KOR), is thought to be involved in neuromodulation in the dentate gyrus. Both dynorphin and its receptor are widely expressed in the dentate gyrus and have been implicated in epilepsy and other complex behaviours such as stress-induced deficits in learning and stress-induced depression-like behaviours. Administration of KOR agonists can prevent both the behavioural and electroencephalographic measures of seizures in several different models of epilepsy. Antagonism of the KORs also prevents stress-induced behaviours. This evidence suggests the KORs as possible therapeutic targets for various pathological conditions. In addition, KOR agonists prevent the induction of LTP. Although there are several mechanisms through which dynorphin could mediate these effects, no studies to date investigated the effects of KOR activation on intrinsic membrane properties and cell excitability. We used whole-cell, patch-clamp recordings from acute mouse hippocampus slices to investigate the effect of KOR activation on dentate gyrus granule cell excitability. The agonist U69,593 (U6, 1 µM) resulted in a lower spike threshold, a decreased latency to first spike, an increased spike half-width, and an overall increase in spike number with current injections ranging from 15 to 45 pA. There was also a reduction in the interspike interval (ISI) both early and late in the spike train, with no change in membrane potential or input resistance. Preincubation of the slice with the selective KOR antagonist, nor-binalthorphimine (BNI, 1 µM) inhibited the effect of U6 on the latency to first spike and spike half-width suggesting that these effects are mediated through KORs. The inclusion of GDP-ßS (1 mM) in the recording pipette prevented all of the U6 effects, suggesting that all effects are mediated via a G-protein-dependent mechanism. Inclusion of the A-type K+ current blocker, 4-aminopyridine (4-AP, 5 mM) in the pipette also antagonised the effects of U6. Kv4.2 is one of the channel α subunits thought to be responsible for carrying the A-type K+ current. Incubation of hippocampus slices with U6 resulted in a decrease in the Kv4.2 subunit protein at the cell surface. These results are consistent with an increase in cell excitability in response to KOR activation and may reflect new possibilities for additional opioid functions.
Assuntos
Giro Denteado/efeitos dos fármacos , Neurônios/efeitos dos fármacos , Receptores Opioides kappa/metabolismo , 4-Aminopiridina/farmacologia , Potenciais de Ação/efeitos dos fármacos , Animais , Benzenoacetamidas/farmacologia , Giro Denteado/metabolismo , Dinorfinas/metabolismo , Proteínas de Ligação ao GTP/metabolismo , Guanosina Difosfato/análogos & derivados , Guanosina Difosfato/farmacologia , Masculino , Camundongos , Camundongos da Linhagem 129 , Naltrexona/análogos & derivados , Naltrexona/farmacologia , Neurônios/metabolismo , Técnicas de Patch-Clamp/métodos , Canais de Potássio/metabolismo , Pirrolidinas/farmacologia , Receptores Opioides kappa/agonistas , Receptores Opioides kappa/antagonistas & inibidores , Tionucleotídeos/farmacologiaRESUMO
The multiple memory systems hypothesis proposes that different types of learning strategies are mediated by distinct neural systems in the brain. Male and female mice were tested on a water plus-maze task that could be solved by either a place or response strategy. One group of mice was pre-exposed to the same context as training and testing (PTC) and the other group was pre-exposed to a different context (PDC). Our results show that the PTC condition biased mice to place strategy use in males, but this bias was dependent on the presence of ovarian hormones in females.
