RESUMO
We report on a fiber laser-based stimulated emission-depletion microscope providing down to â¼20 nm resolution in raw data images as well as 15-19 nm diameter probing areas in fluorescence correlation spectroscopy. Stimulated emission depletion pulses of nanosecond duration and 775 nm wavelength are used to silence two fluorophores simultaneously, ensuring offset-free colocalization analysis. The versatility of this superresolution method is exemplified by revealing the octameric arrangement of Xenopus nuclear pore complexes and by quantifying the diffusion of labeled lipid molecules in artificial and living cell membranes.
Assuntos
Difusão , Microscopia de Fluorescência/métodos , Nanotecnologia/métodos , Animais , Sobrevivência Celular , Cor , Lasers , Microscopia de Fluorescência/instrumentação , Nanotecnologia/instrumentação , Fibras Ópticas , XenopusRESUMO
Neuregulin-1 (NRG1) gene variants are associated with increased genetic risk for schizophrenia. It is unclear whether risk haplotypes cause elevated or decreased expression of NRG1 in the brains of schizophrenia patients, given that both findings have been reported from autopsy studies. To study NRG1 functions in vivo, we generated mouse mutants with reduced and elevated NRG1 levels and analyzed the impact on cortical functions. Loss of NRG1 from cortical projection neurons resulted in increased inhibitory neurotransmission, reduced synaptic plasticity, and hypoactivity. Neuronal overexpression of cysteine-rich domain (CRD)-NRG1, the major brain isoform, caused unbalanced excitatory-inhibitory neurotransmission, reduced synaptic plasticity, abnormal spine growth, altered steady-state levels of synaptic plasticity-related proteins, and impaired sensorimotor gating. We conclude that an "optimal" level of NRG1 signaling balances excitatory and inhibitory neurotransmission in the cortex. Our data provide a potential pathomechanism for impaired synaptic plasticity and suggest that human NRG1 risk haplotypes exert a gain-of-function effect.
Assuntos
Neuregulina-1/metabolismo , Plasticidade Neuronal , Células Piramidais/fisiologia , Animais , Região CA1 Hipocampal/citologia , Região CA1 Hipocampal/fisiologia , Movimento Celular , Condicionamento Psicológico , Espinhas Dendríticas/fisiologia , Medo , Feminino , Expressão Gênica , Interneurônios/fisiologia , Masculino , Camundongos Transgênicos , Rede Nervosa , Neuregulina-1/genética , Transmissão SinápticaRESUMO
We demonstrated superresolution optical microscopy in a living higher animal. Stimulated emission depletion (STED) fluorescence nanoscopy reveals neurons in the cerebral cortex of a mouse with <70-nanometer resolution. Dendritic spines and their subtle changes can be observed at their relevant scales over extended periods of time.