RESUMEN
BACKGROUND: Amyotrophic lateral sclerosis (ALS) is relatively rare, yet the economic and social burden is substantial. Having accurate incidence and prevalence estimates would facilitate efficient allocation of healthcare resources. OBJECTIVE: To provide a comprehensive and critical review of the epidemiological literature on ALS. METHODS: MEDLINE and EMBASE (1995-2011) databases of population-based studies on ALS incidence and prevalence reporting quantitative data were analyzed. Data extracted included study location and time, design and data sources, case ascertainment methods and incidence and/or prevalence rates. Medians and interquartile ranges (IQRs) were calculated, and ALS case estimates were derived using 2010 population estimates. RESULTS: In all, 37 articles met the inclusion criteria. In Europe, the median incidence rate (/100,000 population) was 2.08 (IQR 1.47-2.43), corresponding to an estimated 15,355 (10,852-17,938) cases. Median prevalence (/100,000 population) was 5.40 (IQR 4.06-7.89), or 39,863 (29,971-58,244) prevalent cases. CONCLUSIONS: Disparity in rates among ALS incidence and prevalence studies may be due to differences in study design or true variations in population demographics such as age and geography, including environmental factors and genetic predisposition. Additional large-scale studies that use standardized case ascertainment methods are needed to more accurately assess the true global burden of ALS.
Asunto(s)
Esclerosis Amiotrófica Lateral/diagnóstico , Esclerosis Amiotrófica Lateral/epidemiología , Salud Global , Salud Global/tendencias , HumanosRESUMEN
TRAIL-induced apoptosis has been considered a promising therapeutic approach for tumors that are resistant to chemotherapy, which is usually mediated via mitochondrial apoptotic cascades. Recent studies have shown that in certain cancer cells, TRAIL-mediated apoptosis is also dependent on mitochondrial involvement, suggesting that similar mechanisms of resistance to chemotherapy might be implicated in the resistance of tumor cells to TRAIL. We have used TRAIL-resistant leukemic cells that are deficient in both Bax and Bak to determine the roles of these Bcl-2 members in TRAIL-mediated apoptosis. Exposure of these cells to TRAIL did not have an impact on cell viability, although it induced the processing of caspase-3 to its active p20 subunit. The activity of the p20 caspase-3 appeared to be inhibited as no autoprocessing of this p20 subunit or cleavage of known caspase-3 substrates were detected. Also, in the absence of Bax and Bak, no release of mitochondrial apoptogenic proteins was observed following TRAIL treatment. Adenoviral transduction of the Bax, but not the Bak gene, to the Bax/Bak-deficient leukemic cells rendered them TRAIL-sensitive as assessed by enhanced apoptotic death and caspase-3 processing. These findings demonstrate preferential utilization of Bax over Bak in leukemic cell response to specific apoptotic stimulation.
Asunto(s)
Apoptosis/efectos de los fármacos , Leucemia/metabolismo , Glicoproteínas de Membrana/farmacología , Proteínas de la Membrana/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2 , Proteínas Proto-Oncogénicas/metabolismo , Linfocitos T/metabolismo , Factor de Necrosis Tumoral alfa/farmacología , Adenoviridae/genética , Apoptosis/fisiología , Proteínas Reguladoras de la Apoptosis , Caspasa 3 , Caspasas/metabolismo , Activación Enzimática/efectos de los fármacos , Humanos , Leucemia/patología , Proteínas de la Membrana/genética , Mitocondrias/efectos de los fármacos , Mitocondrias/metabolismo , Proteínas Mitocondriales/metabolismo , Proteínas Proto-Oncogénicas/genética , Linfocitos T/efectos de los fármacos , Ligando Inductor de Apoptosis Relacionado con TNF , Proteína Destructora del Antagonista Homólogo bcl-2 , Proteína X Asociada a bcl-2RESUMEN
