RESUMEN
A critical element of physician-scientist training is the development and practice of core competencies that promote success in research careers. The ability to develop compelling training and research proposals is one such foundational skill. The NIH Ruth L. Kirschstein National Research Service Award (NRSA) individual fellowship for dual-degree students (F30, F31, or F31-Diversity) creates an ideal opportunity to provide formal instruction in grant-writing skills to physician-scientists early in training. In the guided process of preparing a predoctoral fellowship application, students learn to formulate clear short- and long-term research and training goals; construct a comprehensive, well-reasoned, and rigorous proposal; become familiar with funding agency priorities; and gain strategic insights into the peer review system. Beyond building scientific writing skills, the application process for an NRSA F30 or F31 is an opportunity for trainees to strengthen mentor-mentee relationships, identify learning opportunities key to their scientific development, and build effective research and mentoring teams. These skills also apply to developing future postdoctoral mentored K applications or faculty research program grants. Here, we outline key features of the structured proposal development training developed for students in the Yale MD-PhD Program and review outcomes associated with its implementation.
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Distinciones y Premios , Médicos , Humanos , Becas , Mentores , DocentesRESUMEN
The average age when physician-scientists begin their career has been rising. Here, we focused on one contributor to this change: the increasingly common decision by candidates to postpone applying to MD-PhD programs until after college. This creates a time gap between college and medical school. Data were obtained from 3544 trainees in 73 programs, 72 program directors, and AAMC databases. From 2013 to 2020, the prevalence of gaps rose from 53% to 75%, with the time usually spent doing research. Gap prevalence for MD students also increased but not to the same extent and for different reasons. Differences by gender, underrepresented status, and program size were minimal. Most candidates who took a gap did so because they believed it would improve their chances of admission, but gaps were as common among those not accepted to MD-PhD programs as among those who were. Many program directors preferred candidates with gaps, believing without evidence that gaps reflects greater commitment. Although candidates with gaps were more likely to have a publication at the time of admission, gaps were not associated with a shorter time to degree nor have they been shown to improve outcomes. Together, these observations raise concerns that, by promoting gaps after college, current admissions practices have had unintended consequences without commensurate advantages.
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Médicos , Educación de Postgrado en Medicina , Humanos , InvestigadoresRESUMEN
The cellular mechanisms contributing to long-term potentiation and activity-induced formation of glutamatergic synapses have been intensely debated. Recent studies have sparked renewed interest in the role of presynaptic components in these processes. Based on the present evidence, it appears likely that long-term plasticity utilizes both pre- and postsynaptic expression mechanisms.
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Plasticidad Neuronal , Terminales Presinápticos/fisiología , Potenciales de Acción , Animales , Ácido Glutámico/metabolismo , Ácido Glutámico/fisiología , Humanos , Neurotransmisores/metabolismo , Neurotransmisores/fisiología , Sinapsis/fisiologíaRESUMEN
Nicotinic acetylcholine receptors (nAChRs) affect a wide array of biological processes, including learning and memory, attention, and addiction. lynx1, the founding member of a family of mammalian prototoxins, modulates nAChR function in vitro by altering agonist sensitivity and desensitization kinetics. Here we demonstrate, through the generation of lynx1 null mutant mice, that lynx1 modulates nAChR signaling in vivo. Its loss decreases the EC(50) for nicotine by approximately 10-fold, decreases receptor desensitization, elevates intracellular calcium levels in response to nicotine, and enhances synaptic efficacy. lynx1 null mutant mice exhibit enhanced performance in specific tests of learning and memory. Consistent with reports that mutations resulting in hyperactivation of nAChRs can lead to neurodegeneration, aging lynx1 null mutant mice exhibit a vacuolating degeneration that is exacerbated by nicotine and ameliorated by null mutations in nAChRs. We conclude that lynx1 functions as an allosteric modulator of nAChR function in vivo, balancing neuronal activity and survival in the CNS.
