RESUMO
Background: Treatment options for soft tissue sarcoma (STS) patients aged ≥65 years (elderly) can be limited by concerns regarding the increased risk of toxicity associated with standard systemic therapies. Trabectedin has demonstrated improved disease control in a phase III trial (ET743-SAR-3007) of patients with advanced liposarcoma or leiomyosarcoma after failure of anthracycline-based chemotherapy. Since previous retrospective analyses have suggested that trabectedin has similar safety and efficacy outcomes regardless of patient age, we carried out a subgroup analysis of the safety and efficacy observed in elderly patients enrolled in this trial. Patients and methods: Patients were randomized 2 : 1 to trabectedin (n = 384) or dacarbazine (n = 193) administered intravenously every-3-weeks. The primary end point was overall survival (OS); secondary end points were progression-free survival (PFS), time-to-progression, objective response rate (ORR), duration of response, symptom severity, and safety. A post hoc analysis was conducted in the elderly patient subgroup. Results: Among 131 (trabectedin = 94; dacarbazine = 37) elderly patients, disease characteristics were well-balanced and consistent with those of the total study population. Treatment exposure was longer in patients treated with trabectedin versus dacarbazine (median four versus two cycles, respectively), with a significantly higher proportion receiving prolonged therapy (≥6 cycles) in the trabectedin arm (43% versus 23%, respectively; P = 0.04). Elderly patients treated with trabectedin showed significantly improved PFS [4.9 versus 1.5 months, respectively; hazard ratio (HR)=0.40; P = 0.0002] but no statistically significant improvement in OS (15.1 versus 8.0 months, respectively; HR = 0.72; P = 0.18) or ORR (9% versus 3%, respectively; P = 0.43). The safety profile for elderly trabectedin-treated patients was comparable to that of the overall trabectedin-treated study population. Conclusions: This subgroup analysis of the elderly population of ET743-SAR-3007 suggests that elderly patients with STS and good performance status can expect clinical benefit from trabectedin similar to that observed in younger patients. Trial registration: www.clinicaltrials.gov, NCT01343277.
Assuntos
Antineoplásicos Alquilantes/efeitos adversos , Dacarbazina/administração & dosagem , Leiomiossarcoma/tratamento farmacológico , Lipossarcoma/tratamento farmacológico , Trabectedina/administração & dosagem , Administração Intravenosa , Adolescente , Adulto , Fatores Etários , Idoso , Idoso de 80 Anos ou mais , Antineoplásicos Alquilantes/administração & dosagem , Dacarbazina/efeitos adversos , Esquema de Medicação , Feminino , Humanos , Estimativa de Kaplan-Meier , Leiomiossarcoma/mortalidade , Leiomiossarcoma/patologia , Lipossarcoma/mortalidade , Lipossarcoma/patologia , Masculino , Pessoa de Meia-Idade , Intervalo Livre de Progressão , Fatores de Tempo , Trabectedina/efeitos adversos , Adulto JovemRESUMO
BACKGROUND: Insulin-like growth factor receptor (IGF-1R) has been studied as an oncologic target in soft tissue sarcoma (STS), but its role in sarcoma biology is unclear. Anti-IGF-1R antibody cixutumumab demonstrated acceptable toxicity but limited activity as a single agent in STS. We carried out a dose-escalation study of cixutumumab with doxorubicin to evaluate safety and dosing of the combination. PATIENTS AND METHODS: Eligible patients with advanced STS were treated with cixutumumab intravenously on days 1/8/15 at one of three dose levels (A: 1 mg/kg, B: 3 mg/kg, C: 6 mg/kg) with doxorubicin at 75 mg/m(2) as a 48 h infusion on day 1 of a 21 day cycle. After