Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 7 de 7
Filtrar
Mais filtros










Base de dados
Intervalo de ano de publicação
1.
J Physiol ; 598(12): 2431-2452, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32304329

RESUMO

KEY POINTS: CAST/ELKS are positive regulators of presynaptic growth and are suppressors of active zone expansion at the developing mouse calyx of Held. CAST/ELKS regulate all three CaV 2 subtype channel levels in the presynaptic terminal and not just CaV 2.1. The half-life of ELKS is on the timescale of days and not weeks. Synaptic transmission was not impacted by the loss of CAST/ELKS. CAST/ELKS are involved in pathways regulating morphological properties of presynaptic terminals during an early stage of circuit maturation. ABSTRACT: Many presynaptic active zone (AZ) proteins have multiple regulatory roles that vary during distinct stages of neuronal circuit development. The CAST/ELKS protein family are evolutionarily conserved presynaptic AZ molecules that regulate presynaptic calcium channels, synaptic transmission and plasticity in the mammalian CNS. However, how these proteins regulate synapse development and presynaptic function in a developing neuronal circuit in its native environment is unclear. To unravel the roles of CAST/ELKS in glutamatergic synapse development and in presynaptic function, we used CAST knockout (KO) and ELKS conditional KO (CKO) mice to examine how their loss during the early stages of circuit maturation impacted the calyx of Held presynaptic terminal development and function. Morphological analysis from confocal z-stacks revealed that combined deletion of CAST/ELKS resulted in a reduction in the surface area and volume of the calyx. Analysis of AZ ultrastructure showed that AZ size was increased in the absence of CAST/ELKS. Patch clamp recordings demonstrated a reduction of all presynaptic CaV 2 channel subtype currents that correlated with a loss in presynaptic CaV 2 channel numbers. However, these changes did not impair synaptic transmission and plasticity and synaptic vesicle release kinetics. We conclude that CAST/ELKS proteins are positive regulators of presynaptic growth and are suppressors of AZ expansion and CaV 2 subtype currents and levels during calyx of Held development. We propose that CAST/ELKS are involved in pathways regulating presynaptic morphological properties and CaV 2 channel subtypes and suggest there is developmental compensation to preserve synaptic transmission during early stages of neuronal circuit maturation.


Assuntos
Terminações Pré-Sinápticas , Sinapses , Animais , Canais de Cálcio , Camundongos , Transmissão Sináptica , Vesículas Sinápticas
2.
Neuron ; 101(2): 260-273.e6, 2019 01 16.
Artigo em Inglês | MEDLINE | ID: mdl-30545599

RESUMO

The abundance of presynaptic CaV2 voltage-gated Ca2+ channels (CaV2) at mammalian active zones (AZs) regulates the efficacy of synaptic transmission. It is proposed that presynaptic CaV2 levels are saturated in AZs due to a finite number of slots that set CaV2 subtype abundance and that CaV2.1 cannot compete for CaV2.2 slots. However, at most AZs, CaV2.1 levels are highest and CaV2.2 levels are developmentally reduced. To investigate CaV2.1 saturation states and preference in AZs, we overexpressed the CaV2.1 and CaV2.2 α1 subunits at the calyx of Held at immature and mature developmental stages. We found that AZs prefer CaV2.1 to CaV2.2. Remarkably, CaV2.1 α1 subunit overexpression drove increased CaV2.1 currents and channel numbers and increased synaptic strength at both developmental stages examined. Therefore, we propose that CaV2.1 levels in the AZ are not saturated and that synaptic strength can be modulated by increasing CaV2.1 levels to regulate neuronal circuit output. VIDEO ABSTRACT.


Assuntos
Tronco Encefálico/citologia , Canais de Cálcio Tipo N/metabolismo , Terminações Pré-Sinápticas/fisiologia , Sinapses/fisiologia , Transmissão Sináptica/genética , Animais , Animais Recém-Nascidos , Biofísica , Cloreto de Cádmio/farmacologia , Cálcio/metabolismo , Bloqueadores dos Canais de Cálcio/farmacologia , Canais de Cálcio Tipo N/genética , Estimulação Elétrica , Feminino , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Neurotransmissores/metabolismo , Terminações Pré-Sinápticas/ultraestrutura , Sinapses/ultraestrutura
3.
Elife ; 62017 08 08.
Artigo em Inglês | MEDLINE | ID: mdl-28786379

RESUMO

In central nervous system (CNS) synapses, action potential-evoked neurotransmitter release is principally mediated by CaV2.1 calcium channels (CaV2.1) and is highly dependent on the physical distance between CaV2.1 and synaptic vesicles (coupling). Although various active zone proteins are proposed to control coupling and abundance of CaV2.1 through direct interactions with the CaV2.1 α1 subunit C-terminus at the active zone, the role of these interaction partners is controversial. To define the intrinsic motifs that regulate coupling, we expressed mutant CaV2.1 α1 subunits on a CaV2.1 null background at the calyx of Held presynaptic terminal. Our results identified a region that directly controlled fast synaptic vesicle release and vesicle docking at the active zone independent of CaV2.1 abundance. In addition, proposed individual direct interactions with active zone proteins are insufficient for CaV2.1 abundance and coupling. Therefore, our work advances our molecular understanding of CaV2.1 regulation of neurotransmitter release in mammalian CNS synapses.


