RESUMEN
Sudden unexpected death in infancy is defined as sudden unexpected death occurring before 12 months of age. The common causes of sudden unexpected death in infancy are infection, cardiovascular anomaly, child abuse, and metabolic disorders. However, the many potential inherited metabolic disorders are difficult to diagnose at autopsy and may therefore be underdiagnosed as a cause of sudden unexpected death in infancy. In the present study we retrospectively reviewed 30 Japanese sudden unexpected death in infancy cases encountered between 2006 and 2009 at our institute. With postmortem blood acylcarnitine analysis and histological examination of the liver, we found two cases of long-chain fatty acid oxidation defects. Molecular analysis revealed that the one patient had a compound heterozygote for a novel mutation (p.L644S) and a disease-causing mutation (p.F383Y) in the carnitine palmitoyltransferase 2 gene. Furthermore, retrospective acylcarnitine analysis of the newborn screening card of this patient was consistent with carnitine palmitoyltransferase II deficiency. Metabolic autopsy and expanded newborn screening would be helpful for forensic scientists and pediatricians to diagnose fatty acid oxidation disorders and prevent sudden unexpected death in infancy.
Asunto(s)
Tamizaje Neonatal/métodos , Muerte Súbita del Lactante/epidemiología , Acil-CoA Deshidrogenasa de Cadena Larga/genética , Carnitina/análogos & derivados , Carnitina/sangre , Carnitina Aciltransferasas/genética , Carnitina O-Palmitoiltransferasa/deficiencia , Carnitina O-Palmitoiltransferasa/genética , Estudios de Casos y Controles , Femenino , Haplotipos , Humanos , Lactante , Recién Nacido , Japón , Hígado/patología , Masculino , Proteína Trifuncional Mitocondrial , Complejos Multienzimáticos/genética , Mutación Missense , Estudios Retrospectivos , Muerte Súbita del Lactante/etiologíaRESUMEN
Methamphetamine induces several cardiac dysfunctions, which leads to arrhythmia, cardiac failure and sudden cardiac death. Although these cardiac alterations elicited by methamphetamine were thought to be due to an indirect action of methamphetamine, namely, an excessive catecholamine release from synaptic terminals, while it seems likely that methamphetamine directly modulates the functioning of cardiomyocytes independent of neurotransmitters. However, the direct effects of methamphetamine on cardiomyocytes are still not clear. We show that methamphetamine directly accelerates the beating rate and alters Ca(2+) oscillation pattern in cultured neonatal rat cardiomyocytes. Adrenergic receptor antagonists did not block the methamphetamine-induced alterations in cardiomyocytes. Treatment with a ryanodine receptor type 2 inhibitor and a sarcoplasmic reticulum Ca(2+)-ATPase inhibitor did not affect these responses, either. In contrast, the L-type Ca(2+) channel inhibitor nifedipine eradicated these responses. Furthermore, methamphetamine elevated the internal free Ca(2+) concentration in HEK-293T cells stably transfected with the L-type Ca(2+) channel alpha1C subunit. In neonatal rat cardiomyocytes, methamphetamine accelerates beating rate and alters Ca(2+) oscillation pattern by increasing Ca(2+) entry via the L-type Ca(2+) channels independent of any neurotransmitters.
