The effects of cutting solutions on the viability of GABAergic interneurons in cerebral cortical slices of adult mice.
To obtain viable GABAergic interneurons in cerebral cortical slices of adult mice, we investigated the effects of slice cutting solutions on the viability of green fluorescent protein (GFP)-expressing cortical neurons in GAD67-GFP knock-in mice. Almost no nuclei of GFP-positive neurons were labeled with propidium iodide in incubated slices, suggesting that GFP fluorescence was a useful indicator for the viability of GABAergic interneurons. When several cutting solutions were compared with saline-based solution, N-methyl-d-glucamine-based sodium-free solution was most effective to keep the number of GFP-positive neurons near the level of perfusion-fixed brain. GFP-positive neurons in slices cut with sodium-free solution were more numerous in cortical layers V-VI, at 30-60 microm depth from the cut surface and 1-6h after cutting than those with saline-based solution. Furthermore, the number of GFP-positive neurons decreased in the cutting condition of high calcium concentration (5mM) or high temperature (37 degrees C), and GFP fluorescence decreased when cut at 0 degrees C. The present results indicate that cutting the brain at 20 degrees C in sodium-free solution is a method for preparing cortical slices with GABAergic interneurons viable. This method would thus be useful for electrophysiological and morphological studies of cortical interneurons in slice preparations of the adult brain.
Tanaka
Yasuyo
Y
Department of Morphological Brain Science, Graduate School of Medicine, Kyoto University, Kyoto 606-8501, Japan.
Tanaka
Yasuhiro
Y
Furuta
Takahiro
T
Yanagawa
Yuchio
Y
Kaneko
Takeshi
T
D000704:Analysis of Variance; D000818:Animals; D002470:Cell Survival; D002540:Cerebral Cortex / Q000166:cytology*; D003470:Culture Media / Q000737:chemistry / Q000494:pharmacology*; D005260:Female; D005968:Glutamate Decarboxylase / Q000235:genetics; D049452:Green Fluorescent Proteins / Q000096:biosynthesis / Q000235:genetics; D066298:In Vitro Techniques; D007395:Interneurons / Q000187:drug effects* / Q000378:metabolism*; D008297:Male; D051379:Mice; D008822:Mice, Transgenic; D013997:Time Factors; D005680:gamma-Aminobutyric Acid / Q000378:metabolism*
D003470:Culture Media; D049452:Green Fluorescent Proteins; D005680:gamma-Aminobutyric Acid; D005968:Glutamate Decarboxylase; C402565:glutamate decarboxylase 1