Assuntos
Sinais (Psicologia) , Aprendizagem em Labirinto/fisiologia , Memória/fisiologia , Percepção Espacial/fisiologia , Análise de Variância , Animais , Comportamento Animal , Estradiol/farmacologia , Feminino , Masculino , Aprendizagem em Labirinto/efeitos dos fármacos , Memória/efeitos dos fármacos , Camundongos , Ovariectomia/métodos , Tempo de Reação/efeitos dos fármacos , Tempo de Reação/fisiologiaRESUMO
Transient outward K+ currents are particularly important for the regulation of membrane excitability of neurons and repolarization of action potentials in cardiac myocytes. These currents are modulated by PKC (protein kinase C) activation, and the K+- channel subunit Kv4.2 is a major contributor to these currents. Furthermore, the current recorded from Kv4.2 channels expressed in oocytes is reduced by PKC activation. The mechanism underlying PKC regulation of Kv4.2 currents is unknown. In the present study, we determined that PKC directly phosphorylates the Kv4.2 channel protein. In vitro phosphorylation of the intracellular N- and C-termini of Kv4.2 GST (glutathione transferase) tagged fusion protein revealed that the C-terminal of Kv4.2 was phosphorylated by PKC, whereas the N-terminal was not. Amino acid mapping and site-directed mutagenesis revealed that the phosphorylated residues on the Kv4.2 C-terminal were Ser447 and Ser537. A phospho-site-specific antibody showed that phosphorylation at the Ser537 site was increased in the hippocampus in response to PKC activation. Surface biotinylation experiments revealed that mutation to alanine of both Ser447 and Ser537 in order to block phosphorylation at both of the PKC sites increased surface expression compared with wild-type Kv4.2. Electrophysiological recordings of the wild-type and both the alanine and aspartate mutant Kv4.2 channels expressed with KChIP3 (Kv4 channel-interacting protein 3) revealed no significant difference in the half-activation or half-inactivation voltage of the channel. Interestingly, Ser537 lies within a possible ERK (extracellular-signal-regulated kinase)/MAPK (mitogen-activated protein kinase) recognition (docking) domain in the Kv4.2 C-terminal sequence. We found that phosphorylation of Kv4.2 by PKC enhanced ERK phosphorylation of the channel in vitro. These findings suggest the possibility that Kv4.2 is a locus for PKC and ERK cross-talk.
Assuntos
MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Proteína Quinase C/metabolismo , Canais de Potássio Shal/metabolismo , Transdução de Sinais , Sequência de Aminoácidos , Animais , Sítios de Ligação , Células COS , Chlorocebus aethiops , Citoplasma/metabolismo , DNA Complementar/genética , DNA Complementar/metabolismo , Humanos , Proteínas Interatuantes com Canais de Kv/metabolismo , Quinases de Proteína Quinase Ativadas por Mitógeno/metabolismo , Modelos Biológicos , Dados de Sequência Molecular , Oócitos/metabolismo , Fosforilação , Ratos , Proteínas Repressoras/metabolismo , XenopusRESUMO
Potassium channel interacting proteins (KChIPs) are members of a family of calcium binding proteins that interact with Kv4 potassium (K(+)) channel primary subunits and also act as transcription factors. The Kv4 subunit is a primary K(+) channel pore-forming subunit, which contributes to the somatic and dendritic A-type currents throughout the nervous system. These A-type currents play a key role in the regulation of neuronal excitability and dendritic processing of incoming synaptic information. KChIP3 is also known as calsenilin and as the transcription factor, downstream regulatory element antagonist modulator (DREAM), which regulates a number of genes including prodynorphin. KChIP3 and Kv4 primary channel subunits are highly expressed in hippocampus, an area of the brain important for learning and memory. Through its various functions, KChIP3 may play a role in the regulation of synaptic plasticity and learning and memory. We evaluated the role of KChIP3 in a hippocampus-dependent memory task, contextual fear conditioning. Male KChIP3 knockout (KO) mice showed significantly enhanced memory 24 hours after training as measured by percent freezing. In addition, we found that membrane association and interaction with Kv4.2 of KChIP3 protein was significantly decreased and nuclear KChIP3 expression was increased six hours after the fear conditioning training paradigm with no significant change in KChIP3 mRNA. In addition, prodynorphin mRNA expression was significantly decreased six hours after fear conditioning training in wild-type (WT) but not in KO animals. These data suggest a role for regulation of gene expression by KChIP3/DREAM/calsenilin in consolidation of contextual fear conditioning memories.