six cycles of the combination, patients could receive cixutumumab alone. The Time-to-Event Continual Reassessment Method was used to estimate the probability of dose-limiting toxicity (DLT) and to assign patients to the dose with an estimated probability of DLT≤20%. RESULTS: Between September 2008 and January 2012, 30 patients with advanced STS received a median of six cycles of therapy (range <1-22). Two DLTs were observed, grade 3 mucositis (dose level B) and grade 4 hyperglycemia (dose level C). Grade 2 and 3 reduced left ventricular ejection fraction was seen in three and two patients, respectively. Five partial responses were observed, and estimated progression-free survival was 5.3 months (95% confidence interval 3.0-6.3) in 26 response-assessable patients. Immunohistochemical staining of 11 available tumor samples for IGF-1R and phospho-IGF-1R was not significantly different among responders and non-responders, and serum analysis of select single-nucleotide polymorphisms did not predict for cardiotoxicity. CONCLUSION: The maximum tolerated dose was doxorubicin 75 mg/m(2) on day 1 and cixitumumab 6 mg/kg on days 1/8/15 of a 21 day cycle. Cardiac toxicity was observed and should be monitored in subsequent studies, which should be considered in STS only if a predictive biomarker of benefit to anti-IGF-1R therapy is identified. TRIAL REGISTRATION: ClinicalTrials.gov:NCT00720174.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Modelos Estatísticos , Receptor IGF Tipo 1/antagonistas & inibidores , Sarcoma/tratamento farmacológico , Adulto , Idoso , Idoso de 80 Anos ou mais , Anticorpos Monoclonais/administração & dosagem , Anticorpos Monoclonais Humanizados , Doxorrubicina/administração & dosagem , Feminino , Seguimentos , Humanos , Masculino , Dose Máxima Tolerável , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Prognóstico , Receptor IGF Tipo 1/genética , Receptor IGF Tipo 1/metabolismo , Sarcoma/mortalidade , Sarcoma/patologia , Taxa de Sobrevida , Adulto JovemRESUMO
BACKGROUND: The use of potential surrogate end points for overall survival, such as disease-free survival (DFS) or time-to-treatment failure (TTF) is increasingly common in randomized controlled trials (RCTs) in cancer. However, the definition of time-to-event (TTE) end points is rarely precise and lacks uniformity across trials. End point definition can impact trial results by affecting estimation of treatment effect and statistical power. The DATECAN initiative (Definition for the Assessment of Time-to-event End points in CANcer trials) aims to provide recommendations for definitions of TTE end points. We report guidelines for RCT in sarcomas and gastrointestinal stromal tumors (GIST). METHODS: We first carried out a literature review to identify TTE end points (primary or secondary) reported in publications of RCT. An international multidisciplinary panel of experts proposed recommendations for the definitions of these end points. Recommendations were developed through a validated consensus method formalizing the degree of agreement among experts. RESULTS: Recommended guidelines for the definition of TTE end points commonly used in RCT for sarcomas and GIST are provided for adjuvant and metastatic settings, including DFS, TTF, time to progression and others. CONCLUSION: Use of standardized definitions should facilitate comparison of trials' results, and improve the quality of trial design and reporting. These guidelines could be of particular interest to research scientists involved in the design, conduct, reporting or assessment of RCT such as investigators, statisticians, reviewers, editors or regulatory authorities.
