ABSTRACT
Understanding synaptic connectivity and plasticity within brain circuits and their relationship to learning and behavior is a fundamental quest in neuroscience. Visualizing the fine details of synapses using optical microscopy remains however a major technical challenge. Super resolution microscopy opens the possibility to reveal molecular features of synapses beyond the diffraction limit. With direct stochastic optical reconstruction microscopy, dSTORM, we image synaptic proteins in the brain tissue of the fruit fly, Drosophila melanogaster. Super resolution imaging of brain tissue harbors difficulties due to light scattering and the density of signals. In order to reduce out of focus signal, we take advantage of the genetic tools available in the Drosophila and have fluorescently tagged synaptic proteins expressed in only a small number of neurons. These neurons form synapses within the calyx of the mushroom body, a distinct brain region involved in associative memory formation. Our results show that super resolution microscopy, in combination with genetically labeled synaptic proteins, is a powerful tool to investigate synapses in a quantitative fashion providing an entry point for studies on synaptic plasticity during learning and memory formation.
ABSTRACT
In common with other areas of the prefrontal cortex, activity in frontopolar area 10 is probably modulated by dopamine. We studied the dopaminergic innervation of monkey prefrontal area 10 by immunostaining with tyrosine hydroxylase (TH) antibodies. TH-positive axons in layer 3 were examined by electron microscopy of series of ultrathin sections. TH-positive boutons containing vesicles were sparse (2 × 10(-4) per µm(3)) and the majority (94%, n = 52) had no identifiable synaptic specialization, which supports the hypothesis that dopamine is released non-synaptically and raises the question of whether the local microenvironment surrounding the boutons is special. Compared with unlabelled boutons TH-positive boutons had a higher proportion of their perimeter in contact with dendritic shafts and were more often in continuous contact with pairs of pre- and postsynaptic structures. However, this may result from exclusion from sites preferred by glutamatergic and GABAergic synapses as the density of all synapses in the closer vicinity was no different from any randomly selected site in the neuropil. This quantitative ultrastructural study presents basic features of the dopaminergic innervation in prefrontal area 10 and provides a more detailed understanding of the structural basis of dopamine signalling in the cortex.