Assuntos
Condicionamento Clássico/fisiologia , Medo , Regulação da Expressão Gênica/fisiologia , Proteínas Interatuantes com Canais de Kv/fisiologia , Proteínas Repressoras/fisiologia , Análise de Variância , Animais , Comportamento Animal , Nucléolo Celular/metabolismo , Sinais (Psicologia) , Encefalinas/genética , Comportamento Exploratório/fisiologia , Reação de Congelamento Cataléptica/fisiologia , Regulação da Expressão Gênica/genética , Hipocampo/metabolismo , Imunoprecipitação/métodos , Proteínas Interatuantes com Canais de Kv/deficiência , Masculino , Aprendizagem em Labirinto/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Atividade Motora/genética , Precursores de Proteínas/genética , RNA Mensageiro/metabolismo , Teste de Desempenho do Rota-Rod , Limiar Sensorial/fisiologia , Canais de Potássio Shal/metabolismo , Fatores de TempoRESUMO
Spatial memory processing requires functional interaction between the hippocampus and the medial entorhinal cortex (MEC). The grid cells of the MEC are most abundant in layer II and rely on a complex network of local inhibitory interneurons to generate spatial firing properties. Stress can cause spatial memory deficits in males, but the specific underlying mechanisms affecting the known memory pathways remain unclear. Stress activates both the autonomic nervous system and the hypothalamic-pituitary-adrenal axis to release norepinephrine (NE) and glucocorticoids, respectively. Given that adrenergic receptor (AR) and glucocorticoid receptor (GR) expression is abundant in the MEC, both glucocorticoids and NE released in response to stress may have rapid effects on MEC-LII networks. We used whole-cell patch clamp electrophysiology in MEC slice preparations from male mice to test the effects of NE and glucocorticoids on inhibitory synaptic inputs of MEC-LII principal cells. Application of NE (100 µM) increased the frequency and amplitude of spontaneous inhibitory post-synaptic currents (sIPSCs) in approximately 75% of the principal cells tested. Unlike NE, bath application of dexamethasone (Dex, 1 µM), a synthetic glucocorticoid, or corticosterone (1 µM) the glucocorticoid in rodents, rapidly decreased the frequency of sIPSCs, but not miniature (mIPSCs) in MEC-LII principal cells. Interestingly, pre-treatment with Dex prior to NE application led to an NE-induced increase in sIPSC frequency in all cells tested. This effect was mediated by the α1-AR, as application of an α1-AR agonist, phenylephrine (PHE) yielded the same results, suggesting that a subset of cells in MEC-LII are unresponsive to α1-AR activation without prior activation of GR. We conclude that activation of GRs primes a subset of principal cells that were previously insensitive to NE to become responsive to α1-AR activation in a transcription-independent manner. These findings demonstrate the ability of stress hormones to markedly alter inhibitory signaling within MEC-LII circuits and suggest the intriguing possibility of modulation of network processing upstream of the hippocampus.
RESUMO
Previous genomic studies in humans indicate that SIRT1, a nicotinamide adenine dinucleotide (NAD+)-dependent protein deacetylase, is involved in anxiety and depression, but the mechanisms are unclear. We previously showed that SIRT1 is highly activated in the nuclear fraction of the dentate gyrus of the chronically stressed animals and inhibits memory formation and increases anhedonic behavior during chronic stress, but specific functional targets of cytoplasmic SIRT1 are unknown. Here, we demonstrate that SIRT1 activity rapidly modulates intrinsic and synaptic properties of the dentate gyrus granule cells and anxiety behaviors through deacetylation of BK channel α subunits in control animals. Chronic stress decreases BKα channel membrane expression, and SIRT1 activity has no rapid effects on synaptic transmission or intrinsic properties in the chronically stressed animal. These results suggest SIRT1 activity rapidly modulates the physiological function of the dentate gyrus, and this modulation participates in the maladaptive stress response.
RESUMO
A-type channels, encoded by the pore-forming alpha-subunits of the Kv4.x family, are particularly important in regulating membrane excitability in the CNS and the heart. Given the key role of modulation of A currents by kinases, we sought to investigate the protein structure-function relationships underlying the regulation of these currents by PKA. We have previously shown the existence of two PKA phosphorylation sites in the Kv4.2 sequence; therefore, we focused this study on the Kv4.2 primary subunit. In the present studies we made the surprising finding that PKA phosphorylation of the Kv4.2 alpha-subunit is necessary but not sufficient for channel modulation; channel modulation by PKA required the presence of an ancillary subunit, the K+ channel interacting protein (KChIP3). Therefore, these findings indicate a surprising complexity to kinase regulation of A currents, in that an interaction of two separate molecular events, alpha-subunit phosphorylation and the association of an ancillary subunit (KChIP3), are necessary for phosphorylation-dependent regulation of Kv4.2-encoded A channels by PKA. Overall, our studies indicate that PKA must of necessity act on a supramolecular complex of pore-forming alpha-subunits plus ancillary subunits to alter channel properties.