Granzyme B (GrB), a serine protease with substrate specificity similar to the caspase family, is a major component of granule-mediated cytotoxicity of T lymphocytes. Although GrB can directly activate caspases, it induces apoptosis predominantly via Bid cleavage, mitochondrial outer membrane permeabilization, and cytochrome c release. To study the molecular regulators for GrB-mediated mitochondrial apoptotic events, we used a CTL-free cytotoxicity system, wherein target cells are treated with purified GrB and replication-deficient adenovirus (Ad). We report here that the Bcl-2 proapoptotic family member, Bak, plays a dominant role in GrB-mediated mitochondrial apoptotic events. A variant of Jurkat cells, deficient in Bak expression, was resistant to GrB/Ad-mediated apoptosis, as determined by lack of membranous phosphatidylserine exposure, lack of DNA breaks, lack of mitochondrial outer membrane permeabilization, and unchanged expression of inner mitochondrial membrane cardiolipin. The resistance of Bak-deficient cells to GrB/Ad cytotoxicity was reversed by transduction of the Bak gene into these cells. The requirement for both Bid and Bak, was further demonstrated in a cell-free system using purified mitochondria and S-100 cytosol. Purified mitochondria from Bid knockout mice, but not from Bax knockout mice, failed to release cytochrome c in response to autologous S-100 and GrB. Also, Bak-deficient mitochondria did not release cytochrome c in response to GrB-treated cytosol unless recombinant Bak protein was added. These results are the first to report a role for Bak in GrB-mediated mitochondrial apoptosis. This study demonstrates that GrB-cleaved Bid, which differs in size and site of cleavage from caspase-8-cleaved Bid, utilizes Bak for cytochrome c release, and therefore, suggests that deficiency in Bak may serve as a mechanism of immune evasion for tumor or viral infected cells.
Asunto(s)
Apoptosis , Grupo Citocromo c/metabolismo , Proteínas de la Membrana/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Serina Endopeptidasas/metabolismo , Adenoviridae , Proteína Proapoptótica que Interacciona Mediante Dominios BH3 , Proteínas Portadoras/genética , Proteínas Portadoras/metabolismo , Vectores Genéticos , Granzimas , Humanos , Células Jurkat , Mitocondrias/metabolismo , Proteínas Proto-Oncogénicas/metabolismo , Proteína Destructora del Antagonista Homólogo bcl-2 , Proteína X Asociada a bcl-2RESUMEN
In the present study a clonal Jurkat cell line deficient in expression of Bak was used to analyze the role of Bak in cytochrome c release from mitochondria. The Bak-deficient T leukemic cells were resistant to apoptosis induced by UV, staurosporin, VP-16, bleomycin, or cisplatin. In contrast to wild type Jurkat cells, these Bak-deficient cells did not respond to UV or treatment with these anticancer drugs by membranous phosphatidylserine exposure, DNA breaks, activation of caspases, or release of mitochondrial cytochrome c. The block in the apoptotic cascade was in the mitochondrial mechanism for cytochrome c release because purified mitochondria from Bak-deficient cells failed to release cytochrome c or apoptosis-inducing factor in response to recombinant Bax or truncated Bid. The resistance of Bak-deficient cells to VP-16 was reversed by transduction of the Bak gene into these cells. Also, the cytochrome c releasing capability of the Bak-deficient mitochondria was restored by insertion of recombinant Bak protein into purified mitochondria. Following mitochondrial localization, low dose recombinant Bak restored the mitochondrial release of cytochrome c in response to Bax; at increased doses it induced cytochrome c release itself. The function of Bak is independent of Bid and Bax because recombinant Bak induced cytochrome c release from mitochondria purified from Bax(-/-), Bid(-/-), or Bid(-/-) Bax(-/-) mice. Together, our findings suggest that Bak plays a key role in the apoptotic machinery of cytochrome c release and thus in the chemoresistance of human T leukemic cells.