Assuntos
Determinação de Ponto Final/normas , Tumores do Estroma Gastrointestinal/terapia , Ensaios Clínicos Controlados Aleatórios como Assunto/normas , Projetos de Pesquisa/normas , Sarcoma/terapia , Terminologia como Assunto , Consenso , Técnica Delphi , Progressão da Doença , Intervalo Livre de Doença , Determinação de Ponto Final/classificação , Tumores do Estroma Gastrointestinal/diagnóstico , Tumores do Estroma Gastrointestinal/mortalidade , Humanos , Ensaios Clínicos Controlados Aleatórios como Assunto/classificação , Sarcoma/diagnóstico , Sarcoma/mortalidade , Fatores de Tempo , Falha de TratamentoRESUMO
BACKGROUND: This retrospective pooled analysis assessed the effect of age on the efficacy and safety of trabectedin in young and elderly patients with recurrent advanced soft tissue sarcoma (STS). METHODS: Data from 350 adults with STS treated in five phase II trials with trabectedin were divided in the younger (<60 years; n=267) and the older cohort (≥60 years; n=83). RESULTS: The response rate did not differ with age (younger: 10.1% vs elderly 9.6%). No significant differences were found in median progression-free survival (PFS) in younger (2.5 months) and older (3.7 months) cohort with a comparable PFS rates at 3 (45.1% vs 55.1%) and 6 months (29.5% vs 36.4%). Similar median overall survival was observed in both cohorts (13.0 vs 14.0 months). Reversible neutropenia and aspartate aminotransferase/alanine aminotransferase elevation were the most common abnormalities. A higher incidence of grade 3/4 neutropenia (43.6% vs 60.2%) and fatigue (6.3% vs 14.4%) was observed in older patients. In 24 patients aged ≥70 years, no significant differences in efficacy or safety outcomes were found. CONCLUSION: This analysis demonstrated that trabectedin is a feasible treatment in young and elderly patients with STS, with meaningful clinical benefits and an acceptable safety profile, essential in palliative treatment of elderly patients.
Assuntos
Antineoplásicos Alquilantes/uso terapêutico , Dioxóis/uso terapêutico , Sarcoma/tratamento farmacológico , Sarcoma/mortalidade , Tetra-Hidroisoquinolinas/uso terapêutico , Adulto , Fatores Etários , Idoso , Idoso de 80 Anos ou mais , Antineoplásicos Alquilantes/efeitos adversos , Dioxóis/efeitos adversos , Intervalo Livre de Doença , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Estudos Retrospectivos , Tetra-Hidroisoquinolinas/efeitos adversos , Trabectedina , Resultado do Tratamento , Adulto JovemRESUMO
BACKGROUND: This expanded access program (EAP) was designed to provide trabectedin access for patients with incurable soft tissue sarcoma (STS) following progression of disease with standard therapy. The outcomes of trial participants accrued over approximately 5 years are reported. PATIENTS AND METHODS: Adult patients with advanced STS of multiple histologies, including leiomyosarcoma and liposarcoma (L-sarcomas), following relapse or disease progression following standard-of-care chemotherapy, were enrolled. Trabectedin treatment cycles (1.5 mg/m(2), intravenously over 24 h) were repeated q21 days. Objective response, overall survival (OS), and safety were evaluated. RESULTS: Of 1895 patients enrolled, 807 (43%) had evaluable objective response data, with stable disease reported in 343 (43%) as best response. L-sarcoma patients exhibited longer, OS compared with other histologies [16.2 months (95% confidence interval (CI) 14.1-19.5) versus 8.4 months (95% CI 7.1-10.7)], and a slightly higher objective response rate [6.9% (95% CI 4.8-9.6) versus 4.0% (95% CI 2.1-6.8)]. The median treatment duration was 70 days representing a median of three treatment cycles; 30% of patients received ≥ 6 cycles. Safety and tolerability in this EAP were consistent with prior clinical trial data. CONCLUSION: Results of this EAP are consistent with previous reports of trabectedin, demonstrating disease control despite a low incidence of objective responses in advanced STS patients after failure of standard chemotherapy. CLINICALTRIALS.GOV: NCT00210665.
