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Briefly, transverse cerebellar slices (350 µm) were made at 4°C from brains of 30-day-old mice and maintained in artificial CSF 23°C.
Animals were maintained in artificial water and were starved at least seven days prior to experimentation.
The animals were maintained in artificial sea water at 15 to 18°C under constant light to avoid spawning.
The collected animals were either extracted directly for subsequent chemical analysis of CHCs, or they were maintained in artificial laboratory nests for various experiments (see below).
By "established" we mean the presence of self-sustaining populations in natural bodies of water or man-made canals and lakes, excluding aquaculture facilities and other locations where the snails may have been maintained in artificial environments.
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It has to be modified, joint by joint, muscle by muscle, and thereafter maintained in its artificial state.
C. borealis were purchased from Commercial Lobster (Boston, MA) and maintained in recirculating artificial seawater tanks at 10 13°C for 1 day to several weeks.
After brains were removed from mice following decapitation under deep anesthesia with isoflurane, sagittal slices (400 µm thick) between 1.0 2.0 mm from the midline were prepared using a vibratome (VT1200, Leica Microsystems, Wetzlar, Germany) and were maintained in an artificial cerebral spinal fluid (aCSF -filled holding chaCSF -filledm temperature for 1 holding
The Daphnia were maintained in an artificial freshwater medium [ 55] modified as in [ 41].
In the EGAS technique, the GBM specimen was cut into round thin-slice-core and was maintained in the artificial cerebral spinal fluid (aCSF) with bubbling carbogen gas.
It is worth noting that the exclusive role of layer 5 neurons in the generation of active states was demonstrated only in slices maintained in "modified" artificial cerebrospinal fluid.
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