Assuntos
Canais de Cálcio Tipo N/metabolismo , Fusão de Membrana , Terminações Pré-Sinápticas/enzimologia , Vesículas Sinápticas/metabolismo , Animais , Canais de Cálcio Tipo N/genética , Análise Mutacional de DNA , Camundongos , Neurotransmissores/metabolismo
4.
PLoS One ; 8(10): e77998, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-24205061

RESUMO

Primary sensory afferents of the dorsal root and trigeminal ganglia constantly transmit sensory information depicting the individual's physical and chemical environment to higher brain regions. Beyond the typical trigeminal stimuli (e.g. irritants), environmental stimuli comprise a plethora of volatile chemicals with olfactory components (odorants). In spite of a complete loss of their sense of smell, anosmic patients may retain the ability to roughly discriminate between different volatile compounds. While the detailed mechanisms remain elusive, sensory structures belonging to the trigeminal system seem to be responsible for this phenomenon. In order to gain a better understanding of the mechanisms underlying the activation of the trigeminal system by volatile chemicals, we investigated odorant-induced membrane potential changes in cultured rat trigeminal neurons induced by the odorants vanillin, heliotropyl acetone, helional, and geraniol. We observed the dose-dependent depolarization of trigeminal neurons upon application of these substances occurring in a stimulus-specific manner and could show that distinct neuronal populations respond to different odorants. Using specific antagonists, we found evidence that TRPA1, TRPM8, and/or TRPV1 contribute to the activation. In order to further test this hypothesis, we used recombinantly expressed rat and human variants of these channels to investigate whether they are indeed activated by the odorants tested. We additionally found that the odorants dose-dependently inhibit two-pore potassium channels TASK1 and TASK3 heterologously expressed In Xenopus laevis oocytes. We suggest that the capability of various odorants to activate different TRP channels and to inhibit potassium channels causes neuronal depolarization and activation of distinct subpopulations of trigeminal sensory neurons, forming the basis for a specific representation of volatile chemicals in the trigeminal ganglia.


Assuntos
Neurônios/metabolismo , Canais de Potencial de Receptor Transitório/metabolismo , Gânglio Trigeminal/citologia , Compostos Orgânicos Voláteis/farmacologia , Monoterpenos Acíclicos , Animais , Benzaldeídos/farmacologia , Células Cultivadas , Humanos , Proteínas do Tecido Nervoso , Neurônios/efeitos dos fármacos , Canais de Potássio de Domínios Poros em Tandem/antagonistas & inibidores , Canais de Potássio de Domínios Poros em Tandem/genética , Canais de Potássio de Domínios Poros em Tandem/metabolismo , Ratos , Canal de Cátion TRPA1 , Canais de Cátion TRPC/antagonistas & inibidores , Canais de Cátion TRPC/genética , Canais de Cátion TRPC/metabolismo , Canais de Cátion TRPM/antagonistas & inibidores , Canais de Cátion TRPM/genética , Canais de Cátion TRPM/metabolismo , Canais de Cátion TRPV/antagonistas & inibidores , Canais de Cátion TRPV/genética , Canais de Cátion TRPV/metabolismo , Terpenos/farmacologia , Canais de Potencial de Receptor Transitório/antagonistas & inibidores , Canais de Potencial de Receptor Transitório/genética
5.
Front Syst Neurosci ; 7: 64, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-24115922

RESUMO

Albeit lacking a sense of smell, anosmic patients maintain a reduced ability to distinguish different volatile chemicals by relying exclusively on their trigeminal system (TS). To elucidate differences in the neuronal representation of these volatile substances in the TS, we performed voltage-sensitive dye imaging (VSDI) in the rat trigeminal ganglion (TG) in vivo. We demonstrated that stimulus-specific patterns of bioelectrical activity occur within the TG upon nasal administration of ten different volatile chemicals. With regard to spatial differences between the evoked trigeminal response patterns, these substances could be sorted into three groups. Signal intensity and onset latencies were also dependent on the administered stimulus and its concentration. We conclude that particular compounds detected by the TS are represented by (1) a specific spatial response pattern, (2) the signal intensity, and (3) onset latencies within the pattern. Jointly, these trigeminal representations may contribute to the surprisingly high discriminative skills of anosmic patients.

