RESUMO
Cav1.2 channels play crucial roles in various neuronal and physiological processes. Here, we present cryo-EM structures of human Cav1.2, both in its apo form and in complex with several drugs, as well as the peptide neurotoxin calciseptine. Most structures, apo or bound to calciseptine, amlodipine, or a combination of amiodarone and sofosbuvir, exhibit a consistent inactivated conformation with a sealed gate, three up voltage-sensing domains (VSDs), and a down VSDII. Calciseptine sits on the shoulder of the pore domain, away from the permeation path. In contrast, when pinaverium bromide, an antispasmodic drug, is inserted into a cavity reminiscent of the IFM-binding site in Nav channels, a series of structural changes occur, including upward movement of VSDII coupled with dilation of the selectivity filter and its surrounding segments in repeat III. Meanwhile, S4-5III merges with S5III to become a single helix, resulting in a widened but still non-conductive intracellular gate.
Assuntos
Canais de Cálcio Tipo L , Venenos Elapídicos , Humanos , Canais de Cálcio Tipo L/química , Canais de Cálcio Tipo L/metabolismo , Neurotoxinas , Domínios Proteicos , Microscopia CrioeletrônicaRESUMO
Drug-drug interaction of the antiviral sofosbuvir and the antiarrhythmics amiodarone has been reported to cause fatal heartbeat slowing. Sofosbuvir and its analog, MNI-1, were reported to potentiate the inhibition of cardiomyocyte calcium handling by amiodarone, which functions as a multi-channel antagonist, and implicate its inhibitory effect on L-type Cav channels, but the molecular mechanism has remained unclear. Here we present systematic cryo-EM structural analysis of Cav1.1 and Cav1.3 treated with amiodarone or sofosbuvir alone, or sofosbuvir/MNI-1 combined with amiodarone. Whereas amiodarone alone occupies the dihydropyridine binding site, sofosbuvir is not found in the channel when applied on its own. In the presence of amiodarone, sofosbuvir/MNI-1 is anchored in the central cavity of the pore domain through specific interaction with amiodarone and directly obstructs the ion permeation path. Our study reveals the molecular basis for the physical, pharmacodynamic interaction of two drugs on the scaffold of Cav channels.
Assuntos
Amiodarona , Sofosbuvir , Sofosbuvir/efeitos adversos , Amiodarona/farmacologia , Antivirais/farmacologia , Miócitos Cardíacos/metabolismo , Sítios de Ligação , Canais de Cálcio Tipo L/metabolismo , Cálcio/metabolismoRESUMO
The mechanism that initiates autophagosome formation on the ER in multicellular organisms is elusive. Here, we showed that autophagy stimuli trigger Ca2+ transients on the outer surface of the ER membrane, whose amplitude, frequency, and duration are controlled by the metazoan-specific ER transmembrane autophagy protein EPG-4/EI24. Persistent Ca2+ transients/oscillations on the cytosolic ER surface in EI24-depleted cells cause accumulation of FIP200 autophagosome initiation complexes on the ER. This defect is suppressed by attenuating ER Ca2+ transients. Multi-modal SIM analysis revealed that Ca2+ transients on the ER trigger the formation of dynamic and fusion-prone liquid-like FIP200 puncta. Starvation-induced Ca2+ transients on lysosomes also induce FIP200 puncta that further move to the ER. Multiple FIP200 puncta on the ER, whose association depends on the ER proteins VAPA/B and ATL2/3, assemble into autophagosome formation sites. Thus, Ca2+ transients are crucial for triggering phase separation of FIP200 to specify autophagosome initiation sites in metazoans.
Assuntos
Autofagossomos , Cálcio , Animais , Autofagossomos/metabolismo , Cálcio/metabolismo , Retículo Endoplasmático/metabolismo , Proteínas Relacionadas à Autofagia/metabolismo , Autofagia , Proteínas de Ciclo Celular/metabolismoRESUMO
Interferons (IFNs) induce an antimicrobial state, protecting tissues from infection. Many viruses inhibit IFN signaling, but whether bacterial pathogens evade IFN responses remains unclear. Here, we demonstrate that the Shigella OspC family of type-III-secreted effectors blocks IFN signaling independently of its cell death inhibitory activity. Rather, IFN inhibition was mediated by the binding of OspC1 and OspC3 to the Ca2+ sensor calmodulin (CaM), blocking CaM kinase II and downstream JAK/STAT signaling. The growth of Shigella lacking OspC1 and OspC3 was attenuated in epithelial cells and in a murine model of infection. This phenotype was rescued in both models by the depletion of IFN receptors. OspC homologs conserved in additional pathogens not only bound CaM but also inhibited IFN, suggesting a widespread virulence strategy. These findings reveal a conserved but previously undescribed molecular mechanism of IFN inhibition and demonstrate the critical role of Ca2+ and IFN targeting in bacterial pathogenesis.