Assuntos
Antineoplásicos Alquilantes/administração & dosagem , Ensaios de Uso Compassivo/tendências , Dioxóis/administração & dosagem , Saúde Global/tendências , Sarcoma/tratamento farmacológico , Sarcoma/patologia , Tetra-Hidroisoquinolinas/administração & dosagem , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Antineoplásicos Alquilantes/efeitos adversos , Ensaios de Uso Compassivo/mortalidade , Dioxóis/efeitos adversos , Progressão da Doença , Feminino , Acessibilidade aos Serviços de Saúde/tendências , Humanos , Estimativa de Kaplan-Meier , Masculino , Pessoa de Meia-Idade , Sarcoma/mortalidade , Tetra-Hidroisoquinolinas/efeitos adversos , Trabectedina , Falha de Tratamento , Resultado do Tratamento , Adulto JovemAssuntos
Antígenos CD/análise , Doença de Hodgkin/patologia , Leucemia Linfocítica Crônica de Células B/patologia , Linfoma Difuso de Grandes Células B/patologia , Segunda Neoplasia Primária/patologia , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Medula Óssea/patologia , Terapia Combinada , Ciclofosfamida/administração & dosagem , Doxorrubicina/administração & dosagem , Doença de Hodgkin/tratamento farmacológico , Doença de Hodgkin/imunologia , Humanos , Imuno-Histoquímica , Leucemia Linfocítica Crônica de Células B/tratamento farmacológico , Leucemia Linfocítica Crônica de Células B/imunologia , Linfoma Difuso de Grandes Células B/tratamento farmacológico , Linfoma Difuso de Grandes Células B/imunologia , Masculino , Pessoa de Meia-Idade , Segunda Neoplasia Primária/tratamento farmacológico , Segunda Neoplasia Primária/imunologia , Prednisona/administração & dosagem , Vincristina/administração & dosagemRESUMO
BACKGROUND: Orthotopic liver transplantation (OLT) is frequently complicated by thrombotic events that may threaten the viability of the allograft and severely compromise the overall outcome in these patients. Although multiple prothrombotic pathogenic mechanisms may be involved, a role for inadequate natural anticoagulant levels in the early postoperative period has been postulated. METHODS: We describe a case of a woman who suffered multiple thrombotic complications after a second OLT. Prospective assays of procoagulant and natural anticoagulant factor levels, in addition to screening tests for a variety of inherited and acquired hypercoagulable states, were carried out. RESULTS: Serial studies confirmed an acquired, isolated deficiency of Protein S associated with the second transplanted liver. Protein S levels were normal after the patient's first and third OLTs. There was no laboratory evidence of other underlying prothrombotic conditions. CONCLUSIONS: This unusual case of acquired Protein S deficiency demonstrates that the hypercoagulable phenotype may develop in the recipient of a liver from a heterozygous deficient donor. Furthermore, isolated low Protein S may be causally associated with hepatic artery thrombosis after OLT.
Assuntos
Arteriopatias Oclusivas/etiologia , Artéria Hepática , Transplante de Fígado/efeitos adversos , Deficiência de Proteína S/etiologia , Adulto , Arteriopatias Oclusivas/complicações , Feminino , Heterozigoto , Humanos , Deficiência de Proteína S/complicações , Deficiência de Proteína S/genéticaRESUMO
In vivo studies of Friend virus erythroleukemia have implied that proviral integrations adjacent to the gene for the Ets-related transcription factor PU.1 may inhibit the commitment of erythroblasts to differentiate and cause their capability for indefinite transplantation (C. Spiro, B. Gliniak, and D. Kabat, J. Virol. 62:4129-4135, 1988; R. Paul, S. Schuetze, S. L. Kozak, C. Kozak, and D. Kabat, J. Virol. 65:464-467, 1991). To test this hypothesis, we ligated PU.1 cDNA into a retroviral vector and studied its effects on cultured cells. Infection of fibroblasts with PU.1-encoding retrovirus resulted in PU.1 synthesis followed by nuclear pyknosis, cell rounding, and degeneration. In contrast, in long-term bone marrow cultures, erythroblasts were efficiently and rapidly immortalized. The resulting cell lines were polyclonal populations that contained PU.1, were morphologically blast-like, required erythropoietin and bone marrow stromal cells for survival and proliferation, and spontaneously differentiated at low frequency to synthesize hemoglobin. After 9 months in culture, erythroblasts became stroma independent, and they then grew as clonal cell lines. We conclude that PU.1 perturbs the pathway(s) that controls potential for indefinite proliferation and that it can be used to generate permanent erythroblast cell lines.
