Green fluorescent protein expression and colocalization with calretinin, parvalbumin, and somatostatin in the GAD67-GFP knock-in mouse.
Gamma-aminobutyric acid (GABA)ergic neurons in the central nervous system regulate the activity of other neurons and play a crucial role in information processing. To assist an advance in the research of GABAergic neurons, here we produced two lines of glutamic acid decarboxylase-green fluorescence protein (GAD67-GFP) knock-in mouse. The distribution pattern of GFP-positive somata was the same as that of the GAD67 in situ hybridization signal in the central nervous system. We encountered neither any apparent ectopic GFP expression in GAD67-negative cells nor any apparent lack of GFP expression in GAD67-positive neurons in the two GAD67-GFP knock-in mouse lines. The timing of GFP expression also paralleled that of GAD67 expression. Hence, we constructed a map of GFP distribution in the knock-in mouse brain. Moreover, we used the knock-in mice to investigate the colocalization of GFP with NeuN, calretinin (CR), parvalbumin (PV), and somatostatin (SS) in the frontal motor cortex. The proportion of GFP-positive cells among NeuN-positive cells (neocortical neurons) was approximately 19.5%. All the CR-, PV-, and SS-positive cells appeared positive for GFP. The CR-, PV, and SS-positive cells emitted GFP fluorescence at various intensities characteristics to them. The proportions of CR-, PV-, and SS-positive cells among GFP-positive cells were 13.9%, 40.1%, and 23.4%, respectively. Thus, the three subtypes of GABAergic neurons accounted for 77.4% of the GFP-positive cells. They accounted for 6.5% in layer I. In accord with unidentified GFP-positive cells, many medium-sized spherical somata emitting intense GFP fluorescence were observed in layer I.
Tamamaki
Nobuaki
N
Department of Morphological Brain Science, Graduate School of Medicine, Kyoto University, Kyoto 606-8501, Japan. tamamaki@mbs.med.kyoto-u.ac.jp
Yanagawa
Yuchio
Y
Tomioka
Ryohei
R
Miyazaki
Jun-Ichi
J
Obata
Kunihiko
K
Kaneko
Takeshi
T
D000818:Animals; D015153:Blotting, Western; D064032:Calbindin 2; D002490:Central Nervous System / Q000737:chemistry*; D015870:Gene Expression; D005968:Glutamate Decarboxylase / Q000032:analysis* / Q000235:genetics; D049452:Green Fluorescent Proteins; D007150:Immunohistochemistry; D017403:In Situ Hybridization; D007527:Isoenzymes / Q000032:analysis* / Q000235:genetics; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D008818:Mice, Neurologic Mutants; D009044:Motor Cortex / Q000737:chemistry; D009474:Neurons / Q000737:chemistry; D010320:Parvalbumins / Q000032:analysis*; D064030:S100 Calcium Binding Protein G / Q000032:analysis*; D013004:Somatostatin / Q000032:analysis*; D005680:gamma-Aminobutyric Acid
C576511:Calb2 protein, mouse; D064032:Calbindin 2; D007527:Isoenzymes; D008164:Luminescent Proteins; D010320:Parvalbumins; D064030:S100 Calcium Binding Protein G; D049452:Green Fluorescent Proteins; D013004:Somatostatin; D005680:gamma-Aminobutyric Acid; D005968:Glutamate Decarboxylase; C402565:glutamate decarboxylase 1