Asunto(s)
Antineoplásicos/farmacología , Apoptosis , Grupo Citocromo c/metabolismo , Proteínas de la Membrana/fisiología , Mitocondrias/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2 , Linfocitos T/metabolismo , Adenoviridae/genética , Antimetabolitos Antineoplásicos/farmacología , Antineoplásicos Fitogénicos/farmacología , Proteína Proapoptótica que Interacciona Mediante Dominios BH3 , Bleomicina/farmacología , Western Blotting , Proteínas Portadoras/metabolismo , Caspasas/metabolismo , Cisplatino/farmacología , Daño del ADN , Inhibidores Enzimáticos/farmacología , Etopósido/farmacología , Citometría de Flujo , Humanos , Células Jurkat , Inhibidores de la Síntesis del Ácido Nucleico/farmacología , Plásmidos/metabolismo , Estructura Terciaria de Proteína , Proteínas Proto-Oncogénicas/metabolismo , Fármacos Sensibilizantes a Radiaciones/farmacología , Proteínas Recombinantes/metabolismo , Estaurosporina/farmacología , Factores de Tiempo , Transducción Genética , Rayos Ultravioleta , Proteína Destructora del Antagonista Homólogo bcl-2 , Proteína X Asociada a bcl-2RESUMEN
Jurkat leukemic T cells are highly sensitive to the extrinsic pathways of apoptosis induced via the death receptor Fas or tumor necrosis factor-related apoptosis-inducing ligand as well as to the intrinsic/mitochondrial pathways of death induced by VP-16 or staurosporin. We report here that clonal Jurkat cell lines selected for resistance to Fas-induced apoptosis were cross-resistant to VP-16 or staurosporin. Each of the apoptotic pathways was blocked at an apical phase, where common regulators of apoptosis have not yet been defined. The Fas pathway was blocked at the level of caspase-8, whereas the intrinsic pathway was blocked at the mitochondria. No processing or activity of caspases was detected in resistant cells in response to either Fas-cross-linking or VP-16 treatment. Also, no apoptosis-associated alterations in the mitochondrial inner membrane, outer membrane, or matrix were detected in resistant Jurkat cells treated with VP-16. Thus, no changes in permeability transition, loss in inner membrane cardiolipin, generation of reactive oxygen species, or release of cytochrome c were observed in resistant cells treated with VP-16. Further, unlike purified mitochondria from wild type cells, those obtained from resistant cells did not release cytochrome c or apoptosis-inducing factor in response to recombinant Bax or truncated Bid. These results identify a defect in mitochondria ability to release intermembrane proteins in response to Bid or Bax as a mechanism of resistance to chemotherapeuetic drugs. Further, the selection of VP-16-resistant mitochondria via elimination of Fas-susceptible cells may suggest the existence of a shared regulatory component between the extrinsic and intrinsic pathways of apoptosis.
Asunto(s)
Apoptosis/fisiología , Mitocondrias/fisiología , Subgrupos de Linfocitos T/citología , Linfocitos T/citología , Receptor fas/inmunología , Antineoplásicos/farmacología , Caspasas/metabolismo , Activación Enzimática , Etopósido/farmacología , Humanos , Transducción de Señal/fisiología , Subgrupos de Linfocitos T/efectos de los fármacos , Subgrupos de Linfocitos T/fisiología , Linfocitos T/efectos de los fármacos , Linfocitos T/fisiología , Células Tumorales Cultivadas , Receptor fas/farmacologíaRESUMEN
Seprase is a homodimeric 170-kDa integral membrane gelatinase that is related to the ectoenzyme dipeptidyl peptidase IV. We have identified an alternatively spliced seprase messenger from the human melanoma cell line LOX that encodes a novel truncated isoform, seprase-s. The splice variant mRNA is generated by an out-of-frame deletion of a 1223-base pair exonic region that encodes part of the cytoplasmic tail, transmembrane, and the membrane proximal-central regions of the extracellular domain (Val(5) through Ser(412)) of the seprase 97-kDa subunit (seprase-l). The seprase-s mRNA has an elongated 5' leader (548 nucleotides) that harbors at least two upstream open reading frames that inhibit seprase-s expression from a downstream major open reading frame. Deletion mutagenesis of the wild type splice variant cDNA confirms that initiation of the seprase-s coding sequence begins with an ATG codon that corresponds to Met(522) of seprase-l. The seprase-s open reading frame encodes a 239-amino acid polypeptide with an M(r) approximately 27,000 that precisely overlaps the carboxyl-terminal catalytic region of seprase-l.
