Analysis of SNAP-25 immunoreactivity in hippocampal inhibitory neurons during development in culture and in situ.
Synaptosomal associated protein of 25 kDa (SNAP-25) is a component of the soluble N-ethylmaleimide-sensitive fusion protein (NSF) attachment protein receptor (SNARE) complex which plays a central role in synaptic vesicle exocytosis. We have previously demonstrated that adult rat hippocampal GABAergic synapses, both in culture and in brain, are virtually devoid of SNAP-25 immunoreactivity and are less sensitive to the action of botulinum toxin type A, which cleaves this SNARE protein [Neuron 41 (2004) 599]. In the present study, we extend our findings to the adult mouse hippocampus and we also provide demonstration that hippocampal inhibitory synapses lacking SNAP-25 labeling belong to parvalbumin-, calretinin- and cholecystokinin-positive interneurons. A partial colocalization between SNAP-25 and glutamic acid decarboxylase is instead detectable in developing mouse hippocampus at P0 and, at a lesser extent, at P5. In rat embryonic hippocampal cultures at early developmental stages, SNAP-25 immunoreactivity is detectable in a percentage of GABAergic neurons, which progressively reduces with time in culture. Consistent with the presence of the substrate, botulinum toxin type A is partially effective in inhibiting synaptic vesicle recycling in immature GABAergic neurons. Since SNAP-25, beside its role as a SNARE protein, is involved in additional processes, such as neurite outgrowth and regulation of calcium dynamics, the presence of higher levels of the protein at specific stages of neuronal differentiation may have implications for the construction and for the functional properties of brain circuits.
Frassoni
C
C
Department of Medical Pharmacology, CNR Institute of Neuroscience, University of Milano, Center of Excellence on Neurodegenerative Diseases, Milano, Italy.
Inverardi
F
F
Coco
S
S
Ortino
B
B
Grumelli
C
C
Pozzi
D
D
Verderio
C
C
Matteoli
M
M
D000818:Animals; D015415:Biomarkers; D019274:Botulinum Toxins, Type A / Q000494:pharmacology; D002454:Cell Differentiation / Q000187:drug effects / Q000502:physiology; D002478:Cells, Cultured; D019084:Fluorescent Antibody Technique, Indirect; D006624:Hippocampus / Q000166:cytology / Q000378:metabolism*; D007150:Immunohistochemistry; D007395:Interneurons / Q000378:metabolism; D008297:Male; D008565:Membrane Proteins / Q000276:immunology / Q000378:metabolism*; D051379:Mice; D009419:Nerve Tissue Proteins / Q000276:immunology / Q000378:metabolism*; D009465:Neuromuscular Agents / Q000494:pharmacology; D009474:Neurons / Q000378:metabolism*; D012333:RNA, Messenger / Q000096:biosynthesis / Q000302:isolation & purification; D051381:Rats; D017207:Rats, Sprague-Dawley; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013569:Synapses / Q000187:drug effects / Q000502:physiology; D013572:Synaptic Vesicles / Q000187:drug effects; D050825:Synaptosomal-Associated Protein 25; D005680:gamma-Aminobutyric Acid / Q000502:physiology
D015415:Biomarkers; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D009465:Neuromuscular Agents; D012333:RNA, Messenger; C494209:Snap25 protein, mouse; C494210:Snap25 protein, rat; D050825:Synaptosomal-Associated Protein 25; D005680:gamma-Aminobutyric Acid; D019274:Botulinum Toxins, Type A