Assuntos
Interferons , Fatores de Virulência , Animais , Antivirais , Sinalização do Cálcio , Células Epiteliais/metabolismo , Interferons/metabolismo , Camundongos , Fatores de Virulência/metabolismoRESUMO
Plant immunity is activated upon pathogen perception and often affects growth and yield when it is constitutively active. How plants fine-tune immune homeostasis in their natural habitats remains elusive. Here, we discover a conserved immune suppression network in cereals that orchestrates immune homeostasis, centering on a Ca2+-sensor, RESISTANCE OF RICE TO DISEASES1 (ROD1). ROD1 promotes reactive oxygen species (ROS) scavenging by stimulating catalase activity, and its protein stability is regulated by ubiquitination. ROD1 disruption confers resistance to multiple pathogens, whereas a natural ROD1 allele prevalent in indica rice with agroecology-specific distribution enhances resistance without yield penalty. The fungal effector AvrPiz-t structurally mimics ROD1 and activates the same ROS-scavenging cascade to suppress host immunity and promote virulence. We thus reveal a molecular framework adopted by both host and pathogen that integrates Ca2+ sensing and ROS homeostasis to suppress plant immunity, suggesting a principle for breeding disease-resistant, high-yield crops.
Assuntos
Cálcio/metabolismo , Sequestradores de Radicais Livres/metabolismo , Proteínas Fúngicas/metabolismo , Oryza/imunologia , Imunidade Vegetal , Proteínas de Plantas/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Sistemas CRISPR-Cas/genética , Membrana Celular/metabolismo , Resistência à Doença/genética , Modelos Biológicos , Oryza/genética , Doenças das Plantas/imunologia , Proteínas de Plantas/genética , Ligação Proteica , Estabilidade Proteica , Reprodução , Especificidade da Espécie , Ubiquitina-Proteína Ligases/metabolismo , Ubiquitinação , Zea mays/imunologiaRESUMO
Neural stem cells (NSCs) in the adult brain transit from the quiescent state to proliferation to produce new neurons. The mechanisms regulating this transition in freely behaving animals are, however, poorly understood. We customized in vivo imaging protocols to follow NSCs for several days up to months, observing their activation kinetics in freely behaving mice. Strikingly, NSC division is more frequent during daylight and is inhibited by darkness-induced melatonin signaling. The inhibition of melatonin receptors affected intracellular Ca2+ dynamics and promoted NSC activation. We further discovered a Ca2+ signature of quiescent versus activated NSCs and showed that several microenvironmental signals converge on intracellular Ca2+ pathways to regulate NSC quiescence and activation. In vivo NSC-specific optogenetic modulation of Ca2+ fluxes to mimic quiescent-state-like Ca2+ dynamics in freely behaving mice blocked NSC activation and maintained their quiescence, pointing to the regulatory mechanisms mediating NSC activation in freely behaving animals.
Assuntos
Células-Tronco Adultas/metabolismo , Cálcio/metabolismo , Ritmo Circadiano , Espaço Intracelular/metabolismo , Células-Tronco Neurais/metabolismo , Células-Tronco Adultas/citologia , Células-Tronco Adultas/efeitos dos fármacos , Animais , Astrócitos/efeitos dos fármacos , Astrócitos/metabolismo , Comportamento Animal/efeitos dos fármacos , Divisão Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Ritmo Circadiano/efeitos dos fármacos , Citosol/metabolismo , Fator de Crescimento Epidérmico/farmacologia , Receptores de Inositol 1,4,5-Trifosfato/metabolismo , Melatonina/metabolismo , Camundongos , Células-Tronco Neurais/citologia , Células-Tronco Neurais/efeitos dos fármacos , Optogenética , Transdução de Sinais/efeitos dos fármacos , Triptaminas/farmacologiaRESUMO
Single-cell transcriptomics has been widely applied to classify neurons in the mammalian brain, while systems neuroscience has historically analyzed the encoding properties of cortical neurons without considering cell types. Here we examine how specific transcriptomic types of mouse prefrontal cortex (PFC) projection neurons relate to axonal projections and encoding properties across multiple cognitive tasks. We found that most types projected to multiple targets, and most targets received projections from multiple types, except PFCâPAG (periaqueductal gray). By comparing Ca2+ activity of the molecularly homogeneous PFCâPAG type against two heterogeneous classes in several two-alternative choice tasks in freely moving mice, we found that all task-related signals assayed were qualitatively present in all examined classes. However, PAG-projecting neurons most potently encoded choice in cued tasks, whereas contralateral PFC-projecting neurons most potently encoded reward context in an uncued task. Thus, task signals are organized redundantly, but with clear quantitative biases across cells of specific molecular-anatomical characteristics.
