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1.
Sci Rep ; 10(1): 11715, 2020 07 16.
Artigo em Inglês | MEDLINE | ID: mdl-32678166

RESUMO

Although convincing experimental evidence indicates that Na+/K+/Cl- cotransporter 1 (NKCC1) is involved in spinal nociceptive information processing and in the generation of hyperalgesia and allodynia in chronic pain states, the cellular distribution of NKCC1 in the superficial spinal dorsal horn is still poorly understood. Because this important piece of knowledge is missing, the effect of NKCC1 on pain processing is still open to conflicting interpretations. In this study, to provide the missing experimental data, we investigated the cellular distribution of NKCC1 in the superficial spinal dorsal horn by immunohistochemical methods. We demonstrated for the first time that almost all spinal axon terminals of peptidergic nociceptive primary afferents express NKCC1. In contrast, virtually all spinal axon terminals of nonpeptidergic nociceptive primary afferents were negative for NKCC1. Data on the colocalization of NKCC1 with axonal and glial markers indicated that it is almost exclusively expressed by axon terminals and glial cells in laminae I-IIo. In lamina IIi, however, we observed a strong immunostaining for NKCC1 also in the dendrites and cell bodies of PV-containing inhibitory neurons and a weak staining in PKCγ-containing excitatory neurons. Our results facilitate further thinking about the role of NKCC1 in spinal pain processing.


Assuntos
Neuroglia/metabolismo , Células do Corno Posterior/metabolismo , Transdução de Sinais/genética , Membro 2 da Família 12 de Carreador de Soluto/metabolismo , Corno Dorsal da Medula Espinal/citologia , Animais , Dor Crônica/metabolismo , Técnicas de Inativação de Genes , Hiperalgesia/metabolismo , Imuno-Histoquímica , Camundongos , Camundongos Knockout , Terminações Pré-Sinápticas/metabolismo , Ratos , Ratos Wistar , Membro 2 da Família 12 de Carreador de Soluto/genética
2.
J Comp Neurol ; 523(13): 1967-83, 2015 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-25764511

RESUMO

γ-Aminobutyric acid (GABA)- and glycine-mediated hyperpolarizing inhibition is associated with a chloride influx that depends on the inwardly directed chloride electrochemical gradient. In neurons, the extrusion of chloride from the cytosol primarily depends on the expression of an isoform of potassium-chloride cotransporters (KCC2s). KCC2 is crucial in the regulation of the inhibitory tone of neural circuits, including pain processing neural assemblies. Thus we investigated the cellular distribution of KCC2 in neurons underlying pain processing in the superficial spinal dorsal horn of rats by using high-resolution immunocytochemical methods. We demonstrated that perikarya and dendrites widely expressed KCC2, but axon terminals proved to be negative for KCC2. In single ultrathin sections, silver deposits labeling KCC2 molecules showed different densities on the surface of dendritic profiles, some of which were negative for KCC2. In freeze fracture replicas and tissue sections double stained for the ß3-subunit of GABAA receptors and KCC2, GABAA receptors were revealed on dendritic segments with high and also with low KCC2 densities. By measuring the distances between spots immunoreactive for gephyrin (a scaffolding protein of GABAA and glycine receptors) and KCC2 on the surface of neurokinin 1 (NK1) receptor-immunoreactive dendrites, we found that gephyrin-immunoreactive spots were located at various distances from KCC2 cotransporters; 5.7 % of them were recovered in the middle of 4-10-µm-long dendritic segments that were free of KCC2 immunostaining. The variable local densities of KCC2 may result in variable postsynaptic potentials evoked by the activation of GABAA and glycine receptors along the dendrites of spinal neurons.


Assuntos
Células do Corno Posterior/metabolismo , Corno Dorsal da Medula Espinal/citologia , Simportadores/metabolismo , Animais , Peptídeo Relacionado com Gene de Calcitonina/metabolismo , Proteínas de Transporte/metabolismo , Glutamato Descarboxilase/metabolismo , Masculino , Proteínas de Membrana/metabolismo , Microscopia Imunoeletrônica , Células do Corno Posterior/citologia , Células do Corno Posterior/diagnóstico por imagem , Terminações Pré-Sinápticas/metabolismo , Terminações Pré-Sinápticas/ultraestrutura , Ratos , Ratos Wistar , Receptores de GABA-A/metabolismo , Receptores da Neurocinina-1/metabolismo , Ultrassonografia , Proteína Vesicular 2 de Transporte de Glutamato/metabolismo , Proteína Vesicular 2 de Transporte de Glutamato/ultraestrutura , Cotransportadores de K e Cl-
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