| ```markdown | |
| **Goal/Experiment:** | |
| The goal of this protocol is to provide a detailed, step-by-step method for performing ALM (anterior lateral motor) window surgery in mice. This includes preparation, craniotomy, window installation, recovery, and post-operative care. This method is designed for researchers conducting in vivo imaging and other experimental studies on cortical structures. | |
| # ALM Window Surgery | |
| **Authors:** | |
| Kayvon Daie¹, Tim Wang¹, Amrita Singh¹, Arseny Finkelstein¹, Marton Rozsa¹, JJ Kim², Karel Svoboda¹ | |
| ¹Janelia Research Campus; ²HHMI Janelia Research Campus, Johns Hopkins University School of Medicine | |
| **Publication Date:** Dec 15, 2020 | |
| **Last Modified:** Oct 23, 2023 | |
| **Protocol ID:** 45619 | |
| ## Abstract | |
| Protocol for Head Post and Cranial Window Surgery developed at Janelia Research Campus in the Svoboda Lab. | |
| Developed and improved by: | |
| - Karel Svoboda for Trachtenberg et al 2002 | |
| - Anthony Holtmaat for Holtmaat et al 2006 and Holtmaat et al 2009 | |
| - Daniel Huber for Huber et al 2012 | |
| Compiled and modified by various researchers over the years, the latest version was compiled by JJ Kim in 2019. | |
| **Note:** This outline provides practical step-by-step advice. Surgeons must adhere to the official animal protocol and current procedures posted on the Svoboda lab wiki. | |
| ## Guidelines | |
| ### ALM Craniotomy Considerations | |
| - **Coordinates**: 1.5 mm lateral and 2.5 mm anterior to Bregma. The medial edge of the circle should fall over the midline. | |
| - **Head post placement**: Just posterior to the outer edge of the glass window. | |
| - **Skull preparation**: Rotate the skull when drilling and measuring. | |
| - **Blood vessel management**: Handle major blood vessels carefully to minimize bleeding. | |
| - **Triple glass window**: Use 2.5 mm/2.5 mm/3 mm glasses. | |
| - **Thinning the skull**: The skull varies in thickness; lateral edges are particularly thick. | |
| - **Avoid dural thickening**: Thin down the skull adequately. | |
| - **Bleeding management**: Blood coagulates in ~90 seconds. | |
| ## Protocol | |
| ### Animal Preparation | |
| 1. **Weigh Mouse**: Transfer mouse to a new cage if needed. Note ear tags or mark first mouse. | |
| 2. **Prepare Cage**: Allow mouse to adjust to a new cage several hours before surgery. | |
| 3. **Food Preparation**: Remove wirebar, provide food pellets, and add dietgel to the cage. | |
| ### Drug Preparation | |
| 4. **Buprenex dilution**: Dose 0.1 mg/kg, draw up using an insulin syringe. Administer dexamethasone (optional), marcaine (50 µL), ketoprofen (5 mg/kg). | |
| ### Preparation and Mounting | |
| 5. **Surgical Prep**: | |
| - **Gloves and Gown**: Wear protective clothing. | |
| - **Prepare Viruses**: If needed. | |
| - **Surgery Area**: Sterilize with Virkon-s and set up the necessary equipment. | |
| 6. **Induction**: | |
| - **Isoflurane**: Turn it to 3%, place the mouse in the induction chamber, and monitor. | |
| 7. **Securing the mouse**: | |
| - **Nose Cone**: Gently secure the mouse ensuring comfort. | |
| 8. **Prepare for Surgery**: | |
| - Cover eyes with artificial tears and inject marcaine subcutaneously. | |
| - Apply betadine and perform a clean incision to remove scalp and expose skull. | |
| ### Craniotomy and Window Installation | |
| 18. **Drill Window**: | |
| - *Mark and drill*: Carefully thin the skull without penetrating fully. | |
| - *Install Window*: Place and secure the triple glass window. | |
| ### Recovery | |
| 38. **Post-Surgery**: | |
| - Lower isoflurane and clean the surgical site. | |
| - Secure mouse on a heating pad until recovery reflexes are observed. | |
| - Administer ketoprofen and buprenex, record the end time and details. | |
| ### Virus Injections (If Applicable) | |
| 44. **Virus Preparation**: | |
| - Backfill pipette with mineral oil, prepare injection apparatus, and spin down the virus if needed. | |
| 48. **Performing Injections**: | |
| - Use stereotactic equipment to inject virus precisely. | |
| ### Post-Operation Care | |
| 41. **Noting Care**: Ensure post-op care instructions are clear for follow-up. | |
| ## End Notes | |
| - **References**: Include scholarly articles for detailed readings. | |
| - **Attachments**: Detailed diagrams and supplementary PDF for visualization. | |
| **endofoutput** | |
| ``` |