Assuntos
Proteínas de Ligação a DNA/fisiologia , Eritroblastos/fisiologia , Eritropoese , Células 3T3 , Animais , Células da Medula Óssea , Transformação Celular Neoplásica , Células Cultivadas , Expressão Gênica , Globinas/genética , Técnicas In Vitro , Camundongos , RNA Mensageiro/genética , Receptores da Eritropoetina/genética , Proteínas Oncogênicas de Retroviridae , TransfecçãoRESUMO
Both viral and cellular genes have been directly implicated in pathogenesis of Friend viral erythroleukemia. The virus-encoded gp55 glycoprotein binds to erythropoietin receptors to cause mitogenesis and differentiation of erythroblasts. However, if the provirus integrates adjacent to the gene for the PU.1 transcription factor, the cell loses its commitment to terminally differentiate and becomes immortal, as indicated by its transplantability and by its potential for indefinite growth in culture (C. Spiro, B. Gliniak, and D. Kabat, J. Virol. 63:4434-4437, 1989; R. Paul, S. Schuetze, S. L. Kozak, and D. Kabat, J. Virol. 65:464-467, 1991). To test the implications of these results, we produced polyclonal antiserum to bacterially synthesized PU.1, and we used it to analyze PU.1 expression throughout leukemic progression and during chemically induced differentiation of Friend erythroleukemia (F-MEL) cell lines. This antiserum identified three electrophoretically distinct PU.1 components in extracts of F-MEL cells and demonstrated their nuclear localization. Although PU.1 proteins are abundant in F-MEL cells, they are absent or present in only trace amounts in normal erythroblasts or in differentiating erythroblasts from the preleukemic stage of Friend disease. Furthermore, chemicals (dimethyl sulfoxide or N,N'-hexamethylenebisacetamide) that overcome the blocked differentiation of F-MEL cells induce rapid declines of PU.1 mRNA and PU.1 proteins. The elimination of PU.1 proteins coincides with recommitment to the program of erythroid differentiation and with loss of immortality. These results support the hypothesis that PU.1 interferes with the commitment of erythroblasts to differentiate and that chemicals that reduce PU.1 expression reinstate the erythropoietic program.
Assuntos
Diferenciação Celular , Proteínas de Ligação a DNA/metabolismo , Leucemia Eritroblástica Aguda/metabolismo , RNA Mensageiro/metabolismo , Fatores de Transcrição/metabolismo , Animais , Proteínas de Ligação a DNA/imunologia , Proteínas de Ligação a DNA/isolamento & purificação , Eritropoese/fisiologia , Vírus da Leucemia Murina de Friend , Camundongos , Proteínas Oncogênicas de Retroviridae , Fatores de Transcrição/imunologia , Fatores de Transcrição/isolamento & purificação , Células Tumorais CultivadasRESUMO
Previous studies identified a common site (Sfpi-1) for proviral integration in immortalized Friend erythroleukemias. cDNAs corresponding to a 1.5-kb Sfpi-1 mRNA were isolated and sequenced. These were larger than an independently isolated Sfpi-1 cDNA described by researchers from another laboratory, and they contained common differences from that sequence, including in the coding region four extra nucleotides that altered the reading frame. The properly translated protein is identical to Pu.1, a transcription activation factor that is related to the ets oncogene family. Genetic methods were used to map Sfpi-1 with respect to other loci on mouse chromosome 2. Our results suggest that Pu.1 blocks erythroblast differentiation and thereby causes immortalization.
Assuntos
Proteínas de Ligação a DNA/genética , Vírus da Leucemia Murina de Friend/genética , Leucemia Experimental/genética , Provírus/genética , Proteínas Oncogênicas de Retroviridae/genética , Fatores de Transcrição/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Linhagem Celular , DNA Viral/genética , Genes Virais , Leucemia Experimental/microbiologia , Camundongos , Dados de Sequência Molecular , Proteínas Tirosina Quinases/genética , RNA Mensageiro/genética , Mapeamento por RestriçãoRESUMO
Substance P (SP) is present in avian sympathetic ganglia and accelerates the decay rate of acetylcholine (ACh)-evoked macroscopic currents in sympathetic neurons. We demonstrate here that SP modulates ACh-elicited single channels in a manner consistent with an enhancement of ACh receptor (AChR) desensitization. Furthermore, since AChR channel function was monitored in cell-attached patches with SP applied to the extra-patch membrane, the peptide must act via a second messenger mechanism. SP specifically decreases the net ACh-activated single-channel current across the patch membrane by decreasing both channel opening frequency and mean open time kinetics. These experiments demonstrate that a peptide can modulate neuronal AChR function by a second messenger mechanism.
