RESUMO
Twelve patients with 3,4-methylenedioxymethamphetamine (MDMA) toxicity from a single rave event presented to multiple San Francisco Bay area hospitals with various life-threatening complications including seizures and hyperthermia. Eight required emergent endotracheal intubation and six had hypotension. Hyperkalemia, acute kidney injury, and rhabdomyolysis were present in most of the patients. In all, 2 patients died, 4 survived with permanent neurologic, musculoskeletal, and/or renal sequelae, and 6 survived without any apparent lasting deficits. Hyperthermia was present in 10 patients and was severe (40.9-43° C) in 7. Using multiple cooling methods, the average time to achieve cooling was 2.7 hours. Serum drug analysis was performed on 3 patients, demonstrating toxic MDMA concentrations without the presence of other xenobiotics. Two capsules confiscated by police at the event contained 82% and 98% MDMA, respectively, without other pharmacologically active compounds. Capsule #2 contained 270 mg MDMA, which is more than twice the amount of MDMA usually contained in 1 dose. The MDMA-induced hyperthermia significantly contributed to the morbidity and mortality in this case series. Factors contributing to the severity of the hyperthermia include ingestion of large doses of MDMA, a warm ambient environment, and physical exertion.
Assuntos
Overdose de Drogas/terapia , Alucinógenos/efeitos adversos , N-Metil-3,4-Metilenodioxianfetamina/efeitos adversos , Adulto , Cuidados Críticos/métodos , Overdose de Drogas/complicações , Overdose de Drogas/epidemiologia , Serviço Hospitalar de Emergência/estatística & dados numéricos , Serviço Hospitalar de Emergência/tendências , Feminino , Humanos , Tempo de Internação/estatística & dados numéricos , Masculino , São Francisco/epidemiologia , Resultado do Tratamento , Adulto JovemRESUMO
CAPS-1 is required for Ca2+-triggered fusion of dense-core vesicles with the plasma membrane, but its site of action and mechanism are unknown. We analyzed the kinetics of Ca2+-triggered exocytosis reconstituted in permeable PC12 cells. CAPS-1 increased the initial rate of Ca2+-triggered vesicle exocytosis by acting at a rate-limiting, Ca2+-dependent prefusion step. CAPS-1 activity depended upon prior ATP-dependent priming during which PIP2 synthesis occurs. CAPS-1 activity and binding to the plasma membrane depended upon PIP2. Ca2+ was ineffective in triggering vesicle fusion in the absence of CAPS-1 but instead promoted desensitization to CAPS-1 resulting from decreased plasma membrane PIP2. We conclude that CAPS-1 functions following ATP-dependent priming as a PIP2 binding protein to enhance Ca2+-dependent DCV exocytosis. Essential prefusion steps in dense-core vesicle exocytosis involve sequential ATP-dependent synthesis of PIP2 and the subsequent PIP2-dependent action of CAPS-1. Regulation of PIP2 levels and CAPS-1 activity would control the secretion of neuropeptides and monoaminergic transmitters.
Assuntos
Proteínas de Ligação ao Cálcio/fisiologia , Exocitose/fisiologia , Fosfatidilinositol 4,5-Difosfato/metabolismo , Vesículas Secretórias/metabolismo , Animais , Cálcio/metabolismo , Cálcio/fisiologia , Proteínas de Ligação ao Cálcio/biossíntese , Proteínas de Ligação ao Cálcio/genética , Células PC12 , Fosfatidilinositol 4,5-Difosfato/antagonistas & inibidores , Ligação Proteica/fisiologia , Ratos , Ratos Endogâmicos , Proteínas de Transporte VesicularRESUMO
Phosphoinositides (PI) are important signaling molecules involved in the regulation of vesicular trafficking. We found that phosphatidylinositol 4-phosphate (PI4P) and phosphatidylinositol 4,5-biphosphate [PI(4,5)P(2)] increase the secretory response triggered by 10 mum Ca(2+) in streptolysin-O-permeabilized insulin-secreting INS-1E cells. In addition, nutrient-induced exocytosis was diminished in intact cells expressing constructs that sequester PI(4,5)P(2) and in cells transfected with constructs that reduce by RNA interference the level of two enzymes involved in PI(4,5)P(2) production, type III PI4-kinase beta and type I phosphatidylinositol 4-bisphosphate 5-kinase-gamma. To clarify the mechanism of action of PI, we investigated the involvement in the regulation of insulin exocytosis of three potential PI targets, phospholipase D1, the Ca(2+)-dependent activator protein for secretion 1, and Munc18-interacting protein 1. Transfection of insulin-secreting cells with plasmids that direct the synthesis of small interfering RNAs capable of reducing the endogenous levels of these proteins inhibited hormone release elicited by glucose- and cAMP-elevating agents without affecting basal release. Our data indicate that the production of PI(4,5)P(2) is necessary for proper control of beta-cell secretion and suggest that at least part of the effect of PI on insulin exocytosis could be exerted through the activation of phospholipase D1, Ca(2+)-dependent activator protein for secretion 1, and Munc18-interacting protein 1.
Assuntos
1-Fosfatidilinositol 4-Quinase/metabolismo , Células Secretoras de Insulina/metabolismo , Insulina/metabolismo , Fosfatidilinositol 4,5-Difosfato/fisiologia , Fosfatos de Fosfatidilinositol/fisiologia , Fosfotransferases (Aceptor do Grupo Álcool)/metabolismo , 1-Fosfatidilinositol 4-Quinase/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Proteínas de Ligação ao Cálcio/metabolismo , Exocitose/efeitos dos fármacos , Exocitose/genética , Exocitose/fisiologia , Secreção de Insulina , Células Secretoras de Insulina/efeitos dos fármacos , Células Secretoras de Insulina/enzimologia , Proteínas de Membrana , Proteínas do Tecido Nervoso/metabolismo , Fosfatidilinositol 4,5-Difosfato/biossíntese , Fosfatidilinositol 4,5-Difosfato/farmacologia , Fosfatos de Fosfatidilinositol/biossíntese , Fosfatos de Fosfatidilinositol/farmacologia , Fosfolipase D/metabolismo , Fosfotransferases (Aceptor do Grupo Álcool)/genética , Interferência de RNA , Ratos , Transdução de SinaisRESUMO
In regulated vesicle exocytosis, SNARE protein complexes drive membrane fusion to connect the vesicle lumen with the extracellular space. The triggering of fusion pore formation by Ca(2+) is mediated by specific isoforms of synaptotagmin (Syt), which employ both SNARE complex and membrane binding. Ca(2+) also promotes fusion pore expansion and Syts have been implicated in this process but the mechanisms involved are unclear. We determined the role of Ca(2+)-dependent Syt-effector interactions in fusion pore expansion by expressing Syt-1 mutants selectively altered in Ca(2+)-dependent SNARE binding or in Ca(2+)-dependent membrane insertion in PC12 cells that lack vesicle Syts. The release of different-sized fluorescent peptide-EGFP vesicle cargo or the vesicle capture of different-sized external fluorescent probes was used to assess the extent of fusion pore dilation. We found that PC12 cells expressing partial loss-of-function Syt-1 mutants impaired in Ca(2+)-dependent SNARE binding exhibited reduced fusion pore opening probabilities and reduced fusion pore expansion. Cells with gain-of-function Syt-1 mutants for Ca(2+)-dependent membrane insertion exhibited normal fusion pore opening probabilities but the fusion pores dilated extensively. The results indicate that Syt-1 uses both Ca(2+)-dependent membrane insertion and SNARE binding to drive fusion pore expansion.