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Table 2 Potential problems and troubleshooting

From: Craniula: A cranial window technique for prolonged imaging of brain surface vasculature with simultaneous adjacent intracerebral injection

Observation Possible reason Solution
Mouse is not breathing after being placed on stereotactic stage Ear bars are too tight Loosen the ear bars to allow air to flow freely through the trachea.
  Possible respiratory failure Gently release the mouse from the stage and administer oxygen inhalation using an inhaled anesthesia set up (by opening the oxygen flow meter without turning on the isoflurane vaporizer). Depending on the type of anesthesia equipment available, it might be necessary to purge the system before oxygen therapy can be administered. We encourage investigators to consult with a qualified veterinarian for more detailed advice
Vetbond™ on the foramen Excess applied when window was being attached Using a double-pronged tissue pick tool, gently scratch the Vetbond™ surrounding the cannula foramen. The Vetbond™ can be detached smoothly to clean out a space to place the cannula
Cannula misplacement Cannula was not held in place for enough time to allow Vetbond™ to dry Slight movement of the cannula when it is released from the forceps may indicate misplacement of the cannula. Remove the cannula from its position and clean it with sharp forceps to remove Vetbond™ from the foramen and surrounding area. Vetbond™ can be easily removed up to1-2 min following application. Care must be taken because any rough movement can permanently damage the craniula. Clean the foramen completely, taking care that the stainless steel part of the cannula is also free of Vetbond™. Once cleaned, the cannula can be reinserted
Unable to manage syringe containing Super Glue LocTite® Thickness of the Super Glue LocTite® When adding the second layer of Super Glue LocTite®, hold the needle hub firmly and push the plunger of the syringe slowly until a drop of glue forms on the needle bevel; then, gently apply the glue as described in corresponding section
Super Glue LocTite® on top of the cranial window Mistake when adding the second layer of glue Remove the glue with a scalpel by moving it towards the edges of the cover slip. Clean the edge of the scalpel after each stroke, thereby preventing glue from being placed back onto the cover slip
Dummy glued to the cannula Vetbond™ in the groove of the cannula When it is necessary to unscrew a glued dummy, it is possible that the cannula may detach from the skull, and possibly the cranial window will crack. Therefore, gentle and careful movement is essential. To unscrew a glued dummy, two straight mosquito hemostats can be used. Hold the bottom of the cannula with one hemostat. While holding the pedestal with the second hemostat, gently unscrew the dummy. This procedure must be done with the mouse under anesthesia