Assuntos
Cognição/fisiologia , Neurônios/fisiologia , Córtex Pré-Frontal/fisiologia , Análise e Desempenho de Tarefas , Animais , Cálcio/metabolismo , Comportamento de Escolha , Sinais (Psicologia) , Imageamento Tridimensional , Integrases/metabolismo , Camundongos Endogâmicos C57BL , Odorantes , Optogenética , Substância Cinzenta Periaquedutal/fisiologia , Recompensa , Análise de Célula Única , Transcriptoma/genéticaRESUMO
P-type ATPases are found in all kingdoms of life and constitute a wide range of cation transporters, primarily for H+, Na+, K+, Ca2+, and transition metal ions such as Cu(I), Zn(II), and Cd(II). They have been studied through a wide range of techniques, and research has gained very significant insight on their transport mechanism and regulation. Here, we review the structure, function, and dynamics of P2-ATPases including Ca2+-ATPases and Na,K-ATPase. We highlight mechanisms of functional transitions that are associated with ion exchange on either side of the membrane and how the functional cycle is regulated by interaction partners, autoregulatory domains, and off-cycle states. Finally, we discuss future perspectives based on emerging techniques and insights.
Assuntos
Trifosfato de Adenosina/química , ATPases Transportadoras de Cobre/química , ATPase Trocadora de Hidrogênio-Potássio/química , ATPases Transportadoras de Cálcio do Retículo Sarcoplasmático/química , ATPase Trocadora de Sódio-Potássio/química , Trifosfato de Adenosina/metabolismo , Animais , Sítios de Ligação , Cátions Bivalentes , Cátions Monovalentes , ATPases Transportadoras de Cobre/genética , ATPases Transportadoras de Cobre/metabolismo , ATPase Trocadora de Hidrogênio-Potássio/genética , ATPase Trocadora de Hidrogênio-Potássio/metabolismo , Humanos , Transporte de Íons , Modelos Moleculares , Ligação Proteica , Domínios e Motivos de Interação entre Proteínas , Estrutura Secundária de Proteína , Prótons , ATPases Transportadoras de Cálcio do Retículo Sarcoplasmático/genética , ATPases Transportadoras de Cálcio do Retículo Sarcoplasmático/metabolismo , Imagem Individual de Molécula , ATPase Trocadora de Sódio-Potássio/genética , ATPase Trocadora de Sódio-Potássio/metabolismo , Especificidade por SubstratoRESUMO
Cognitive faculties such as imagination, planning, and decision-making entail the ability to represent hypothetical experience. Crucially, animal behavior in natural settings implies that the brain can represent hypothetical future experience not only quickly but also constantly over time, as external events continually unfold. To determine how this is possible, we recorded neural activity in the hippocampus of rats navigating a maze with multiple spatial paths. We found neural activity encoding two possible future scenarios (two upcoming maze paths) in constant alternation at 8 Hz: one scenario per â¼125-ms cycle. Further, we found that the underlying dynamics of cycling (both inter- and intra-cycle dynamics) generalized across qualitatively different representational correlates (location and direction). Notably, cycling occurred across moving behaviors, including during running. These findings identify a general dynamic process capable of quickly and continually representing hypothetical experience, including that of multiple possible futures.