Assuntos
Colforsina/análogos & derivados , Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , Canais de Potássio de Abertura Dependente da Tensão da Membrana , Canais de Potássio/metabolismo , Proteínas Repressoras , 8-Bromo Monofosfato de Adenosina Cíclica/farmacologia , Substituição de Aminoácidos , Animais , Sítios de Ligação/genética , Células COS , Proteínas de Ligação ao Cálcio/genética , Proteínas de Ligação ao Cálcio/metabolismo , Colforsina/farmacologia , Proteínas Quinases Dependentes de AMP Cíclico/antagonistas & inibidores , Ativação Enzimática/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Proteínas Interatuantes com Canais de Kv , Substâncias Macromoleculares , Mutagênese Sítio-Dirigida , Oócitos/efeitos dos fármacos , Oócitos/metabolismo , Técnicas de Patch-Clamp , Fosfopeptídeos/metabolismo , Fosforilação , Canais de Potássio/genética , Subunidades Proteicas/metabolismo , Canais de Potássio Shal , Relação Estrutura-Atividade , Transfecção , Xenopus laevisRESUMO
Calcium-calmodulin-dependent kinase II (CaMKII) has a long history of involvement in synaptic plasticity, yet little focus has been given to potassium channels as CaMKII targets despite their importance in repolarizing EPSPs and action potentials and regulating neuronal membrane excitability. We now show that Kv4.2 acts as a substrate for CaMKII in vitro and have identified CaMKII phosphorylation sites as Ser438 and Ser459. To test whether CaMKII phosphorylation of Kv4.2 affects channel biophysics, we expressed wild-type or mutant Kv4.2 and the K(+) channel interacting protein, KChIP3, with or without a constitutively active form of CaMKII in Xenopus oocytes and measured the voltage dependence of activation and inactivation in each of these conditions. CaMKII phosphorylation had no effect on channel biophysical properties. However, we found that levels of Kv4.2 protein are increased with CaMKII phosphorylation in transfected COS cells, an effect attributable to direct channel phosphorylation based on site-directed mutagenesis studies. We also obtained corroborating physiological data showing increased surface A-type channel expression as revealed by increases in peak K(+) current amplitudes with CaMKII phosphorylation. Furthermore, endogenous A-currents in hippocampal pyramidal neurons were increased in amplitude after introduction of constitutively active CaMKII, which results in a decrease in neuronal excitability in response to current injections. Thus CaMKII can directly modulate neuronal excitability by increasing cell-surface expression of A-type K(+) channels.
Assuntos
Proteínas Quinases Dependentes de Cálcio-Calmodulina/metabolismo , Regulação da Expressão Gênica/fisiologia , Neurônios/metabolismo , Canais de Potássio de Abertura Dependente da Tensão da Membrana , Canais de Potássio/metabolismo , Potássio/metabolismo , Animais , Proteínas de Ligação ao Cálcio/genética , Proteínas de Ligação ao Cálcio/metabolismo , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina , Proteínas Quinases Dependentes de Cálcio-Calmodulina/genética , Células Cultivadas , Chlorocebus aethiops , Cricetinae , Técnicas de Transferência de Genes , Hipocampo/citologia , Proteínas Interatuantes com Canais de Kv , Mutagênese Sítio-Dirigida , Neurônios/citologia , Oócitos/metabolismo , Técnicas de Patch-Clamp , Fosforilação , Canais de Potássio/genética , Células Piramidais/citologia , Células Piramidais/metabolismo , Ratos , Ratos Sprague-Dawley , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Canais de Potássio Shal , Regulação para Cima/fisiologia , XenopusRESUMO
Historically, much attention has focused on the mechanisms of activity-dependent plasticity since the description of long-term potentiation by Bliss and Lomo in the early 1970s, while extrasynaptic changes have received much less interest. However, recent work has concentrated on the role of back-propagating action potentials in hippocampal dendrites in synaptic plasticity. In this review, we focus on the modulation of back-propagating action potentials by K+ currents in the dendrites of hippocampal cells. We described the primary K+-channel subunits and their interacting subunits that most likely contribute to these currents, and how these sites can be regulated by phosphorylation and other mechanisms. In conclusion, we provide a model for an alternative form of coincidence detection through K+ channels in the hippocampus.