Assuntos
Gânglios Simpáticos/fisiologia , Canais Iônicos/fisiologia , Neurônios/fisiologia , Receptores Nicotínicos/fisiologia , Substância P/farmacologia , Acetilcolina/farmacologia , Alcaloides/farmacologia , Animais , Membrana Celular/fisiologia , Células Cultivadas , Embrião de Galinha , Condutividade Elétrica , Eletrofisiologia/métodos , Potenciais Evocados/efeitos dos fármacos , Canais Iônicos/efeitos dos fármacos , Neurônios/efeitos dos fármacos , Inibidores de Proteínas Quinases , Receptores Nicotínicos/efeitos dos fármacos , Sistemas do Segundo Mensageiro , EstaurosporinaRESUMO
By using a tagged derivative of Friend spleen focus-forming virus, we previously obtained evidence that proviral integration(s) in the host genome can cause erythroblast immortality by abrogating the commitment of the cell to differentiate (C. Spiro, B. Gliniak, and D. Kabat, J. Virol. 62:4129-4135, 1988). Exploiting the fact that each leukemia was a single clone that contained one tagged provirus, we have now molecularly cloned and characterized one common genomic site for immortalizing proviral integrations.
Assuntos
Clonagem Molecular , Vírus da Leucemia Murina de Friend/genética , Vírus da Leucemia Murina/genética , Leucemia Eritroblástica Aguda/microbiologia , Lisogenia , Vírus Formadores de Foco no Baço/genética , Animais , Southern Blotting , DNA Viral/genética , Genes Virais , Camundongos , Mapeamento por Restrição , Baço/microbiologia , Vírus Formadores de Foco no Baço/isolamento & purificaçãoRESUMO
Measurement of acetylcholine (ACh)-induced currents indicates that the sensitivity of embryonic sympathetic neurons increases following innervation in vivo and in vitro. We have used single-channel recording to assess the contribution of changes in ACh receptor properties to this increase. Early in development (before synaptogenesis), we detect three classes of ACh-activated channels that differ in their conductance and kinetics. Molecular studies indicating a variety of neuronal receptor subunit clones suggest a similar diversity. Later in development (after innervation), changes in functional properties include increases in conductance and apparent mean open time, the addition of a new conductance class, as well as apparent clustering and segregation of channel types. These changes in channel function are compatible with the developmental increase in ACh sensitivity.
Assuntos
Desenvolvimento Embrionário e Fetal , Neurônios/fisiologia , Receptores Nicotínicos/fisiologia , Sistema Nervoso Simpático/embriologia , Acetilcolina/farmacologia , Animais , Embrião de Galinha , Condutividade Elétrica , Neurônios/efeitos dos fármacos , Tempo de Reação , Sistema Nervoso Simpático/citologiaRESUMO
Many proteins are expressed in distinct embryonic and adult forms. However, in most cases we do not know why the embryonic form of proteins is required. This question can be readily addressed for the acetylcholine receptor (AChR) because developmentally specified modifications of this ligand-gated ion channel can be directly related to changes in membrane currents. In developing rat soleus muscle, spontaneous transmitter release causes miniature end-plate currents (m.e.p.cs) to flow into the muscle cell. We show here that these m.e.p.cs in neonatal soleus trigger spontaneous contractions. By injecting m.e.p.cs into young fibres, we showed that only embryonic m.e.p.cs can trigger such contractions; adult m.e.p.cs do not last long enough. Developing muscle fibres must be active for synapse and muscle differentiation. Our experiments indicate that the embryonic form of the AChR is essential for spontaneous contractile activity and may therefore be required for normal neuromuscular development.
