Glycinergic neurons expressing enhanced green fluorescent protein in bacterial artificial chromosome transgenic mice.
Although glycine is a major inhibitory transmitter in the mammalian CNS, the role of glycinergic neurons in defined neuronal circuits remains ill defined. This is due in part to difficulties in identifying these cells in living slice preparations for electrophysiological recordings and visualizing their axonal projections. To facilitate the morphological and functional analysis of glycinergic neurons, we generated bacterial artificial chromosome (BAC) transgenic mice, which specifically express enhanced green fluorescent protein (EGFP) under the control of the promotor of the glycine transporter (GlyT) 2 gene, which is a reliable marker for glycinergic neurons. Neurons expressing GlyT2-EGFP were intensely fluorescent, and their dendrites and axons could be visualized in great detail. Numerous positive neurons were detected in the spinal cord, brainstem, and cerebellum. The hypothalamus, intralaminar nuclei of the thalamus, and basal forebrain also received a dense GlyT2-EGFP innervation, whereas in the olfactory bulb, striatum, neocortex, hippocampus, and amygdala positive fibers were much less abundant. No GlyT2-EGFP-positive cell bodies were seen in the forebrain. On the subcellular level, GlyT2-EGFP fluorescence was colocalized extensively with glycine immunoreactivity in somata and dendrites and with both glycine and GlyT2 immunoreactivity in axon terminals, as shown by triple staining at all levels of the neuraxis, confirming the selective expression of the transgene in glycinergic neurons. In slice preparations of the spinal cord, no difference between the functional properties of EGFP-positive and negative neurons could be detected, confirming the utility of visually identifying glycinergic neurons to investigate their functional role in electrophysiological studies.
Zeilhofer
Hanns Ulrich
HU
Department of Experimental and Clinical Pharmacology and Toxicology, University of Erlangen-Nürnberg, 91054 Erlangen, Germany.
Studler
Barbara
B
Arabadzisz
Dimitrula
D
Schweizer
Claude
C
Ahmadi
Seifollah
S
Layh
Beate
B
Bösl
Michael R
MR
Fritschy
Jean-Marc
JM
D026921:Amino Acid Transport Systems, Neutral / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D015415:Biomarkers / Q000378:metabolism; D001921:Brain / Q000166:cytology / Q000378:metabolism*; D022202:Chromosomes, Artificial, Bacterial / Q000378:metabolism*; D005260:Female; D005786:Gene Expression Regulation; D005818:Genetic Engineering / Q000379:methods; D005998:Glycine / Q000378:metabolism*; D050487:Glycine Plasma Membrane Transport Proteins; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism*; D007150:Immunohistochemistry; D007395:Interneurons / Q000378:metabolism; D049408:Luminescent Agents / Q000378:metabolism; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D009434:Neural Pathways / Q000166:cytology / Q000378:metabolism*; D009474:Neurons / Q000166:cytology / Q000378:metabolism*; D011401:Promoter Regions, Genetic / Q000235:genetics; D013116:Spinal Cord / Q000166:cytology / Q000378:metabolism
D026921:Amino Acid Transport Systems, Neutral; D015415:Biomarkers; D050487:Glycine Plasma Membrane Transport Proteins; D049408:Luminescent Agents; C493270:Slc6a5 protein, mouse; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D005998:Glycine