Assuntos
Hipocampo/fisiologia , Proteínas do Tecido Nervoso/fisiologia , Plasticidade Neuronal/fisiologia , Canais de Potássio de Abertura Dependente da Tensão da Membrana , Canais de Potássio/fisiologia , Potássio/fisiologia , Potenciais de Ação , Animais , Proteínas do Citoesqueleto/fisiologia , Dendritos/fisiologia , Hipocampo/citologia , Humanos , Transporte de Íons , Sistema de Sinalização das MAP Quinases , Camundongos , Modelos Moleculares , Modelos Neurológicos , Proteínas do Tecido Nervoso/química , Fosforilação , Canais de Potássio/química , Canais de Potássio/classificação , Proteínas Quinases/fisiologia , Processamento de Proteína Pós-Traducional , Subunidades Proteicas , Canais de Potássio Shal , Sinapses/fisiologiaRESUMO
Evidence suggests that when presented with novel acute stress, animals previously exposed to chronic homotypic or heterotypic stressors exhibit normal or enhanced hypothalamic-pituitary-adrenal (HPA) response compared with animals exposed solely to that acute stressor. The molecular mechanisms involved in this effect remain unknown. The extracellular signal-regulated kinase (ERK) is one of the key pathways regulated in the hippocampus in both acute and chronic stress. The aim of this study was to examine the interaction of prior chronic stress, using the chronic variable stress model (CVS), with exposure to a novel acute stressor (2,5-dihydro-2,4,5-trimethyl thiazoline; TMT) on ERK activation, expression of the downstream protein BCL-2, and the glucocorticoid receptor co-chaperone BAG-1 in control and chronically stressed male rats. TMT exposure after chronic stress resulted in a significant interaction of chronic and acute stress in all 3 hippocampus subregions on ERK activation and BCL-2 expression. Significantly, acute stress increased ERK activation, BCL-2 and BAG-1 protein expression in the dentate gyrus (DG) of CVS-treated rats compared with control, CVS-treated alone, and TMT-only animals. Furthermore, CVS significantly increased ERK activation in medial prefrontal cortex, but acute stress had no significant effect. Inhibition of corticosterone synthesis with metyrapone had no significant effect on ERK activation in the hippocampus; therefore, glucocorticoids alone do not mediate the molecular effects. Finally, because post-translational modifications of histones are believed to play an important role in the stress response, we examined changes in histone acetylation. We found that, in general, chronic stress decreased K12H4 acetylation, whereas acute stress increased acetylation. These results indicate a molecular mechanism by which chronic stress-induced HPA axis plasticity can lead to neurochemical alterations in the hippocampus that influence reactivity to subsequent stress exposure. This may represent an important site of dysfunction that contributes to stress-induced pathology such as depression, anxiety disorders, and posttraumatic stress disorder.
Assuntos
Giro Denteado/metabolismo , Sistema Hipotálamo-Hipofisário/metabolismo , Sistema de Sinalização das MAP Quinases , Sistema Hipófise-Suprarrenal/metabolismo , Estresse Psicológico/metabolismo , Acetilação , Acetiltransferases/metabolismo , Animais , Histonas/metabolismo , Masculino , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Distribuição Aleatória , Ratos , Ratos Wistar , Estresse Fisiológico , TiazóisRESUMO
Memory acquisition and synaptic plasticity are accompanied by changes in the intrinsic excitability of CA1 pyramidal neurons. These activity-dependent changes in excitability are mediated by modulation of intrinsic currents which alters the responsiveness of the cell to synaptic inputs. The afterhyperpolarization (AHP), a major contributor to the regulation of neuronal excitability, is reduced in animals that have acquired several types of hippocampus-dependent memory tasks and also following synaptic potentiation by high frequency stimulation. BK channels underlie the fast AHP and contribute to spike repolarization, and this AHP is reduced in animals that successfully acquired trace-eyeblink conditioning. This suggests that BK channel function is activity-dependent, but the mechanisms are unknown. In this study, we found that blockade of BK channels with paxilline (10 µM) decreased I AHP amplitude and increased spike half-width and instantaneous frequency in response to a +100 pA depolarization. In addition, induction of long term potentiation (LTP) by theta burst stimulation (TBS) in CA1 pyramidal neurons reduced BK channel's contribution to I AHP, spike repolarization, and instantaneous frequency. This result indicates that BK channel activity is decreased following synaptic potentiation. Interestingly, blockade of mammalian target of rapamycin (MTORC1) with rapamycin (400 nM) following synaptic potentiation restored BK channel function, suggesting a role for protein translation in signaling events which decreased postsynaptic BK channel activity following synaptic potentiation.
