Neuromuscular

PERIPHERAL NERVE: Normal features

Age changes
  Pi granules
Blood-Nerve barrier
Elzholz body
Endoneurium
Epineurium
Perineurium

Morphology
  Sural nerve
    Adults
      Axons
        Unmyelinated
        Myelinated
          Schmidt-Lanterman Cleft
            Morphology
          Node of Raniver
            Ultrastructure
      Epineurium
      Perineurium
      Schwann cells
      Vessels
        Endoneurial
        Epineurial
    Children
  Infraorbital nerve
  Sciatic nerve (Adult)
  Myelin
    Sciatic
    Sural

Mouse

NCAM stain
Non-Myelinating (Remak) Schwann cells in clusters (NCAM stain)
Mild staining is also present in some myelin sheaths


Epineurium
Perineurium
  • Structure
    • Lamellated structure surrounding all nerve fascicles
    • Width
      • Thicker
        • Larger fascicles
        • More proximal nerve: Up to 15 layers
        • At nerve branches
      • Thin: Near distal nerve terminals
    • Continues in
      • CNS: Pia-arachnoid
      • PNS
        • Capsule of some sensory receptors
        • Not: Muscle basal lamina
    • Vessels: Trans-perineurial
      • Smooth muscle layer
        • Continuous in arterioles
        • Incomplete in veins
      • Arteriole size: 10–25 μM
      • May have associated axons
    • Separated from endoneurium by: Subperineurial space
  • Embryology
    • Development period: 12 to 14 weeks
    • Origin: Mesenchymal cells (Fibroblasts)
    • Neurofibromin
      • Associated with: Arrangement & formation of perineurium
  • Perineurial cells
    • Shape: Flattened cells in concentric layers
    • Surrounded by: Basement membrane
      • Thicker than Schwann cell basement membrane
      • May surround one, or several, neighboring perineurial cells
    • Cytoplasm
      • Rarified except in perinuclear region
      • Contains filaments similar to smooth muscle cells
      • Many pinocytotic vescicles
      • Cytoplasm: Linoleic acid high
      • Other molecules
        • Surface complement regulators & inhibitors
          • C1IHN, C4BP, CD59, Clusterin
    • Tight junctions
      • Link adjacent perineurial cells
      • Blood-Nerve barrier
    • Origin
      • ? Fibroblasts
      • Differentiation maintained by
        • desert hedgehog from Schwann cells
        • Neurofibromin
  • Molecular & Ultrastructure
    • Collagen fibrils
      • Width: 40 to 64 nM
      • Contain: Collagen III > I
      • "Raccoon tail" (Long-spaced collagen, Luse body)
    • Non-branching filaments (10 to 12 nM): Oxytalan
    • Laminin isoforms: A, B2 & S chains
    • Stains
    • Junction proteins in perineurium
      • ZO-1 (TJP1)
      • Tricellulin
      • Occludin: Tight junction
      • Claudin-1
      • Claudin-3
      • Claudin-19: Tight junction
      • JAM-C: Adherens junction
      • Cadherin-5: Adherens junction
  • Functions
    • Modulates external stretching forces
    • Regulates endoneurial pressure & milieu
    • Part of blood-nerve barrier: With endoneurial blood vessels
  • Pathology
 

Early Illustration of Perineurium: Ranvier



Endoneurium
Blood-Nerve Barrier 1

Adults: Sural nerve

Structure
Fascicles

Nerve Structure

H&E stain
Normal Endoneurium, Perineurium, Epineurium & Vessels

Nerve fascicles
Contain normal numbers of myelinated axons in endoneurium

VvG stain

Gomori trichrome stain


VvG stain

H&E stain

Nerve (Sural): Axons
Normal
Large & Small axons in endoneurium
Large axons: Surrounded by clear myelin halos
Small axons: Clustered among large myelinated axons

Neurofilament stain


Toluidine blue stain
Axon morphology (Adult): 3 size populations
• Large: Thick myelin sheath
• Intermediate: Thin myelin sheath
• Small: Unmyelinated axons
• Compare to: Infant