6.
PLoS One ; 7(11): e48005, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-23144843

RESUMO

Intracellular Cl(-) concentrations ([Cl(-)](i)) of sensory neurons regulate signal transmission and signal amplification. In dorsal root ganglion (DRG) and olfactory sensory neurons (OSNs), Cl(-) is accumulated by the Na(+)-K(+)-2Cl(-) cotransporter 1 (NKCC1), resulting in a [Cl(-)](i) above electrochemical equilibrium and a depolarizing Cl(-) efflux upon Cl(-) channel opening. Here, we investigate the [Cl(-)](i) and function of Cl(-) in primary sensory neurons of trigeminal ganglia (TG) of wild type (WT) and NKCC1(-/-) mice using pharmacological and imaging approaches, patch-clamping, as well as behavioral testing. The [Cl(-)](i) of WT TG neurons indicated active NKCC1-dependent Cl(-) accumulation. Gamma-aminobutyric acid (GABA)(A) receptor activation induced a reduction of [Cl(-)](i) as well as Ca(2+) transients in a corresponding fraction of TG neurons. Ca(2+) transients were sensitive to inhibition of NKCC1 and voltage-gated Ca(2+) channels (VGCCs). Ca(2+) responses induced by capsaicin, a prototypical stimulus of transient receptor potential vanilloid subfamily member-1 (TRPV1) were diminished in NKCC1(-/-) TG neurons, but elevated under conditions of a lowered [Cl(-)](o) suggesting a Cl(-)-dependent amplification of capsaicin-induced responses. Using next generation sequencing (NGS), we found expression of different Ca(2+)-activated Cl(-) channels (CaCCs) in TGs of mice. Pharmacological inhibition of CaCCs reduced the amplitude of capsaicin-induced responses of TG neurons in Ca(2+) imaging and electrophysiological recordings. In a behavioral paradigm, NKCC1(-/-) mice showed less avoidance of the aversive stimulus capsaicin. In summary, our results strongly argue for a Ca(2+)-activated Cl(-)-dependent signal amplification mechanism in TG neurons that requires intracellular Cl(-) accumulation by NKCC1 and the activation of CaCCs.


Assuntos
Capsaicina/farmacologia , Cloretos/metabolismo , Neurônios/metabolismo , Gânglio Trigeminal/citologia , Animais , Agonistas dos Canais de Cálcio/farmacologia , Bloqueadores dos Canais de Cálcio/farmacologia , Sinalização do Cálcio , Células Cultivadas , Canais de Cloreto/genética , Canais de Cloreto/metabolismo , Cloretos/fisiologia , Feminino , Antagonistas de Receptores de GABA-A/farmacologia , Expressão Gênica , Células HEK293 , Humanos , Masculino , Potenciais da Membrana , Camundongos , Camundongos Knockout , Neurônios/efeitos dos fármacos , Cultura Primária de Células , Piridazinas/farmacologia , Receptores de GABA-A/metabolismo , Simportadores de Cloreto de Sódio-Potássio/genética , Simportadores de Cloreto de Sódio-Potássio/metabolismo , Membro 2 da Família 12 de Carreador de Soluto , Transmissão Sináptica , Canais de Cátion TRPV/genética , Canais de Cátion TRPV/metabolismo , Transcriptoma , Gânglio Trigeminal/efeitos dos fármacos
7.
Hum Mol Genet ; 20(6): 1197-211, 2011 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-21212098

RESUMO

We recently described mitochondrial pathology in neurons of transgenic mice with genes associated with Parkinson's disease (PD). Now we describe severe mitochondrial damage in glial cells of the mesencephalon in mice carrying a targeted deletion of parkin (PaKO) or overexpressing doubly mutated human alpha-synuclein (asyn). The number of mitochondria with altered morphology in glial cells is cell type-dependent, but always higher than in neurons. Interestingly, mitochondrial damage also occurs in mesencephalic glia of mice carrying mutated asyn controlled by the tyrosine hydroxylase promoter. Such mice do not show glial expression of the transgene, but show expression in neighboring neurons. However, we found strong overexpression of endogenous asyn in mesencephalic astrocytes from these mice. Cortical astrocytes neither display enhanced asyn expression nor mitochondrial damage. Cultivated mesencephalic astrocytes from newborn transgenic mice display various functional defects along with the morphological damage of mitochondria. First, the mitochondrial Ca(2+)-storage capacity is reduced in asyn transgenic mesencephalic astrocytes, but not in astrocytes from PaKO. Second, the expression of the mitochondrial protein PTEN-induced putative kinase is constitutively increased in asyn transgenic mice, while in PaKO it reacts to oxidative stress by overexpressing this protein along with other mitochondria-related proteins. Third, the neurotrophic effects exerted by control astrocytes, stimulating cortical neurons from healthy mice to develop longer processes and larger neuronal areas, are lacking in co-cultures with transgenic mesencephalic astrocytes. In summary, glial mitochondria from transgenic mice display morphological and functional alterations. Such transgenic astrocytes fail to influence neuronal differentiation, reflecting an important role that glia may play in PD pathogenesis.


Assuntos
Modelos Animais de Doenças , Camundongos , Mitocôndrias/metabolismo , Neuroglia/metabolismo , Doença de Parkinson/metabolismo , Doença de Parkinson/patologia , Animais , Astrócitos/metabolismo , Astrócitos/patologia , Astrócitos/ultraestrutura , Cálcio/metabolismo , Células Cultivadas , Humanos , Masculino , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Mitocôndrias/genética , Mitocôndrias/patologia , Mitocôndrias/ultraestrutura , Neuroglia/patologia , Neuroglia/ultraestrutura , Neurônios/metabolismo , Neurônios/patologia , Neurônios/ultraestrutura , Doença de Parkinson/genética , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/metabolismo
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA
...