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Encéfalo/metabolismo , Glicoproteínas de Membrana/metabolismo , Neuronas/metabolismo , Neuropéptidos/metabolismo , Nicotina/farmacología , Agonistas Nicotínicos/farmacología , Receptores Nicotínicos/metabolismo , Proteínas Adaptadoras Transductoras de Señales , Factores de Edad , Animales , Aprendizaje por Asociación/efectos de los fármacos , Aprendizaje por Asociación/fisiología , Encéfalo/efectos de los fármacos , Encéfalo/patología , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/fisiología , Agonistas de Aminoácidos Excitadores/farmacología , Glicoproteínas de Membrana/efectos de los fármacos , Glicoproteínas de Membrana/genética , Potenciales de la Membrana/efectos de los fármacos , Potenciales de la Membrana/fisiología , Ratones , Ratones Mutantes , Mutación , Degeneración Nerviosa/metabolismo , Degeneración Nerviosa/patología , Neuronas/efectos de los fármacos , Neuronas/patología , Neuropéptidos/efectos de los fármacos , Neuropéptidos/genética , Técnicas de Placa-Clamp , Receptores Nicotínicos/efectos de los fármacosRESUMEN
The use of fluorescent probes such as FM 1-43 or synapto-pHluorin to study the dynamic aspects of synaptic function has dramatically increased in recent years. The analysis of such experiments is both labor intensive and subject to potentially significant experimenter bias. For our analysis of fluorescently labeled synapses in cultured hippocampal neurons, we have developed an automated approach to punctum identification and classification. This automatic selection and processing of fluorescently labeled synaptic puncta not only reduces the chance of subjective bias and improves the quality and reproducibility of the analyses, but also greatly increases the number of release sites that can be rapidly analyzed from a given experiment, increasing the signal-to-noise ratio of the data. An important innovation to the automation of analysis is our method for objectively selecting puncta for analysis, particularly important for studying and comparing dynamic functional properties of a large population of individual synapses. The fluorescence change for each individual punctum is automatically scored according to several criteria, allowing objective assessment of the quality of each site. An entirely automated and thus unbiased analysis of fluorescence in the study of synaptic function is critical to providing a comprehensive understanding of the cellular and molecular underpinnings of neurotransmission and plasticity.
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Procesamiento de Imagen Asistido por Computador/métodos , Sinapsis/fisiología , Animales , Células Cultivadas , Interpretación Estadística de Datos , Colorantes Fluorescentes , Hipocampo/citología , Hipocampo/metabolismo , Ratones , Microscopía Fluorescente , Plasticidad Neuronal/fisiología , Neuronas/metabolismo , Programas Informáticos , Transmisión Sináptica/fisiologíaRESUMEN
Rett syndrome is an X-linked neurodevelopmental disorder caused by mutations in the gene encoding the transcriptional repressor methyl-CpG-binding protein 2 (MeCP2). Here we demonstrate that the Mecp2-null mouse model of Rett syndrome shows an age-dependent impairment in hippocampal CA1 long-term potentiation induced by tetanic or theta-burst stimulation. Long-term depression induced by repetitive low-frequency stimulation is also absent in behaviorally symptomatic Mecp2-null mice. Immunoblot analyses from behaviorally symptomatic Mecp2-null mice reveal altered expression of N-methyl-d-aspartate receptor subunits NR2A and NR2B. Presynaptic function is also affected, as demonstrated by a significant reduction in paired-pulse facilitation. Interestingly, the properties of basal neurotransmission are normal in the Mecp2-null mice, consistent with our observations that the levels of expression of synaptic and cytoskeletal proteins, including glutamate receptor subunits GluR1 and GluR2, PSD95, synaptophysin-1, synaptobrevin-2, synaptotagmin-1, MAP2, betaIII-tubulin and NF200, are not significantly altered. Together, these data provide the first evidence that the loss of Mecp2 expression is accompanied by age-dependent alterations in excitatory synaptic plasticity that are likely to contribute to the cognitive and functional deficits underlying Rett syndrome.
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Hipocampo/fisiología , Proteína 2 de Unión a Metil-CpG/genética , Proteína 2 de Unión a Metil-CpG/metabolismo , Plasticidad Neuronal/fisiología , Síndrome de Rett/fisiopatología , Animales , Proteínas del Citoesqueleto/metabolismo , Modelos Animales de Enfermedad , Potenciales Postsinápticos Excitadores/fisiología , Potenciación a Largo Plazo/fisiología , Ratones , Ratones Endogámicos C57BL , Ratones Mutantes , Receptores de N-Metil-D-Aspartato/metabolismo , Síndrome de Rett/metabolismo , Transmisión Sináptica/fisiología , Ritmo TetaRESUMEN
Dendritic spines, which are the preferred site of excitatory synapses in the mammalian CNS, are actin-rich structures. We hypothesized that dynamic regulation of actin in spines would differentially affects processes that lead to potentiation vs depression of synaptic efficacy. Here, we report that the expression of long-term depression of excitatory synaptic transmission persists in the presence of actin polymerization in rat hippocampal slices. We observe that the reversal of LTD, de-depression, by high-frequency stimulation was completely blocked. Using electron microscopy, dramatic changes in dendritic spine morphology which accompany the sustained, irreversible depression of excitatory synaptic transmission were observed.