Toluidine Blue stained plastic section

Electron micrograph (Robert E Schmidt MD)


Electron micrographs
  (Robert E Schmidt MD)



Unmyelinated axons
  Surrounded by
    Schwann cells

Nerve (Sural): Schwann cells, Non-myelinating

NCAM1 stain
Neural Cell Adhesion Molecule (NCAM1)
  NCAM is abundant in non-myelinating (Remak) Schwann cells that surround Unmyelinated axons
  NCAM is also present in
    Adaxonal Schwann cell cytoplasm around myelinated axons
    Myelin sheaths of some larger axons
Myelinated axons: Are generally smaller than in the more proximal sciatic nerve

NCAM stain

NCAM in Non-myelinating Schwann cells
  Surrounds most small unmyelinated axons (Yellow)
  Not present around large, myelinated axons (Green)
  May occur in the center of larger axons (Yellow)
See: Loss of small axons

Neurofilament stain (Green)
NCAM stain (Red)
Overlap (Yellow)

NCAM & P0
  Normal: Mostly present in different endoneurial cells
  NCAM
    Non-myelinating Schwann cells (Green)
    Small amounts in non-compacted areas of myelinating Schwann cell cytoplasm (Yellow)
  P0: Myelin & Myelinating Schwann cell cytoplasm (Red)
  Schwann Cell Pathology
    Büngner bands: NCAM & P0 present in same Schwann cells
    Onion bulbs

NCAM stain (Green)
p0 stain (Red)

NCAM & MBP
  Normal: Mostly present in different endoneurial cells
  NCAM
    Non-myelinating Schwann cells (Green)
    Small amounts in non-compacted areas of myelinating Schwann cell cytoplasm (Yellow)
  MBP: In Myelin surrounding larger axons (Red)

NCAM stain (Green)
MBP stain (Red)

Epineurial vessels & Supporting cells in endoneurium

ATPase pH 4.3 stain

Epineurial Vessels

VvG stain
Epineurial vessels: Normal
Artery (Black arrow): Linear fibrils near lumen; Thick wall
Vein (White arrow): Interlaced, thin fibrils around outside of vessel; Thin wall


Toluidine blue stain

Gomori trichrome stain

Myelinated axons: Normal numbers
Epineurial vessels
  Intermediate-size & Small
  Alkaline phosphatase: Stains endothelium in some epineurial and endoneurial vessels

Alkaline phosphatase stain

Congo red stain

Nodes of Ranvier
  • Non-myelinated axon segment
      between 2 myelin internodes
  • Anatomy
    • Axon at node
      • Diameter: Reduced
      • Organelle density
        • Increased
      • Length: ~ 1 μm
    • Schwann cells
      • Loops around axon
      • No myelin

Electron micrographs
  (Robert E Schmidt MD)

Schmidt-Lanterman Cleft (Incisure)

Pi (Reich) Granules


Electron micrograph (Robert E Schmidt MD)
Pi Granules

Location
  Myelinating Schwann cells
  Subcellular
    Often adjacent to nucleus
    Some near paranode
Staining
  Metachromatic: Toluidine blue, Hirsch–Peiffer & Methylene blue stains
  Frozen sections: Acid phosphatase, Sudan black & PAS
Size: 1 μM
Structure: Lamellar; 5 nm periodicity
Age
  More common in older patients
  Not present < 8 years of age
Disease associations
  Non-specific
  Not present in recently remyelinated axons
Pi granules: Few or many in cytoplasm of myelinating Schwann cells


From R Schmidt
Elzholz body
  • Location: Schwann cell cytoplasm
  • Structure: Round; Myelin-like periodicity
  • Coomposition: Lipid
  • Cause: ? Myelin remodeling

Mast cell

Myelin & Schwann cells: Sural nerve


P0 (Red)
  Prominent in myelin around most (large & small) myelinated axons
  Smallest axons (Green), often in clusters, have no myelin or associated P0 staining

Neurofilament stain (Green)
P0 stain (Red)