Assuntos
Comportamento Animal/fisiologia , Cognição/fisiologia , Tomada de Decisões/fisiologia , Hipocampo/fisiologia , Potenciais de Ação/fisiologia , Animais , Locomoção/fisiologia , Masculino , Aprendizagem em Labirinto/fisiologia , Rede Nervosa/fisiologia , Neurônios/fisiologia , Ratos , Ratos Long-Evans , Ritmo Teta/fisiologiaRESUMO
Oncogenic transformation is associated with profound changes in cellular metabolism, but whether tracking these can improve disease stratification or influence therapy decision-making is largely unknown. Using the iKnife to sample the aerosol of cauterized specimens, we demonstrate a new mode of real-time diagnosis, coupling metabolic phenotype to mutant PIK3CA genotype. Oncogenic PIK3CA results in an increase in arachidonic acid and a concomitant overproduction of eicosanoids, acting to promote cell proliferation beyond a cell-autonomous manner. Mechanistically, mutant PIK3CA drives a multimodal signaling network involving mTORC2-PKCζ-mediated activation of the calcium-dependent phospholipase A2 (cPLA2). Notably, inhibiting cPLA2 synergizes with fatty acid-free diet to restore immunogenicity and selectively reduce mutant PIK3CA-induced tumorigenicity. Besides highlighting the potential for metabolic phenotyping in stratified medicine, this study reveals an important role for activated PI3K signaling in regulating arachidonic acid metabolism, uncovering a targetable metabolic vulnerability that largely depends on dietary fat restriction. VIDEO ABSTRACT.
Assuntos
Ácido Araquidônico/análise , Classe I de Fosfatidilinositol 3-Quinases/metabolismo , Eicosanoides/metabolismo , Animais , Ácido Araquidônico/metabolismo , Linhagem Celular Tumoral , Classe I de Fosfatidilinositol 3-Quinases/genética , Citosol/metabolismo , Eicosanoides/fisiologia , Ativação Enzimática , Feminino , Humanos , Metabolismo dos Lipídeos/fisiologia , Alvo Mecanístico do Complexo 2 de Rapamicina/metabolismo , Redes e Vias Metabólicas/genética , Redes e Vias Metabólicas/fisiologia , Camundongos Endogâmicos BALB C , Camundongos Nus , Fosfatidilinositol 3-Quinases/metabolismo , Fosfolipases A2/metabolismo , Fosforilação , Proteína Quinase C/metabolismo , Transdução de Sinais , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Varying pH of luminal fluid along the female reproductive tract is a physiological cue that modulates sperm motility. CatSper is a sperm-specific, pH-sensitive calcium channel essential for hyperactivated motility and male fertility. Multi-subunit CatSper channel complexes organize linear Ca2+ signaling nanodomains along the sperm tail. Here, we identify EF-hand calcium-binding domain-containing protein 9 (EFCAB9) as a bifunctional, cytoplasmic machine modulating the channel activity and the domain organization of CatSper. Knockout mice studies demonstrate that EFCAB9, in complex with the CatSper subunit, CATSPERζ, is essential for pH-dependent and Ca2+-sensitive activation of the CatSper channel. In the absence of EFCAB9, sperm motility and fertility is compromised, and the linear arrangement of the Ca2+ signaling domains is disrupted. EFCAB9 interacts directly with CATSPERζ in a Ca2+-dependent manner and dissociates at elevated pH. These observations suggest that EFCAB9 is a long-sought, intracellular, pH-dependent Ca2+ sensor that triggers changes in sperm motility.
Assuntos
Proteínas de Ligação ao Cálcio/metabolismo , Motilidade dos Espermatozoides/fisiologia , Animais , Cálcio/metabolismo , Canais de Cálcio/metabolismo , Sinalização do Cálcio/fisiologia , Proteínas de Ligação ao Cálcio/fisiologia , Linhagem Celular , Membrana Celular/metabolismo , Fertilidade , Células HEK293 , Humanos , Concentração de Íons de Hidrogênio , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Espermatozoides/metabolismoRESUMO
The L-type voltage-gated Ca2+ (Cav) channels are modulated by various compounds exemplified by 1,4-dihydropyridines (DHP), benzothiazepines (BTZ), and phenylalkylamines (PAA), many of which have been used for characterizing channel properties and for treatment of hypertension and other disorders. Here, we report the cryoelectron microscopy (cryo-EM) structures of Cav1.1 in complex with archetypal antagonistic drugs, nifedipine, diltiazem, and verapamil, at resolutions of 2.9 Å, 3.0 Å, and 2.7 Å, respectively, and with a DHP agonist Bay K 8644 at 2.8 Å. Diltiazem and verapamil traverse the central cavity of the pore domain, directly blocking ion permeation. Although nifedipine and Bay K 8644 occupy the same fenestration site at the interface of repeats III and IV, the coordination details support previous functional observations that Bay K 8644 is less favored in the inactivated state. These structures elucidate the modes of action of different Cav ligands and establish a framework for structure-guided drug discovery.