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Actinas/fisiología , Dendritas/metabolismo , Plasticidad Neuronal/fisiología , Neuronas/fisiología , Sinapsis/fisiología , Animales , Compuestos Bicíclicos Heterocíclicos con Puentes/farmacología , Citocalasina D/farmacología , Dendritas/efectos de los fármacos , Dendritas/efectos de la radiación , Estimulación Eléctrica/métodos , Potenciales Postsinápticos Excitadores/efectos de los fármacos , Hipocampo/fisiología , Hipocampo/ultraestructura , Técnicas In Vitro , Masculino , Microscopía Electrónica/métodos , Plasticidad Neuronal/efectos de los fármacos , Plasticidad Neuronal/efectos de la radiación , Neuronas/efectos de los fármacos , Neuronas/efectos de la radiación , Neuronas/ultraestructura , Inhibidores de la Síntesis del Ácido Nucleico/farmacología , Ratas , Ratas Sprague-Dawley , Sinapsis/efectos de los fármacos , Sinapsis/efectos de la radiación , Tiazoles/farmacología , TiazolidinasRESUMEN
Whether contact of an axon with a dendrite is a necessary inductive signal for the assembly of functional presynaptic machinery is controversial. Combining FM1-43 imaging with retrospective immunocytochemistry, we observe many functional synaptic vesicle (SV) release sites lacking postsynaptic specializations in cultured hippocampal neurons. These "orphan" release sites share the same exocytic machinery and mechanisms of endocytic recycling as mature synaptic sites. Moreover, quantitative analysis of FM1-43 destaining at these orphan release sites reveals similar kinetics with slightly lower release probabilities. Time-lapse imaging of FM1-43 reveals that orphans are generated by complete or partial mobilization of synaptic release sites that retain their functionality in transit. Orphan clusters fuse with existing synaptic release sites or form novel release sites onto dendrites. Mobilization and stabilization of orphan boutons to new sites of dendritic contact may represent a necessary presynaptic counterpart to postsynaptic changes observed during development and plasticity in the CNS.
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Axones/metabolismo , Comunicación Celular/fisiología , Dendritas/metabolismo , Terminales Presinápticos/metabolismo , Animales , Separación Celular , Células Cultivadas , Hipocampo/metabolismo , Ratas , Ratas Sprague-DawleyRESUMEN
Abl family nonreceptor tyrosine kinases regulate cell morphogenesis through functional interactions with the actin cytoskeleton. The vertebrate Abl family kinases, Abl and Arg, are expressed in the adult mouse brain, where they may regulate actin cytoskeletal dynamics in mature neurons. Using immunoelectron microscopy, we have localized Abl and Arg to the pre- and postsynaptic compartments of synapses in the mouse hippocampal area CA1. Paired-pulse facilitation (PPF) was significantly reduced at the Schaffer collateral-CA1 (SC-CA1) excitatory synapses in hippocampal slices from abl-/- and arg-/- mice as compared with wild-type mice. Furthermore, treatment of wild-type slices with the specific Abl family kinase inhibitor STI571 also reduced PPF. Basal synaptic transmission, posttetanic potentiation (PTP), long-term potentiation (LTP), and long-term depression (LTD) were similar to wild-type controls in abl-/- and arg-/- slices and in STI571-treated wild-type slices. These results indicate that an important function of Abl and Arg is to modulate synaptic efficacy via a presynaptic mechanism during repetitive activation.
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Hipocampo/fisiología , Plasticidad Neuronal/fisiología , Proteínas Proto-Oncogénicas c-abl/genética , Proteínas Proto-Oncogénicas c-abl/metabolismo , Animales , Benzamidas , Inhibidores Enzimáticos/farmacología , Mesilato de Imatinib , Potenciación a Largo Plazo/fisiología , Depresión Sináptica a Largo Plazo/fisiología , Ratones , Ratones Mutantes , Microscopía Inmunoelectrónica , Técnicas de Cultivo de Órganos , Piperazinas/farmacología , Probabilidad , Proteínas Tirosina Quinasas/antagonistas & inhibidores , Proteínas Tirosina Quinasas/genética , Proteínas Tirosina Quinasas/metabolismo , Proteínas Proto-Oncogénicas c-abl/antagonistas & inhibidores , Pirimidinas/farmacología , Sinapsis/enzimología , Sinapsis/ultraestructura , Transmisión Sináptica/fisiologíaRESUMEN
The acid-sensitive ion channel ASIC1 is a proton-gated ion channel from the mammalian nervous system. Its expression in sensory neurons and activation by low extracellular pH suggest that ASIC is involved in transmitting nociceptive impulses produced by the acidification caused by injury or inflammation. However, ASIC1 expression is not restricted to sensory neurons. To understand the functional role of ASIC1 in the CNS we investigated its expression and subcellular distribution therein. In particular, we examined the presence of ASIC1 in domains where the local pH may drop sufficiently to activate ASIC1 under physiological conditions. Immunostaining with specific antibodies revealed broad expression of ASIC1 in many areas of the adult rat brain including the cerebral cortex, hippocampus and cerebellum. Within cells, ASIC1 was found predominantly throughout the soma and along the branches of axons and dendrites. ASIC1 was not enriched in the microdomains where pH may reach low values, such as in synaptic vesicles or synaptic membranes. Pre- or postsynaptic ASIC1 was not gated by synaptic activity in cultured hippocampal neurons. Blockage or desensitization of ASIC1 with amiloride or pH 6.7, respectively, did not modify postsynaptic currents. Finally, the ontogeny of ASIC1 in mouse brain revealed constant levels of expression of ASIC1 protein from embryonic day 12 to the postnatal period, indicating an early and almost constant level of expression of ASIC1 during brain development.