Stereotactic frame

Mice were anesthetized with isofluorane, immobilized in a stereotactic frame (David Kopf Instruments), and ascetically injected with 2.5µL of AD-tau in the dentate gyrus (Bregma: −2.5mm; lateral: −2.0mm; depth: −2.2mm) and overlying cortex (Bregma: −2.5mm; lateral: −2.0mm; depth: −1.0mm) using a Hamilton syringe (Hamilton, syringe: 80265–1702RNR; needle: 7803–07). To examine both seeding and spread of tau pathology, mice were unilaterally injected with a total of 2 µg AD-tau (1µg at each injection site). Mice were allowed to recover on a 37⁰C heating pad and monitored for the first 48-hours post-surgery. All animals underwent the same procedures, therefore no randomization was necessary.

Mice were anesthetized with 2% isoflurane in oxygen and were placed into a stereotactic frame (David Kopf Instruments). Collagenase (Type VIIS, Sigma-Aldrich, St. Louis, MO) and blood injections followed previously described procedures (Chen et al., 2011 (link)). A 28–33 g needle attached to a glass Hamilton syringe was inserted into the right striatum at the following stereotaxic coordinates relative to bregma: 0.5 mm rostral, 2.5 mm lateral, 3.5 mm below the skull surface. After 10 min, Collagenase was injected in 1 μl artificial CSF (NaCl 148 mmol/L, KCl 3 mmol/L, CaCL₂ 1 mmol/L, MgCL₂ 0.8 mmol/L, Na₂HPO₄ 0.8 mmol/L, and NaH₂PO₄ 0.2 mmol/L) over 1 minute. Due to variability in the potency of different Collagenase lots, the dose of each lot was adjusted to that which produced ~50% reduction in striatal cell viability via MTT assay (Chen-Roetling et al., 2013 (link)). Dose range was 0.014–0.074 units, and all direct comparisons were made between groups receiving the same Collagenase dose. For striatal autologous blood injection, blood was collected from a tail vein and injected at the above coordinates at a rate of 1 μl/minute. Ten minutes after completion of Collagenase or blood injection, the needle was slowly removed and the wound was closed with sutures. Surgical control mice were subjected to needle trauma only. Mice were recovered in a warm environment.

Male and female mice (6–8 weeks) were deeply anesthetized with 5% isoflurane (vol/vol) in oxygen and placed into a stereotactic frame (Kopf Instruments; Tujunga, CA) upon a heating pad. Anesthesia was maintained at 1–2% isoflurane during surgery. An incision was made along the midline of the scalp and small burr holes were drilled over each injection site. Virus or tracer was delivered via microinjection using a Nanoject II injector (Drummond Scientific Company; Broomall, PA). Injection coordinates for DMS were (in mm from bregma): 0.90 anterior, +/− 1.4 lateral, and −3.0 from surface of the brain. Mice were given subcutaneous injections of meloxicam (10 mg/kg) during surgery and 24 & 48 h after surgery. Mice were group-housed before and after surgery and 4–6 weeks were allowed for viral expression before behavioral training or electrophysiology experiments.