RESUMO
BACKGROUND: Exposure to chronic stress produces negative effects on mood and hippocampus-dependent memory formation. Alterations in signaling cascades and histone acetylation present a mechanism of modulation of transcription that may underlie stress-dependent processes in the hippocampus critical to learning and memory and development of depressive behaviors. METHODS: The rat model of chronic variable stress (CVS) was used to investigate the role of changes in protein acetylation and other molecular components of hippocampus-dependent memory formation and anhedonic behavior in response to CVS. RESULTS: Chronic variable stress treatment decreased both extracellular signal-regulated protein kinases 1 and 2 activation and Bcl-2 expression in all three regions of the hippocampus that corresponded behaviorally with a decrease in memory for the novel object location task and increased anhedonia. Extracellular signal-regulated protein kinases 1 and 2 activation was not significantly affected in the amygdala and increased in the medial prefrontal cortex by CVS. Chronic variable stress had no significant effect on activation of Akt in the hippocampus. We investigated molecular and behavioral effects of infusion of the sirtuin inhibitor, sirtinol, into the dentate gyrus (DG). Sirtinol infusion into the DG prevented the CVS-mediated decrease in extracellular signal-regulated protein kinases 1 and 2 activity and Bcl-2 expression, as well as histone acetylation in the DG previously observed following CVS. This corresponded to enhanced performance on the novel object location memory task, as well as reduced anhedonic behavior. CONCLUSIONS: These results suggest that changes in sirtuin activity contribute to changes in molecular cascades and histone acetylation within the hippocampus observed following CVS and may represent a novel therapeutic target for stress-induced depression.
Assuntos
Giro Denteado/metabolismo , Hipocampo/metabolismo , Sirtuína 1/metabolismo , Estresse Psicológico/patologia , Animais , Imunoprecipitação da Cromatina , Doença Crônica , Corticosterona/sangue , Modelos Animais de Doenças , Preferências Alimentares/fisiologia , Masculino , Proteína Quinase 1 Ativada por Mitógeno/genética , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/genética , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , RNA Mensageiro/metabolismo , Ratos , Ratos Wistar , Transdução de Sinais/fisiologia , Estresse Psicológico/sangue , Sacarose/administração & dosagem , Edulcorantes/administração & dosagemRESUMO
Recent research investigating Pavlovian fear conditioning and fear extinction has elucidated the neurocircuitry involved in acquisition and inhibition of fear responses. Modulatory factors that may underlie individual differences in fear acquisition and inhibition, however, are not well understood. Testosterone is known to affect anxiety-like behavior and cognitive processing. In this study, we hypothesized that castration would increase anxiety and reduce memory for contextual fear conditioning in an age-dependent manner. In addition, castration would reduce the rate of extinction to context, as high levels of testosterone correlate with reduced PTSD-like symptoms. We compared behaviors in male mice that were castrated at one of two different time points, either before puberty (at 4 weeks) or after puberty (at 10 weeks) to sham-operated control mice. The behaviors investigated included: anxiety, cued and contextual fear conditioning, and extinction of the fear memory. An interaction of hormone status and age and a significant effect of age were measured in the elevated plus maze, a measure of anxiety. Castration caused a significant reduction of contextual fear memory, but no effect on cued fear memory. There was no significant effect of castration on extinction. Interestingly, a significant effect of age of the mouse at the time of testing was observed on extinction. These results suggest that endogenous androgens during puberty are important for anxiety and fear memory formation. In addition, these results define a late post-adolescent developmental time point for changes in anxiety and fear extinction.
Assuntos
Ansiedade/fisiopatologia , Condicionamento Clássico/fisiologia , Extinção Psicológica/fisiologia , Medo/fisiologia , Testosterona/fisiologia , Fatores Etários , Análise de Variância , Animais , Aprendizagem por Associação/fisiologia , Comportamento Animal/fisiologia , Castração , Inibição Psicológica , Masculino , Memória/fisiologia , Camundongos , Camundongos da Linhagem 129 , Fatores Sexuais , Maturidade Sexual/fisiologiaRESUMO
Corticosterone (CORT) release from the adrenal glands in response to acutely stressful stimuli is well-characterized, however several non-experimental, environmental stressors can also engender a CORT response. The aim of this study was to investigate an acute activation of the HPA axis in pair-housed animals in response to separation. We observed a rapid significant increase in CORT in the animal remaining in the home cage following cage mate removal, that was not caused by cage opening and transient removal of cage mate. In addition, we examined this response in both control, non-stressed animals and in animals subjected to chronic variable stress (CVS) and found that although basal levels of CORT differed between control and CVS animals, there was no significant difference in the acute CORT levels between the control and CVS animals after separation, indicating that this environmental event is perceived as acutely stressful in both conditions. Furthermore, we examined the time course of CORT activation and found that CORT levels rapidly rise within minutes of separation peaking at 15 min and returning to baseline by 90 min. The results of this study demonstrate that separation can induce an acute stress response in the remaining cage mate measured by increased CORT and should be considered in molecular, behavioral, and electrophysiological studies.