RESUMEN
Inhibition of rat neuronal Na(+)/K(+)-ATPase alpha3 isoform at low (100 nM) ouabain concentration led to activation of MAP kinase cascade via PKC and PIP(3) kinase. In contrast to ouabain-sensitive alpha3 isoform of Na(+)/K(+)-ATPase, an ouabain-resistant alpha1 isoform (inhibition with 1 mM of ouabain) of Na(+)/K(+)-ATPase regulates MAP kinase via Src kinase dependent reactions. Using of Annexin V-FITC apoptotic test to determine the cells with early apoptotic features allows to conclude that alpha3 isoform stimulates and alpha1 suppresses apoptotic process in cerebellum neurons. These data are the first demonstration showing participation of ouabain-resistant (alpha1) and ouabain-sensitive (alpha3) Na(+)/K(+)-ATPase isoforms in diverse signaling pathways in neuronal cells.
Asunto(s)
Neuronas/enzimología , Transducción de Señal , ATPasa Intercambiadora de Sodio-Potasio/metabolismo , Animales , Apoptosis , Cerebelo/citología , Proteína Quinasa 1 Activada por Mitógenos/metabolismo , Proteína Quinasa 3 Activada por Mitógenos/metabolismo , Ouabaína/farmacología , Fosfatidilinositol 3-Quinasas/metabolismo , Isoformas de Proteínas/antagonistas & inhibidores , Isoformas de Proteínas/metabolismo , Proteína Quinasa C/metabolismo , Ratas , ATPasa Intercambiadora de Sodio-Potasio/antagonistas & inhibidores , Familia-src Quinasas/metabolismoRESUMEN
Carnosine and related compounds were compared in terms of their abilities to decrease the levels of reactive oxygen species (ROS) in suspensions of isolated neurons activated by N-methyl-D-aspartic acid (NMDA) using both stationary fluorescence measurements and flow cytometry. Carnosine was found to suppress the fluorescent signal induced by ROS production and decreased the proportion of highly fluorescent neurons, while histidine showed opposite effects. N-Acetylated derivatives of both carnosine and histidine demonstrated weak (statistically indistinguishable) suppressive effects on the ROS signal. N-Methylated derivatives of carnosine suppressed intracellular ROS generation to the same extent as carnosine. This rank of effectiveness is distinct from that previously obtained for the anti-radical ability of CRCs (anserine>carnosine>ophidine). These differences suggest that the similar ability of carnosine and its N-methylated derivatives to protect neuronal cells against the excitotoxic effect of NMDA is not solely related to the antioxidant properties of these compounds.
Asunto(s)
Anserina/farmacología , Carnosina/análogos & derivados , Carnosina/farmacología , Neuronas/metabolismo , Especies Reactivas de Oxígeno/antagonistas & inhibidores , Animales , Cerebelo/citología , Cerebelo/metabolismo , Citometría de Flujo , N-Metilaspartato/farmacología , Neuronas/efectos de los fármacos , Ratas , Ratas Wistar , Espectrometría de FluorescenciaRESUMEN
Two glutamate receptor agonists, NMDA (N-methyl-D-aspartic acid) and ACPD (cis-(1S/3R)-1-aminocyclopentane- 1,3-dicarboxylic acid), induce the reactive oxygen species (ROS) production in rat cerebellum granule cells, whereas the third one, 3-HPG (3-hydroxyphenylglycine), decreases this parameter. The simultaneous presence of 3-HPG, together with NMDA or ACPD, prevents the generation of ROS by neuronal cells. A similar effect of these ligands on Na+/K+-ATPase can be demonstrated: NMDA and ACPD inhibited the enzyme activity, but 3-HPG activated Na+/K+-ATPase and prevented its inhibition by NMDA or ACPD. In terms of current classification, NMDA is an agonist of ionotropic glutamate receptors of the so-called NMDA class, whereas ACPD and 3-HPG belong to metabotropic agonists, the former primarily being an activator of metabotropic glutamate receptors (mGluRs) of groups 2 and 3, and the latter, that of mGluRs of groups 1 and 5. Thus, the data presented illustrate the existence of diverse mechanisms of the cross talk between Na+/K+-ATPase and different glutamate receptors, as well as that between glutamate receptors of different classes.