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**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.

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