Asunto(s)
Empalme Alternativo , Gelatinasas/genética , Proteínas de la Membrana , Isoformas de Proteínas/genética , ARN Mensajero/genética , Serina Endopeptidasas , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Células COS , Clonación Molecular , ADN Complementario , Endopeptidasas , Humanos , Datos de Secuencia Molecular , Mutagénesis , Sistemas de Lectura Abierta , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Eliminación de Secuencia , Células Tumorales CultivadasAsunto(s)
Accidentes de Tránsito/legislación & jurisprudencia , Accidentes de Tránsito/mortalidad , Equipo Infantil , Heridas y Lesiones/prevención & control , Preescolar , Educación en Salud , Humanos , Lactante , Equipo Infantil/estadística & datos numéricos , Recién Nacido , Kentucky/epidemiología , Heridas y Lesiones/mortalidadAsunto(s)
Enfermedades de los Trabajadores Agrícolas/etiología , Agricultura/instrumentación , Intoxicación por Monóxido de Carbono/etiología , Emisiones de Vehículos/efectos adversos , Adulto , Enfermedades de los Trabajadores Agrícolas/sangre , Enfermedades de los Trabajadores Agrícolas/terapia , Intoxicación por Monóxido de Carbono/sangre , Intoxicación por Monóxido de Carbono/terapia , Carboxihemoglobina/análisis , Tratamiento de Urgencia , Femenino , Humanos , Terapia por Inhalación de OxígenoRESUMEN
OBJECTIVES: To analyze data from motor vehicle crashes (MVCs) involving teenaged drivers in Kentucky for 1994, and derive cost estimates of these crashes. METHODS: Crash data were obtained from the Kentucky Traffic Accident Facts 1994 Report and the Kentucky Accident Reporting System. The National Highway Traffic Safety Administration's Crash-Cost program was used to generate cost estimates for Kentucky data. RESULTS: Teenaged drivers had significantly higher MVC fatal and non-fatal injury rates than did adult drivers. The deaths rates were 43.6 and 19.0 per 100,000 for teens and adult drivers, respectively. Odds ratios (ORs) were calculated to estimate the relative risk for (1) involvement in an MVC, (2) fatal or incapacitating injury, and (3) fatal injury for teenaged compared with adult drivers. The crude ORs were statistically significant at each age. Cost estimates were calculated on a per person/vehicle basis. A single fatal injury was $642,700. A critical injury was $563,000. In general, unit costs rose with increasing levels of injury severity. For the total number of fatal injuries, costs exceeded $91 million. For non-fatal injuries and property damage only crashes, total costs were $318 million. Overall, the total cost estimate for MVCs involving teenaged drivers was nearly $410 million. CONCLUSIONS: Strategies aimed to reduce the number of MVCs attributed to teenaged drivers should reduce both the number and costs of crash related deaths and injuries. Graduated driver licensing (GDL) systems are one plausible approach toward achieving this goal. By recently enacting a GDL system in Kentucky, it is anticipated that many lives and dollars will be saved.
Asunto(s)
Accidentes de Tránsito/economía , Accidentes de Tránsito/mortalidad , Heridas y Lesiones/economía , Accidentes de Tránsito/prevención & control , Adolescente , Adulto , Distribución por Edad , Causas de Muerte , Costos y Análisis de Costo , Femenino , Costos de la Atención en Salud , Humanos , Puntaje de Gravedad del Traumatismo , Kentucky/epidemiología , Masculino , Oportunidad Relativa , Factores de Riesgo , Tasa de Supervivencia , Heridas y Lesiones/epidemiología , Heridas y Lesiones/prevención & controlRESUMEN
Seprase is a homodimeric 170 kDa integral membrane gelatinase whose expression correlates with the invasiveness of the human melanoma cell line LOX. Here, we report the molecular cloning of a cDNA that encodes the 97 kDa subunit of seprase. Its deduced amino acid sequence predicts a type II integral membrane protein with a cytoplasmic tail of 6 amino acids, followed by a transmembrane domain of 20 amino acids and an extracellular domain of 734 amino acids. The carboxyl terminus contains a putative catalytic region (approximately 200 amino acids) which is homologous (68% identity) to that of the nonclassical serine protease dipeptidyl peptidase IV (DPPIV). The conserved serine protease motif G-X-S-X-G is present as G-W-S-Y-G. However, sequence analysis of seprase cDNA from LOX and other cell lines strongly suggests that seprase and human fibroblast activation protein alpha (FAP alpha) are products of the same gene. We propose that seprase/FAP alpha and DPPIV represent a new subfamily of serine integral membrane proteases (SIMP).