Myelin basic protein (MBP) in Myelin (Red)
  Most prominent in myelin around largest myelinated axons
  In myelin around fewer axons than P0 (Above)

Neurofilament stain (Green)
Myelin basic protein stain (Red)

Non-myelinating Schwann cells

Nerve: Sciatic


Neurofilament stain
Large & small axons: Non-uniform distribution within a fascicle

Neurofilament stain
Large & small axons: Non-uniform distribution among fascicles


Neurofilament stain
  Regions with varied numbers of large & small axons

Neurofilament stain

Neurofilament stain

Neurofilament stain

Neurofilament stain

Non-myelinating Schwann cells

NCAM1 stain
  Regions have varied numbers and distributions of NCAM+ Schwann cells

NCAM1 stain

NCAM1 stain
  Regions with varied numbers of NCAM+ Schwann cells

NCAM1 stain

Myelin Components



Neurofilament stain (Green)
P0 stain (Red)
P0 (Red)
  Prominent in myelin around most (large & small) myelinated axons
  Smallest axons (Green), often in clusters, have no myelin or associated P0 staining

Neurofilament stain (Green)
P0 stain (Red)

Myelin basic protein (MBP) (Red)
  Most prominent in myelin around largest myelinated axons

Neurofilament stain (Green)
Myelin basic protein stain (Red)

P0 vs Myelin Basic Protein (MBP) in Myelin


P0 stain (Green)
Myelin basic protein stain (Red)
P0 protein
  Associated with large & small axons
  More around periphery of myelin sheath
Myelin basic protein
  Mainly associated with largest axons
  More toward inside of myelin sheath

P0 stain (Green)
Myelin basic protein stain (Red)

Nerve: Infraorbital (Branch of Trigeminal)



H&E stain
Myelinated Axons: Abundant all through endoneurium

Gomori trichrome stain

VvG stain
Myelinated Axons: Abundant all through endoneurium

H&E stain

Gomori trichrome stain


Acid phosphatase stain
Infraorbital Nerve
  Acid phosphatase positive cells scattered in endoneurium (Above)
  Alkaline phosphatase positive vessels: Few in endoneurium (Below)

Alkaline phosphatase stain

ATPase activity: Abundant in endoneurial cells

ATPase pH 4.3 stain

Infraorbital Nerve
  Larger myelinated axons: Many
  Small unmyelinated axons: Fewer than in sural nerve

Neurofilament stain


NCAM stain
Infraorbital Nerve
  Non-myelinating (Remak) Schwann cells : Fewer than in sural nerve

NCAM stain
NCAM: Present in Schwann cells
  Non-myelinating (Remak) Schwann cells
  Myelinating Schwann cells: Adaxonal & Abaxonal cytoplasm

NCAM stain

Infraorbital Nerve
  Larger myelinated axons (Green): No associated NCAM; Many
  Small unmyelinated axons (Yellow): Fewer than in sural nerve

Neurofilament stain (Green)
NCAM stain (Red)

Infraorbital Nerve: P0 (Red)
  Prominent in myelin (Red) around most (large & small) myelinated axons
  Smallest axons (Green) have no myelin or associated P0 staining

Neurofilament stain (Green)
P0 stain (Red)

Infraorbital Nerve
  Myelin basic protein (MBP) is markedly reduced in myelin around axons compared to sural nerve

Neurofilament stain (Green)
Myelin basic protein stain (Red)


P0 stain (Green)
Myelin basic protein stain (Red)


VvG stain

Nerve: Infant


Toluidine blue stained plastic sections
Nerves from Infants
  Axons are mildly small
  Myelin sheaths around large axons are moderately thin
  Compare to: Adult

Toluidine blue stained plastic sections


Nerve from neonate
Few axons are myelinated.
Fibroblasts are prominent.

Normal nerve: Mouse

Axon populations: 3
  Myelinated, Large
  Myelinated, Small
  Unmyelinated

From: RE Schmidt

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References
1. Exp Neurol 2020 Mar 3;328:113272

3/15/2023