Assuntos
Bloqueadores dos Canais de Cálcio/metabolismo , Canais de Cálcio Tipo L/metabolismo , Canais de Cálcio Tipo L/ultraestrutura , Éster Metílico do Ácido 3-Piridinacarboxílico, 1,4-Di-Hidro-2,6-Dimetil-5-Nitro-4-(2-(Trifluormetil)fenil) , Sequência de Aminoácidos , Animais , Sítios de Ligação , Canais de Cálcio/metabolismo , Canais de Cálcio/fisiologia , Canais de Cálcio/ultraestrutura , Canais de Cálcio Tipo L/fisiologia , Microscopia Crioeletrônica , Diltiazem , Ligantes , Masculino , Modelos Moleculares , Nifedipino , Coelhos , VerapamilRESUMO
Calcium imaging using two-photon scanning microscopy has become an essential tool in neuroscience. However, in its typical implementation, the tradeoffs between fields of view, acquisition speeds, and depth restrictions in scattering brain tissue pose severe limitations. Here, using an integrated systems-wide optimization approach combined with multiple technical innovations, we introduce a new design paradigm for optical microscopy based on maximizing biological information while maintaining the fidelity of obtained neuron signals. Our modular design utilizes hybrid multi-photon acquisition and allows volumetric recording of neuroactivity at single-cell resolution within up to 1 × 1 × 1.22 mm volumes at up to 17 Hz in awake behaving mice. We establish the capabilities and potential of the different configurations of our imaging system at depth and across brain regions by applying it to in vivo recording of up to 12,000 neurons in mouse auditory cortex, posterior parietal cortex, and hippocampus.
Assuntos
Microscopia/métodos , Imagem Molecular/métodos , Neuroimagem/métodos , Animais , Encéfalo/fisiologia , Cálcio/metabolismo , Feminino , Hipocampo/fisiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Neurônios/fisiologia , Análise de Célula Única/métodosRESUMO
Expansion of CAG trinucleotide repeats in ATXN1 causes spinocerebellar ataxia type 1 (SCA1), a neurodegenerative disease that impairs coordination and cognition. While ATXN1 is associated with increased Alzheimer's disease (AD) risk, CAG repeat number in AD patients is not changed. Here, we investigated the consequences of ataxin-1 loss of function and discovered that knockout of Atxn1 reduced CIC-ETV4/5-mediated inhibition of Bace1 transcription, leading to increased BACE1 levels and enhanced amyloidogenic cleavage of APP, selectively in AD-vulnerable brain regions. Elevated BACE1 expression exacerbated Aß deposition and gliosis in AD mouse models and impaired hippocampal neurogenesis and olfactory axonal targeting. In SCA1 mice, polyglutamine-expanded mutant ataxin-1 led to the increase of BACE1 post-transcriptionally, both in cerebrum and cerebellum, and caused axonal-targeting deficit and neurodegeneration in the hippocampal CA2 region. These findings suggest that loss of ataxin-1 elevates BACE1 expression and Aß pathology, rendering it a potential contributor to AD risk and pathogenesis.
Assuntos
Doença de Alzheimer/patologia , Secretases da Proteína Precursora do Amiloide/metabolismo , Ataxina-1/metabolismo , Encéfalo/metabolismo , Doença de Alzheimer/metabolismo , Secretases da Proteína Precursora do Amiloide/genética , Precursor de Proteína beta-Amiloide/metabolismo , Animais , Ataxina-1/deficiência , Ataxina-1/genética , Encéfalo/patologia , Região CA2 Hipocampal/metabolismo , Região CA2 Hipocampal/patologia , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Modelos Animais de Doenças , Feminino , Frequência do Gene , Humanos , Masculino , Camundongos , Camundongos Transgênicos , Neurogênese , Proteínas Proto-Oncogênicas c-ets/genética , Proteínas Proto-Oncogênicas c-ets/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Transcrição Gênica , Repetições de Trinucleotídeos/genética , Regulação para CimaRESUMO
To decipher dynamic brain information processing, current genetically encoded calcium indicators (GECIs) are limited in single action potential (AP) detection speed, combinatorial spectral compatibility, and two-photon imaging depth. To address this, here, we rationally engineered a next-generation quadricolor GECI suite, XCaMPs. Single AP detection was achieved within 3-10 ms of spike onset, enabling measurements of fast-spike trains in parvalbumin (PV)-positive interneurons in the barrel cortex in vivo and recording three distinct (two inhibitory and one excitatory) ensembles during pre-motion activity in freely moving mice. In vivo paired recording of pre- and postsynaptic firing revealed spatiotemporal constraints of dendritic inhibition in layer 1 in vivo, between axons of somatostatin (SST)-positive interneurons and apical tufts dendrites of excitatory pyramidal neurons. Finally, non-invasive, subcortical imaging using red XCaMP-R uncovered somatosensation-evoked persistent activity in hippocampal CA1 neurons. Thus, the XCaMPs offer a critical enhancement of solution space in studies of complex neuronal circuit dynamics. VIDEO ABSTRACT.