Asunto(s)
Antígenos de Neoplasias , Biomarcadores de Tumor , Gelatinasas/genética , Melanoma/enzimología , Proteínas de la Membrana , Serina Endopeptidasas , Secuencia de Aminoácidos , Secuencia de Bases , Línea Celular , Clonación Molecular , ADN Complementario/química , ADN Complementario/genética , Endopeptidasas , Gelatinasas/química , Sustancias de Crecimiento/genética , Humanos , Datos de Secuencia Molecular , Invasividad Neoplásica , Alineación de SecuenciaRESUMEN
The 170-kDa membrane-bound gelatinase, seprase, is a cell surface protease, the expression of which correlates with the invasive phenotype of human melanoma and carcinoma cells. We have isolated seprase from cell membranes and shed vesicles of LOX human melanoma cells. The active enzyme is a dimer of N-glycosylated 97-kDa subunits. Sequence analysis of three internal proteolytic fragments of the 97-kDa polypeptide revealed up to 87.5% identity to the 95-kDa fibroblast activation protein alpha (FAPalpha), the function of which is unknown. Thus, we used reverse transcription-polymerase chain reaction to generate a 2.4-kilobase cDNA from LOX mRNA with FAPalpha primers. COS-7 cells transfected with this cDNA expressed a 170-kDa gelatinase that is recognized by monoclonal antibodies directed against seprase. Sequence analysis also showed similarities to the 110-kDa subunit of dipeptidyl peptidase IV (DPPIV). Like DPPIV, the gelatinase activity of seprase was completely blocked by serine-protease inhibitors, including diisopropyl fluorophosphate. Seprase could be affinity-labeled by [3H]diisopropyl fluorophosphate, but the proteolytically inactive 97-kDa subunit could not, confirming the existence of a serine protease active site on the dimeric form. Proteolytic activity is lost upon dissociation into its 97-kDa subunit following treatment with acid, heat, or cysteine and histidine-modifying agents. We conclude that seprase, FAPalpha, and DPPIV are related serine integral membrane proteases and that seprase is similar to DPPIV, the proteolytic activities of which are dependent upon subunit association.
Asunto(s)
Gelatinasas/metabolismo , Melanoma/enzimología , Proteínas de la Membrana/metabolismo , Serina Endopeptidasas/metabolismo , Secuencia de Aminoácidos , Sitios de Unión , Catálisis , Gelatinasas/química , Humanos , Proteínas de la Membrana/química , Datos de Secuencia Molecular , Homología de Secuencia de Aminoácido , Serina Endopeptidasas/química , Células Tumorales CultivadasRESUMEN
Mice with a nontargeted transgene insertion at the motor endplate disease (med) locus (med(tg)) contain a deletion of a novel gene encoding a neuronal voltage-gated sodium channel, designated Scn8a. We characterized severe skeletal muscle atrophy beginning by Postnatal Day 10 (P10) and death by P20 in the med(tg) mouse. Denervation was functional, rather than structural, since the Scn8a mutation was not accompanied by retraction of neuromuscular contacts, motoneuron death, or decreased motoneuron soma diameter. Although pathology consistent with denervation was seen in both hindlimb and forelimb musculature, the postnatal maturation of the extraocular muscles was not altered. The onset of paralysis is likely coincident with the time that the Scn8a sodium channel normally assumes a critical role in the initiation and/or propagation of action potentials in spinal motoneurons. By contrast, the lack of consequences for extraocular muscle suggests that the Scn8a voltage-gated sodium channel may be of relatively minor importance for oculomotor motoneurons.
Asunto(s)
Neuronas Motoras/fisiología , Nervio Oculomotor/crecimiento & desarrollo , Canales de Sodio/fisiología , Médula Espinal/crecimiento & desarrollo , Animales , Ratones , Ratones Transgénicos , Músculo Esquelético/ultraestructuraRESUMEN
The role of eosinophils in inflammation and their mode of activation is not well understood. Eosinophil accumulation and subsequent expression of cytokines at the site of inflammation may play a role in exacerbation of inflammatory responses. In the present study, we have examined the role of TNF-alpha in eosinophil activation and chemokine production using a human leukaemic eosinophil cell line, EOL-1. Initial studies demonstrated that TNF-alpha induced the upregulation of IL-8 and MCP-1 mRNA and protein. Kinetic studies indicated production of chemokines, IL-8 and MCP-1, as early as 4 h post-activation, with peak levels of chemokine produced at 8 h, and decreasing by 24 h post-TNF-alpha activation. When IL-10, a suppressive cytokine, was incubated with TNF-alpha and EOL-1 cells, no effect was observed on IL-8 and MCP-1 production. However, dexamethasone, a glucocorticoid, demonstrated potent inhibitory effects on the EOL-1-derived chemokines. These studies indicate that eosinophils may be a significant source of chemokines capable of participating in, and maintaining, leukocyte recruitment during inflammatory responses, such as asthma.
