Exploring synaptic circuits with antibodies and confocal microscopy PartII: Principles and...
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Exploring synaptic circuits with antibodies and confocal microscopy PartII: Principles and applications of confocal microscopy
Marco SassoèDip. Anatomia, Farmacologia e Medicina LegaleIstituto Nazionale di NeuroscienzeTorino, Italy
Doctoral School of Neuroscience
Turin University
June 2009
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3. In a confocal microscope the vertical resolution is:
a. The same as the lateral resolution
b. Better than the lateral resolution
c. Worse than the lateral resolution
2. Compared to a conventional epifluorescence microscope
a confocal microscope affords:
a. A much better resolution
b. A marginally better resolution
c. The same resolution
5. In a confocal microscope the specimen is:
a. Illuminated point-by-point
b. Illuminated line-by-line
c. Completely illuminated
1. The major advantage of a confocal microscope is:
a. The use of laser illumination
b. The elimination of out-of-focus fluorescent light
c. The high resolution
d. The presence of a Z-motor 4. The Airy Unit defines:
a. The size of the pinhole
b. The size of the laser beam
c. The intensity of the laser beam
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Development of GABAergic synapses: from synaptic adhesion to activity-dependent synaptogenesis
Synaptic integration of adult-generated neurons
Synapse organization in a mouse model of Rett syndrome
Synaptic plasticity induced by morphine in the mesolimbic pathway
Laboratory of Synapse Development and Plasticity
Federica Briatore
Laura Viltono
Elena Frola
Alessandro Ciccarelli
Marta Pallotto – Patrizia Panzanelli
Elena Boggio
Annarita Patrizi
Giulio Srubek Tomassy
Maurizio Giustetto
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Synapses: 1014 (100-500 trillion) 1 mm3 of cerebral cortex: one billion synapses!
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Martin Chalfie Osamu Shimomura Roger Y. Tsien
The Nobel Prize in Chemistry 2008
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L’eliminazione genetica di MeCP2 induce alterazioni sinaptiche
neurone-specifiche
VI
I strato
IV
V
II-III
100 µm
WT
GFP / NeuN
KO
WT
Strato
1
KO
WT
Strato
5
Elena BoggioMaurizio Giustetto
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Marta Pallotto
WT KO
Ruolo dell’inibizione GABAergica nell’integrazione sinaptica dei
neuroni neogenerati
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Resolution
R = 0,5 / NA
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GABA GABAAR1
PSD95
Fritschy et al., 2006
Panzanelli et al., 2004
Sassoè-Pognetto et al., 2003
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Microscopio a epifluorescenza
Fluorescenza
excitation
emission
fluorochromes
fluorophores
autofluorescence
Indirect (secondary) fluorescence
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The pinhole is conjugate to the focal point of the lens
Sistema confocale•Sorgente luce puntiforme
•Illuminazione puntiforme del campione (con obiettivo ad elevata AN)
•Pinhole su piano di rilevamento
Vantaggi:
-Eliminazione piani fuori fuoco – immagini più nitide
-Eliminazione di interferenza laterale – aumento contrasto
-Sezioni ottiche (e ricostruzioni 3D)
-Miglior risoluzione (teorico)
TUBE LENS
confocal