Asunto(s)
Canales de Calcio Tipo L/efectos de los fármacos , Señalización del Calcio/efectos de los fármacos , Estimulantes del Sistema Nervioso Central/farmacología , Frecuencia Cardíaca/efectos de los fármacos , Metanfetamina/farmacología , Miocitos Cardíacos/efectos de los fármacos , Animales , Línea Celular , Humanos , Miocitos Cardíacos/metabolismo , Miocitos Cardíacos/fisiología , Ratas , Receptores Adrenérgicos/metabolismoRESUMEN
BACKGROUND: It is unknown whether AQP5 and lipid rafts are released into human unstimulated (resting) saliva and saliva in response to secretagogues. METHODS: In order to quantitate the salivary concentration of AQP5, we produced a polyclonal antibody for human AQP5 and developed an enzyme-like immunosorbent assay (ELISA). RESULTS: AQP5 and lipid rafts were identified in human resting saliva. The amount of AQP5 in resting saliva showed a diurnal variation with high levels during waking hours, and an age-related decrease in AQP5 was coincident with the volume of resting saliva. Cevimeline, a muscarinic acetylcholine receptor (mAChR) agonist, induced the release of AQP5 with lipid rafts, amylase, mucin, and lysozyme. Changes in saliva AQP5 levels after cevimeline administration occurred simultaneously with changes in saliva flow rates. Confocal microscopy revealed that AQP5 was located in the apical plasma membrane and showed a diffuse pattern in parotid glands under resting conditions. Following cevimeline administration, AQP5 was predominantly associated with the APM and was localized in the lumen. GENERAL SIGNIFICANCE: AQP5 and lipid rafts were released with salivary proteins from human salivary glands by the stimulation of M3 mAChRs, and that changes in saliva AQP5 levels can be used as an indicator of salivary flow rate and also as a useful index of M3 mAChR agonist's action on human salivary glands.
Asunto(s)
Acuaporina 5/metabolismo , Microdominios de Membrana/fisiología , Quinuclidinas/farmacología , Receptor Muscarínico M3/agonistas , Saliva/metabolismo , Glándulas Salivales/fisiología , Tiofenos/farmacología , Adulto , Factores de Edad , Anciano , Anciano de 80 o más Años , Amilasas/metabolismo , Animales , Ritmo Circadiano , Femenino , Técnica del Anticuerpo Fluorescente , Humanos , Masculino , Microscopía Confocal , Persona de Mediana Edad , Glándula Parótida/efectos de los fármacos , Glándula Parótida/metabolismo , Glándula Parótida/ultraestructura , Ratas , Ratas Wistar , Saliva/efectos de los fármacos , Glándulas Salivales/efectos de los fármacos , Glándulas Salivales/ultraestructura , Sueño , Vigilia , Adulto JovenRESUMEN
Synaptic activity induces changes in the number of dendritic spines. Here, we report a pathway of regulated endocytosis triggered by arcadlin, a protocadherin induced by electroconvulsive and other excitatory stimuli in hippocampal neurons. The homophilic binding of extracellular arcadlin domains activates TAO2beta, a splice variant of the thousand and one amino acid protein kinase 2, cloned here by virtue of its binding to the arcadlin intracellular domain. TAO2beta is a MAPKKK that activates the MEK3 MAPKK, which phosphorylates the p38 MAPK. Activation of p38 feeds-back on TAO2beta, phosphorylating a key serine required for triggering endocytosis of N-cadherin at the synapse. Arcadlin knockout increases the number of dendritic spines, and the phenotype is rescued by siRNA knockdown of N-cadherin. This pathway of regulated endocytosis of N-cadherin via protocadherin/TAO2beta/MEK3/p38 provides a molecular mechanism for transducing neuronal activity into changes in synaptic morphologies.
Asunto(s)
Cadherinas/metabolismo , Espinas Dendríticas/metabolismo , Quinasas Quinasa Quinasa PAM/metabolismo , Transmisión Sináptica/fisiología , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismo , Animales , Animales Recién Nacidos , Células COS , Células Cultivadas , Chlorocebus aethiops , Espinas Dendríticas/ultraestructura , Estimulación Eléctrica , Endocitosis/fisiología , Activación Enzimática/fisiología , Hipocampo/metabolismo , Hipocampo/ultraestructura , Humanos , Sistema de Señalización de MAP Quinasas/fisiología , Ratones , Datos de Secuencia Molecular , Plasticidad Neuronal/fisiología , Proteínas Serina-Treonina Quinasas , Estructura Terciaria de Proteína/genética , Protocadherinas , Ratas , Sinapsis/metabolismo , Sinapsis/ultraestructuraRESUMEN
Two Puralpha-binding proteins (PurBPs) were found in nuclear extract from mouse brain during P4-P10 by the overlay assay. At P14, they were decreased significantly in nuclear extract and increased in the S3 fraction, indicating their dynamic translocation during development. Western blot analysis also demonstrated concomitant translocation of Puralpha with the PurBPs during P7-P14, when neuronal circuit proceeds. Immunocytochemical study with cultured hippocampal neurons from rat E18 confirmed that nuclear Puralpha was translocated to cytoplasm after plating for 7-14 days. These results suggest that spatiotemporal translocation of Puralpha with the PurBPs from nuclei to cytoplasm has a crucial role in neuronal development.