RESUMEN
The spinal nucleus of the bulbocavernosus (SNB) and the dorsolateral nucleus (DLN) are sexually dimorphic motor nuclei in the rat lumbar spinal cord. During postnatal development, SNB and DLN motoneurons grow substantially in measures of soma size, dendritic length, and radial dendritic extent. SNB motoneurons exhibit a biphasic pattern of dendritic growth, where there is an initial period of exuberant growth followed by a period of retraction to mature lengths by 7 weeks. In this experiment, we examined whether primary afferent input to the SNB nucleus was necessary for the normal postnatal growth of SNB motoneurons. We partially deafferented the SNB via unilateral dorsal rhizotomy of lumbosacral dorsal roots in male rats at 1 week of age. Using cholera toxin horseradish peroxidase (BHRP) to visualize SNB motoneurons, we examined SNB motoneuron morphology at 4 and 7 weeks of age. SNB motoneurons in rhizotomized males developed normally; measures of dendritic length in rhizotomized males were typically exuberant at 4 weeks of age, and declined significantly to mature lengths by 7 weeks of age. In addition, dorsal rhizotomy did not alter the development of SNB motoneuron soma size or radial dendritic extent. These results are discussed in reference to sensorimotor connections in the SNB, the extent of the deafferentation, and dendrodendritic interactions.
Asunto(s)
Ganglios Espinales/fisiología , Neuronas Motoras/fisiología , Neuronas Aferentes/fisiología , Rizotomía , Animales , Toxina del Cólera , Dendritas/ultraestructura , Ganglios Espinales/anatomía & histología , Ganglios Espinales/crecimiento & desarrollo , Histocitoquímica , Peroxidasa de Rábano Silvestre , Masculino , Neuronas Motoras/ultraestructura , Neuronas Aferentes/ultraestructura , Ratas , Ratas Sprague-DawleyRESUMEN
The lumbar spinal cord of the rat contains two sexually dimorphic motor nuclei, the spinal nucleus of the bulbocavernosus (SNB) and the dorsolateral nucleus (DLN). Postnatally, SNB and DLN motoneurons grow substantially and reach their adult morphology by 7 weeks of age. The masculinization of SNB and DLN motoneuron dendrites depends upon steroid hormones. After early castration, the growth of SNB and DLN dendrites is markedly attenuated, but testosterone replacement restores this growth. In the SNB, initial dendritic growth is also supported in castrates treated with estrogen. By using castration and hormone replacement techniques, we examined the development of DLN motoneuron morphology in estrogen-treated castrated rats to determine if estrogen also supports the growth of DLN motoneurons. In addition, given that dorsal root ganglia may be a site of estrogen action, we tested the hypothesis that estrogen acts at primary afferents to support DLN dendritic growth. Thus, we attempted to block the potential trophic effect of estrogen by performing unilateral dorsal rhizotomies in estrogen-treated castrates. DLN motoneuron morphology was analyzed at 4 and 7 weeks of age by using cholera toxin horseradish peroxidase (BHRP) histochemistry. As found for SNB motoneurons, estrogen treatment transiently supported development. DLN motoneurons in estrogen-treated castrates developed normally through 4 weeks of age, but by 7 weeks, DLN motoneuron morphology in estrogen-treated castrates was no longer different from that in oil-treated castrates. Moreover, deafferentation via unilateral dorsal rhizotomy did not inhibit estrogen's ability to masculinize the early development of DLN motoneurons. Thus, the trophic effect of estrogen did not appear to act via the dorsal root ganglia to support the early postnatal development of DLN motoneurons.
