GMC03 project protocol

Dysmorphology and SHIRPA behavioral and neuromuscular assessment in 8 inbred founder strains of the Collaborative Cross   (2020)

German Mouse Clinic and Department of Infection Genetics, HZI
With: Hrabě de Angelis M, Fuchs H, Gailus-Durner V, Becker L, Klopstock T, Hans W, Lengger C, Kollmus H, Schughart K

See also: GMC03 animal documentation


German Mouse Clinic: Phenotyping Pipeline

Modified SHIRPA: 8-10 wks of age

Dysmorphology: 12 wks of age (Supplemental data, see Downloads)

 

  • Project protocol - Contents

    Workflow and sampling

    Step
    Procedure
    Equipment
    Data collected
    1
    Record observations in the viewing jar Viewing jar
    Body position, tremor, palpebral closure, lacrimation, defecation
    2 Record observations in the arena Arena Transfer arousal, locomotor activity, gait, pelvic elevation, tail elevation, startle response, touch escape
    3 Record observations above the arena - Positional passivity, trunk curl, limb grasping
    4 Record observations above the arena on the overlying grid Arena and grid Pinna reflex, corneal reflex
    5 Record observation in the viewing tube Clear plastic tube Contact righting reflex
    6 Record additional observations throughout the procedure All Evidence of biting, vocalization

    Equipment and supplies

    • Viewing jar: Clear Perspex cylindrical vessel (14 cm diameter, 15 cm tall), in-house custom made
    • Arena: Clear Perspex arena
      • 50 x 30 x 18 cm (l x w x h)
      • Floor of arena is a Perspex sheet marked with 15 squares (11 cm).
      • A grid (40 x 20 cm) with 12 mm mesh is secured across the top of the width of the arena
    • Viewing tube: Clear Perspex tube (3.4 cm inner diameter, 22 cm long)
    • Tripod
    • Plastic sheet (sandpapered down to create a surface that is not smooth), 20 x 20 cm
    • IHR click box (20 kHz tone at 90 dB)
    • Stopwatch
    • Dowel rod (14 cm)
    • Fine cotton probe
    • Tissues

    Reagents and solutions

    • Cleaning solution: Alcohol wipes

    Procedure: Modified SHIRPA

      1. Behavior observed in the viewing jar:
        1. The viewing jar is placed upright on top of the plastic sheet.
        2. A mouse is removed from its homecage by gripping the tail between the thumb and forefinger and placed in the viewing jar.
        3. The following behaviors are recorded without disturbing the mouse. Incidents of bizarre behavior, stereotypy and convulsions are recorded as well.
          1. Body position: inactive, active, excessively active
          2. Tremor: trembling (not to be confused with breathing)
          3. Palpebral closure: note
          4. Lacrimation: note
          5. Defecation: note
      2. Behavior observed in the arena:
        1. Each mouse is transferred to the center of the arena inside the viewing jar, using the plastic sheet underneath the jar to contain the mouse. The plastic sheet is carefully removed at ~25 cm above the arena floor.
          1. Transfer arousal: freeze, move immediately, move head, look around, etc.
          2. Locomotor activity: total number of squares the mouse enters with all four feet within 30 s.
          3. Gait: retropulsion, uncoordinated movement, etc.
          4. Pelvic elevation: extent pelvis is elevated
          5. Tail elevation: dragging, horizontally extended, elevated
          6. Startle response: response of mouse following 90 dB tone from click box ~30 cm above arena
          7. Touch escape: response of mouse following a stroke on the back of the neck with a bent index finger (mouse approached from front).
      3. Behavior observed outside (above) the arena
        1. Each mouse is removed from the arena by gripping the tail between the thumb and forefinger to observe the following:
          1. Positional passivity: record immediate response to a sequence of handling positions (once a response is recorded, the sequence is terminated and no more positions are used):
            1. Held by tail: record (if mouse does not struggle, move to next test; if mouse struggles, move to trunk curl, below).
            2. Held by neck (loose scruff between the forefinger and thumb): record (if mouse does not struggle, move to next test; if mouse struggles, move to trunk curl, below).
            3. Held in supine position: record if mouse struggles or not.
          2. Trunk curl: note forward curling of each mouse when held by tail.
          3. Limb grasping: note whether mouse grasps its limbs when held by tail.
      4. Behavior observed above the arena on the overlying grid
        1. Pinna reflex: place each mouse gently on the grid and touch proximal part of inner canthus lightly with the tip of a fine cotton probe; record ear retraction.
        2. Corneal reflex: touch cornea lightly with the tip of a fine cotton probe; record eye-blink response.
      5. Behavior observed in the clear plastic tube: place each mouse into the tube and roll the tube slowly until the mouse is upside down; record righting reflex.
      6. Other behaviors to record during the entire session:
        1. Evidence of biting: note whether mouse attempts to bite handler or equipment.
        2. Vocalization: note whether mouse was vocal at any time.

    Data collected by investigator

    • For SHIRPA, see Workflow table (above)
    • For Dysmorphology, see Supplemental data file (Downloads on project page)
      • Growth/weight/body size
      • Eye
      • Coat
      • Skin
      • Vibrissae
      • Extremities
      • Teeth
      • Ears
      • Musculature
      • Skeletal
      • Neurological/behavioral
      • Respiratory system
      • Reproductive system
      • Other