Functional expression of release-regulating glycine transporters GLYT1 on GABAergic neurons and GLYT2 on astrocytes in mouse spinal cord.
It is widely accepted that glycine transporters of the GLYT1 type are situated on astrocytes whereas GLYT2 are present on glycinergic neuronal terminals where they mediate glycine uptake. We here used purified preparations of mouse spinal cord nerve terminals (synaptosomes) and of astrocyte-derived subcellular particles (gliosomes) to characterize functionally and morphologically the glial versus neuronal distribution of GLYT1 and GLYT2. Both gliosomes and synaptosomes accumulated [3H]GABA through GAT1 transporters and, when exposed to glycine in superfusion conditions, they released the radioactive amino acid not in a receptor-dependent manner, but as a consequence of glycine penetration through selective transporters. The glycine-evoked release of [3H]GABA was exocytotic from synaptosomes but GAT1 carrier-mediated from gliosomes. Based on the sensitivity of the glycine effects to selective GLYT1 and GLYT2 blockers, the two transporters contributed equally to evoke [3H]GABA release from GABAergic synaptosomes; even more surprising, the 'neuronal' GLYT2 contributed more efficiently than the 'glial' GLYT1 to mediate the glycine effect in [3H]GABA releasing gliosomes. These functional results were largely confirmed by confocal microscopy analysis showing co-expression of GAT1 and GLYT2 in GFAP-positive gliosomes and of GAT1 and GLYT1 in MAP2-positive synaptosomes. To conclude, functional GLYT1 are present on neuronal axon terminals and functional GLYT2 are expressed on astrocytes, indicating not complete selectivity of glycine transporters in their glial versus neuronal localization in the spinal cord.
Raiteri
Luca
L
Department of Experimental Medicine, Pharmacology and Toxicology Section, University of Genoa, Italy. raiteri@pharmatox.unige.it
Stigliani
Sara
S
Usai
Cesare
C
Diaspro
Alberto
A
Paluzzi
Silvio
S
Milanese
Marco
M
Raiteri
Maurizio
M
Bonanno
Giambattista
G
D000818:Animals; D001253:Astrocytes / Q000378:metabolism*; D050487:Glycine Plasma Membrane Transport Proteins / Q000378:metabolism*; D051379:Mice; D018613:Microscopy, Confocal; D009474:Neurons / Q000378:metabolism*; D013116:Spinal Cord / Q000166:cytology / Q000378:metabolism*; D005680:gamma-Aminobutyric Acid / Q000378:metabolism*
D050487:Glycine Plasma Membrane Transport Proteins; C493270:Slc6a5 protein, mouse; C493267:Slc6a9 protein, mouse; D005680:gamma-Aminobutyric Acid