Assuntos
Potenciais de Ação/fisiologia , Axônios/metabolismo , Córtex Cerebral/metabolismo , Hipocampo/metabolismo , Interneurônios/metabolismo , Células Piramidais/metabolismo , Animais , Córtex Cerebral/citologia , Feminino , Hipocampo/citologia , Interneurônios/citologia , Camundongos , Camundongos Transgênicos , Células Piramidais/citologia , Ratos , Ratos Sprague-DawleyRESUMO
The endoplasmic reticulum (ER) has a broad localization throughout the cell and forms direct physical contacts with all other classes of membranous organelles, including the plasma membrane (PM). A number of protein tethers that mediate these contacts have been identified, and study of these protein tethers has revealed a multiplicity of roles in cell physiology, including regulation of intracellular Ca2+ dynamics and signaling as well as control of lipid traffic and homeostasis. In this review, we discuss the cross talk between the ER and the PM mediated by direct contacts. We review factors that tether the two membranes, their properties, and their dynamics in response to the functional state of the cell. We focus in particular on the role of ER-PM contacts in nonvesicular lipid transport between the two bilayers mediated by lipid transfer proteins.
Assuntos
Cálcio/metabolismo , Proteínas de Transporte/metabolismo , Membrana Celular/metabolismo , Retículo Endoplasmático/metabolismo , Proteínas de Membrana/metabolismo , Proteína ORAI1/metabolismo , Transporte Biológico , Sinalização do Cálcio , Proteínas de Transporte/genética , Membrana Celular/ultraestrutura , Retículo Endoplasmático/ultraestrutura , Células Eucarióticas/metabolismo , Células Eucarióticas/ultraestrutura , Expressão Gênica , Homeostase , Humanos , Proteínas de Membrana/genética , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Proteína ORAI1/genética , Receptores de Esteroides/genética , Receptores de Esteroides/metabolismo , Molécula 1 de Interação Estromal/genética , Molécula 1 de Interação Estromal/metabolismo , Sinaptotagminas/genética , Sinaptotagminas/metabolismoRESUMO
The medial amygdala (MeA) plays a critical role in processing species- and sex-specific signals that trigger social and defensive behaviors. However, the principles by which this deep brain structure encodes social information is poorly understood. We used a miniature microscope to image the Ca2+ dynamics of large neural ensembles in awake behaving mice and tracked the responses of MeA neurons over several months. These recordings revealed spatially intermingled subsets of MeA neurons with distinct temporal dynamics. The encoding of social information in the MeA differed between males and females and relied on information from both individual cells and neuronal populations. By performing long-term Ca2+ imaging across different social contexts, we found that sexual experience triggers lasting and sex-specific changes in MeA activity, which, in males, involve signaling by oxytocin. These findings reveal basic principles underlying the brain's representation of social information and its modulation by intrinsic and extrinsic factors.