Asunto(s)
Estrógenos/farmacología , Ganglios Espinales/crecimiento & desarrollo , Neuronas Motoras/fisiología , Neuronas Aferentes/fisiología , Rizotomía , Animales , Toxina del Cólera , Dendritas/ultraestructura , Ganglios Espinales/citología , Ganglios Espinales/efectos de los fármacos , Histocitoquímica , Masculino , Neuronas Motoras/ultraestructura , Neuronas Aferentes/ultraestructura , Orquiectomía , Monoéster Fosfórico Hidrolasas , Ratas , Ratas Sprague-DawleyRESUMEN
The rat lumbar spinal cord contains a sexually dimorphic motor nucleus, the spinal nucleus of the bulbocavernosus (SNB), whose motoneurons innervate perineal muscles involved in copulatory reflexes. Dendritic development of SNB motoneurons is biphasic and androgen dependent. During the first 4 postnatal weeks, SNB dendrites grow exuberantly, and subsequently retract to mature lengths by 7 weeks of age. After early postnatal castration, SNB dendrites fail to grow, and testosterone replacement restores this growth. In other systems, testosterone and its metabolites, dihydrotestosterone and estrogen, are important for somatic and neural sexual differentiation. The purpose of the present study was to examine the effects of castration and dihydrotestosterone or estrogen replacement on the growth of SNB motoneuron somata and dendritic arbors. Male rat pups were castrated on postnatal (P) day 7 and treated daily with either dihydrotestosterone propionate (DHTP; 2 mg) or estradiol benzoate (EB; 100 micrograms) until P28 or P49. By using cholera toxin horseradish peroxidase (BHRP) histochemistry, the soma size, dendritic length, dendritic extent, and arbor area of BHRP-labeled SNB motoneurons were measured and analyzed. Both DHTP and EB treatment supported the initial exuberant growth of SNB dendrites through P28, but EB treatment was ineffective in maintaining mature, adult lengths at P49. The possible sites of hormone action and functional implications of these hormonal treatments are discussed.
Asunto(s)
Dihidrotestosterona/análogos & derivados , Estradiol/farmacología , Neuronas Motoras/fisiología , Caracteres Sexuales , Médula Espinal/crecimiento & desarrollo , Animales , Castración , Senescencia Celular/efectos de los fármacos , Toxina del Cólera , Dendritas/ultraestructura , Dihidrotestosterona/farmacología , Femenino , Peroxidasa de Rábano Silvestre , Masculino , Neuronas Motoras/efectos de los fármacos , Ratas , Médula Espinal/citología , Médula Espinal/efectos de los fármacosRESUMEN
The rat lumbar spinal cord contains two sexually dimorphic motor nuclei, the spinal nucleus of the bulbocavernosus (SNB), and the dorsolateral nucleus (DLN). These motor nuclei innervate anatomically distinct perineal muscles that are involved in functionally distinct copulatory reflexes. The motoneurons in the SNB and DLN have different dendritic morphologies. The dendrites of motoneurons in the medially positioned SNB have a radial, overlapping arrangement, whereas the dendrites of the laterally positioned DLN have a bipolar and strictly unilateral organization. During development, SNB motoneuron dendrites grow exuberantly and then retract to their mature lengths. In this experiment we determined whether the adult difference in SNB and DLN motoneuron morphology was reflected in different patterns of dendritic growth during normal development. Furthermore, the development of both these nuclei is under androgenic control. In the absence of androgens, SNB dendrites fail to grow; testosterone replacement supports normal dendritic growth. Thus, we also examined the development of DLN dendrites for similar evidence of androgenic regulation. By using cholera toxin-horseradish peroxidase (BHRP) to label motoneurons retrogradely, we measured the morphology of DLN motoneurons in normal males, and in castrates treated with testosterone or oil/blank implants at postnatal day (P) 7, P28, P49, and P70. Our results demonstrate that in contrast to the biphasic pattern of dendritic development in the SNB, dendritic growth in the DLN was monotonic; the dendritic length of motoneurons increased more than 500% between P7 and P70. However, as in the SNB, development of DLN motoneuron morphology is androgen-dependent. In castrates treated with oil/blank implants, DLN somal and dendritic growth were greatly attenuated compared to those of normal or testosterone-treated males. Thus, while androgens are clearly necessary for the growth of motoneurons in both the SNB and DLN, their different developmental patterns suggest that other factors must be involved in regulating this growth.