Asunto(s)
Núcleo Celular/metabolismo , Proteína de Unión a Elemento de Respuesta al AMP Cíclico/metabolismo , Citoplasma/metabolismo , Hipocampo/citología , Neuronas/metabolismo , Factores de Edad , Animales , Animales Recién Nacidos , Western Blotting/métodos , Células Cultivadas , Proteínas de Unión al ADN , Embrión de Mamíferos , Hipocampo/crecimiento & desarrollo , Inmunohistoquímica/métodos , Indoles/metabolismo , Proteínas del Tejido Nervioso , Ratas , Fracciones Subcelulares/metabolismo , Factores de Tiempo , Factores de TranscripciónRESUMEN
Neural activity induces the remodeling of pre- and postsynaptic membranes, which maintain their apposition through cell adhesion molecules. Among them, N-cadherin is redistributed, undergoes activity-dependent conformational changes, and is required for synaptic plasticity. Here, we show that depolarization induces the enlargement of the width of spine head, and that cadherin activity is essential for this synaptic rearrangement. Dendritic spines visualized with green fluorescent protein in hippocampal neurons showed an expansion by the activation of AMPA receptor, so that the synaptic apposition zone may be expanded. N-cadherin-venus fusion protein laterally dispersed along the expanding spine head. Overexpression of dominant-negative forms of N-cadherin resulted in the abrogation of the spine expansion. Inhibition of actin polymerization with cytochalasin D abolished the spine expansion. Together, our data suggest that cadherin-based adhesion machinery coupled with the actin-cytoskeleton is critical for the remodeling of synaptic apposition zone.
Asunto(s)
Cadherinas/metabolismo , Espinas Dendríticas/metabolismo , Hipocampo/metabolismo , Plasticidad Neuronal/fisiología , Transmisión Sináptica/fisiología , Actinas/antagonistas & inhibidores , Actinas/biosíntesis , Potenciales de Acción/fisiología , Animales , Cadherinas/genética , Células Cultivadas , Citocalasina D/farmacología , Espinas Dendríticas/ultraestructura , Potenciales Postsinápticos Excitadores/fisiología , Proteínas Fluorescentes Verdes/metabolismo , Hipocampo/citología , Microscopía Confocal , Microscopía Fluorescente , Proteínas del Tejido Nervioso/metabolismo , Ratas , Receptores AMPA/metabolismo , Proteínas Recombinantes de Fusión , Membranas Sinápticas/metabolismoRESUMEN
Arc, activity-regulated cytoskeleton-associated gene, is an immediate early gene, and its expression is regulated by a variety of stimuli, such as electric stimulation and methamphetamine. The function of Arc, however, is unknown. To explore this function, we carried out expression experiments by transfecting green fluorescent protein (GFP)-Arc constructs or by using a protein transduction system in hippocampal cultured neurons. We found that the overexpression of Arc as well as Arc induction by seizure in vivo decreased microtubule-associated protein 2 (MAP2) staining in the dendrites by immunocytochemistry, although MAP2 content was not changed on Western blot. Furthermore, Arc interacted with newly polymerized microtubules and MAP2, leading to blocking of the epitope of MAP2. The data suggest that Arc increased by synaptic activities would trigger dendritic remodeling by interacting with cytoskeletal proteins.