Assuntos
Tonsila do Cerebelo/fisiologia , Neurônios/citologia , Vigília , Tonsila do Cerebelo/citologia , Animais , Comportamento Animal , Sinais (Psicologia) , Endoscopia/métodos , Feminino , Masculino , Camundongos , Microscopia/métodos , Ocitocina/fisiologia , Caracteres Sexuais , Comportamento Sexual Animal , Comportamento SocialRESUMO
Reward-seeking behavior is fundamental to survival, but suppression of this behavior can be essential as well, even for rewards of high value. In humans and rodents, the medial prefrontal cortex (mPFC) has been implicated in suppressing reward seeking; however, despite vital significance in health and disease, the neural circuitry through which mPFC regulates reward seeking remains incompletely understood. Here, we show that a specific subset of superficial mPFC projections to a subfield of nucleus accumbens (NAc) neurons naturally encodes the decision to initiate or suppress reward seeking when faced with risk of punishment. A highly resolved subpopulation of these top-down projecting neurons, identified by 2-photon Ca2+ imaging and activity-dependent labeling to recruit the relevant neurons, was found capable of suppressing reward seeking. This natural activity-resolved mPFC-to-NAc projection displayed unique molecular-genetic and microcircuit-level features concordant with a conserved role in the regulation of reward-seeking behavior, providing cellular and anatomical identifiers of behavioral and possible therapeutic significance.
Assuntos
Recompensa , Animais , Comportamento Animal , Feminino , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Vias Neurais , Neuroimagem , Córtex Pré-Frontal/citologia , Córtex Pré-Frontal/metabolismo , PuniçãoRESUMO
In the last 5 years, most of the molecules that control mitochondrial Ca(2+) homeostasis have been finally identified. Mitochondrial Ca(2+) uptake is mediated by the Mitochondrial Calcium Uniporter (MCU) complex, a macromolecular structure that guarantees Ca(2+) accumulation inside mitochondrial matrix upon increases in cytosolic Ca(2+). Conversely, Ca(2+) release is under the control of the Na(+)/Ca(2+) exchanger, encoded by the NCLX gene, and of a H(+)/Ca(2+) antiporter, whose identity is still debated. The low affinity of the MCU complex, coupled to the activity of the efflux systems, protects cells from continuous futile cycles of Ca(2+) across the inner mitochondrial membrane and consequent massive energy dissipation. In this review, we discuss the basic principles that govern mitochondrial Ca(2+) homeostasis and the methods used to investigate the dynamics of Ca(2+) concentration within the organelles. We discuss the functional and structural role of the different molecules involved in mitochondrial Ca(2+) handling and their pathophysiological role.
Assuntos
Canais de Cálcio/metabolismo , Cálcio/metabolismo , Mitocôndrias/metabolismo , Trocador de Sódio e Cálcio/metabolismo , Animais , Canais de Cálcio/química , Canais de Cálcio/genética , Sinalização do Cálcio , Regulação da Expressão Gênica , Homeostase , Humanos , Transporte de Íons , Cinética , Mitocôndrias/genética , Mitocôndrias/ultraestrutura , Membranas Mitocondriais/metabolismo , Membranas Mitocondriais/ultraestrutura , Proteínas Mitocondriais , Modelos Moleculares , Trocador de Sódio e Cálcio/genética , TermodinâmicaRESUMO
The gastrointestinal (GI) tract displays multiple motor patterns that move nutrients and wastes through the body. Smooth muscle cells (SMCs) provide the forces necessary for GI motility, but interstitial cells, electrically coupled to SMCs, tune SMC excitability, transduce inputs from enteric motor neurons, and generate pacemaker activity that underlies major motor patterns, such as peristalsis and segmentation. The interstitial cells regulating SMCs are interstitial cells of Cajal (ICC) and PDGF receptor (PDGFR)α+ cells. Together these cells form the SIP syncytium. ICC and PDGFRα+ cells express signature Ca2+-dependent conductances: ICC express Ca2+-activated Cl- channels, encoded by Ano1, that generate inward current, and PDGFRα+ cells express Ca2+-activated K+ channels, encoded by Kcnn3, that generate outward current. The open probabilities of interstitial cell conductances are controlled by Ca2+ release from the endoplasmic reticulum. The resulting Ca2+ transients occur spontaneously in a stochastic manner. Ca2+ transients in ICC induce spontaneous transient inward currents and spontaneous transient depolarizations (STDs). Neurotransmission increases or decreases Ca2+ transients, and the resulting depolarizing or hyperpolarizing responses conduct to other cells in the SIP syncytium. In pacemaker ICC, STDs activate voltage-dependent Ca2+ influx, which initiates a cluster of Ca2+ transients and sustains activation of ANO1 channels and depolarization during slow waves. Regulation of GI motility has traditionally been described as neurogenic and myogenic. Recent advances in understanding Ca2+ handling mechanisms in interstitial cells and how these mechanisms influence motor patterns of the GI tract suggest that the term "myogenic" should be replaced by the term "SIPgenic," as this review discusses.