Asunto(s)
Andrógenos/fisiología , Neuronas Motoras/ultraestructura , Médula Espinal/citología , Animales , Transporte Axonal , Tamaño de la Célula , Toxina del Cólera , Copulación/fisiología , Dendritas/ultraestructura , Peroxidasa de Rábano Silvestre , Masculino , Neuronas Motoras/efectos de los fármacos , Neuronas Motoras/fisiología , Orquiectomía , Ratas , Ratas Sprague-Dawley , Reflejo/fisiología , Caracteres Sexuales , Médula Espinal/efectos de los fármacos , Médula Espinal/crecimiento & desarrollo , Testosterona/farmacologíaRESUMEN
During normal development, motoneuron dendrites in the spinal nucleus of the bulbocavernosus (SNB) grow exuberantly to almost twice their adult length and then retract. In this study, we retrogradely labeled SNB motoneurons with cholera toxin B-conjugated horseradish peroxidase (BHRP) to examine the maturation of SNB dendritic arbors in more detail, particularly with regard to its spatial distribution and reorganization. The number and orientation of SNB motoneuron primary processes did not change over the first ten weeks of life. In contrast, total dendritic length, radial extent and arbor area increased significantly through the first four postnatal weeks and declined thereafter. The declines in length and extent were restricted to particular portions of the arbor, specifically the dorsal, ipsi- and contralateral projections. Estimates of the degree of overlap between the dendritic arbors from both sides of the SNB reflected these changes, with overlap initially increasing and then decreasing as the SNB established its adult dendritic morphology. To determine if dendritic interactions facilitated by this arbor overlap might be involved in regulating the normal retraction of SNB dendrites, we reduced SNB motoneuron numbers unilaterally by target muscle removal on the day of birth. Somal size, number and orientation of primary processes developed normally in unilateral muscle-extirpated animals. The dendritic morphology of surviving SNB motoneurons in unilateral muscle extirpated males was altered, with significant increases in dendritic length, extent and arbor area relative to those of normal males. These results indicate that substantial changes in dendritic organization of SNB motoneurons occur in normal development and may be influenced by interactions between dendrites from the two halves of the SNB.
Asunto(s)
Dendritas/ultraestructura , Neuronas Motoras/ultraestructura , Médula Espinal/crecimiento & desarrollo , Animales , Toxina del Cólera , Histocitoquímica , Peroxidasa de Rábano Silvestre , Masculino , Desarrollo de Músculos , Músculos/inervación , Músculos/fisiología , Ratas , Ratas Sprague-Dawley , Médula Espinal/anatomía & histología , Médula Espinal/citologíaRESUMEN
The spinal nucleus of the bulbocavernosus (SNB) is a sexually dimorphic motor nucleus in the rat lumbar spinal cord. SNB motoneurons and their perineal target muscles are present in adult males, but reduced or absent in adult females. This dimorphism is due to the presence of androgens during development. Perinatal treatment of females with testosterone (T), or a combination of dihydrotestosterone (DHT) and estrogen (E+D females) from embryonic (E) day 16 through postnatal (P) day 5, results in a masculine number of SNB motoneurons and the retention of the target muscles. Perinatal treatment with estrogen alone does not masculinize the SNB; prenatal treatment with DHT alone from E17-E22 results in a feminine number of SNB motoneurons and a significantly altered motoneuron morphology and connectivity. To determine if masculinization of the SNB involves the interaction of estrogen and DHT or results from a longer exposure to DHT alone, the number, morphology, and connectivity of SNB motoneurons in females treated with DHT both pre- and post-natally (from E16-P5) were examined. At E22, DHTP (E16-P5) females have SNB motoneuron numbers identical to E+D and normal females, but far fewer than normal males, thus indicating that T is essential for prenatal masculinization. After E22, SNB motoneuron number declines precipitously in normal females but remains stable in DHTP (E16-P5) females and E+D females, which do not differ from normal males at P10. These results demonstrate that DHT can completely masculinize SNB motoneuron number without any synergistic actions with estrogen, and suggest that the development of SNB motoneuron number is strictly an androgen-mediated event. In adulthood, horseradish peroxidase histochemistry reveals that the connectivity, dendritic length, and soma size of SNB motoneurons in DHTP (E16-P5) females are identical to those of normal males but differ significantly from those of DHTP (E17-E22) females. These data suggest that the altered connectivity in DHTP (E17-E22) females is not simply a hormone-specific effect, but the result of a truncated hormone exposure. Thus, DHT can fully masculinize SNB morphology and connectivity if given during the appropriate period of development. It is suggested that while T may be required to masculinize the SNB prenatally, DHT may be involved in masculinizing postnatal aspects of SNB development.