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Injections were made as a terminal procedure, after optical imaging and electrophysiological recording, and targeted to patches physiologically identified as object-selective.
In addition, blood sampling to measure metabolite or hormone levels is generally a terminal procedure so repeated sampling is not possible.
As a terminal procedure, the animal was deeply anesthetized and then injected i.v. with saline containing 10% sodium sulfide (200 mg/kg).
In separate experiments, 100 μL of CellTiter-GLO assay reagent (Promega) was added directly to each well of a 96-well plate as a terminal procedure (neurons are lysed).
100 μL caspase3/7-GLO reagent was directly added to each well of a 96-well plate as a terminal procedure (neurons are lysed) and incubated for 1 h.
Blood for all studies was collected from the retroorbital venous plexus under anesthesia using isoflurane as previously described, with the exception of blood for thrombin antithrombin (TAT) evaluation, which was obtained under anesthesia from an open puncture of the inferior vena cava, performed as a terminal procedure.
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For the terminal procedure, mice were anesthetized with Avertin and given a single, i.v. injection of DyLight® 488 labeled tomato lectin (100 µg) (Vector Laboratories)20.
Bronchoalveolar lavage fluid (BALF) was obtained during terminal procedure as previously described (7, 9).
The MPTP intoxication protocol, the chronic L-DOPA treatment, the clinical assessments, the terminal procedure and the characterization of the extent of nigrostriatal denervation were conducted as previously published [3], [11], [12], [13], [14].
As terminal procedure, the knee cavity was washed with PBS (2×5 μl), the total number of leucocytes in the knee lavage was determined by counting the cells in a Neubauer chamber after staining with Turk's solution.
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