Assuntos
Desenvolvimento Muscular , Receptores Colinérgicos/embriologia , Potenciais de Ação , Animais , Animais Recém-Nascidos , Contração Muscular , RatosRESUMO
Incubating skeletal muscle fibers with forskolin, an activator of adenylate cyclase, increases the rate at which nicotinic acetylcholine receptors (AChRs) desensitize when exposed to ACh. Several reports indicate that this is due to the phosphorylation of AChRs by cAMP-dependent protein kinase, but other studies suggest that forskolin interacts with AChRs directly and that second-messenger systems are not required. To help clarify this issue, we studied the effects of forskolin and several other drugs on AChR function in embryonic rat myotubes. AChR function was studied by recording ACh-induced membrane depolarizations and ACh-induced single-channel currents. Our results indicate that forskolin at low concentrations enhances AChR desensitization through the action of a second messenger, most likely cAMP. An analog of forskolin that is much less effective in activating adenylate cyclase (1,9-dideoxyforskolin) is also much less potent in enhancing desensitization. Forskolin at low concentrations does not alter single-channel conductance or mean channel open time. However, when used at concentrations above 20 microM, forskolin may also exert direct drug effects on AChRs.
Assuntos
Colforsina/farmacologia , AMP Cíclico/metabolismo , Membranas Intracelulares/metabolismo , Músculos/metabolismo , Receptores Colinérgicos/metabolismo , Animais , Canais Iônicos/efeitos dos fármacos , Ratos , Receptores Colinérgicos/efeitos dos fármacosRESUMO
1. The burst structures of embryonic-type (low-gamma) and adult-type (high-gamma) nicotinic acetylcholine (ACh) receptors (AChRs) in rat myotubes were investigated with the patch clamp technique. The channels were activated with the agonists ACh and suberyldicholine (SubCh). 2. With either agonist, the distribution of burst durations showed two exponential components for both channel types: a 'long' component that corresponds to bursts of one or more openings and a 'brief' component that includes short, isolated openings. 3. For low-gamma channels, the percentage of all openings associated with the brief component decreased from approximately 40% at 10-100 nM-ACh to less than 10% at 10-100 microM-ACh. 4. Both high-gamma and low-gamma long bursts were interrupted by brief (30-90 microseconds) closures and longer (approximately 1 ms) transitions to a partially open (subconductance) state. The duration of brief closures and partial openings was relatively independent of the agonist, but their frequency within low-gamma bursts was 3-fold higher with SubCh than with ACh. 5. Brief closures are interpreted as transitions to a closed, doubly liganded state from which the channel can reopen. This model predicts that the channel opening rate is greater than 10,000 s-1 for both channel types at room temperature. 6. Estimates of the channel opening rate inferred from the rising phase of miniature end-plate currents recorded from rat soleus fibres are consistent with this interpretation. 7. Both high-gamma and low-gamma channels apparently operate via similar gating mechanisms. Differences in their gating behaviour can be explained in terms of faster kinetic rate constants for high-gamma channels.
Assuntos
Músculos/inervação , Receptores Colinérgicos/fisiologia , Acetilcolina/farmacologia , Potenciais de Ação/efeitos dos fármacos , Animais , Células Cultivadas , Canais Iônicos/fisiologia , Cinética , Músculos/embriologia , RatosRESUMO
During nerve-muscle synapse formation, acetylcholine receptors become localized and modified to allow efficient transfer of information from nerve to muscle. In this paper we summarize our studies on two aspects of receptor modulation--their concentration at synaptic sites and their ability to desensitize in response to prolonged application of agonist. We demonstrate that receptor localization is a complex event which extensively reorganizes the structure of the junctional region. This allows the subsequent influences of contraction to be exerted differently in junctional and extrajunctional regions. We indicate that increases in muscle cell Ca2+ appear to mediate some of the effects of muscle contraction and suggest how regulation of Ca2+ levels may specify junctional and extrajunctional differences. Finally, we discuss the role of receptor phosphorylation in determining the rate of desensitization.