Neuroanatomical Techniques

Neuroanatomical Techniques

Presentation by
Armin Blesch, Ph.D.
Harvey Karten, M.D.

Objectives

Neuroanatomical techniques
History of modern neuroanatomy
Rudolf  Albert von Kölliker  (1817-1905)
nucleus  of Kölliker (Rexed  lamina X), continuity of axon and neuron
Heinrich  Wilhelm Gottfried Waldeyer (1837-1921)
Introduced  the term “neuron”  and “chromosome”

Camilio  Golgi   (1843-1926)
Golgi  method; Golgi cells;  Golgi apparatus; Golgi  tendon organ; Golgi-Mazzoni  corpuscle
Santiago  Ramon y Cajal (1852-1934)
Cajal's gold-sublimate method for astrocytes
horizontal  cell of Cajal (Retzius-Cajal cell in cortex
interstitial  nucleus of Cajal

Golgi Stain

Common immunohistochemical stains
Golgi: selective random neuron and fibers
Hematoxylin/Eosin: cell stain
Nissl (thionin): cell body stain
Kluver Barrera: mixed cell fiber stain
Weil: myelinated fiber stain
Acetycholine-esterase
Anterograde and Retrograde Tracing
Brief History of Tracing
(Grafstein, 1967)
(Kristensson & Olsson, 1971)
Fink-Heimer stain
(Heimer 1999)

Chromatolysis

http://cclcm.ccf.org/vm/VM_cases/neuro_cases_PNS_muscle.htm
Anterograde tracing with radioactive amino acids
Edwards and Hendrickson
in: Neuroanatomical tract tracing
Retrograde labeling of spinal motor neurons with HRP
Van der Want  et al.1997
Types of tracers
Application of tracers
Uptake Mechanisms
Active uptake:
Passive incorporation: lipophilic substances
Intracellular injection
Transport
Detection
Fluorescence
Enzyme reaction: HRP (WGA-HRP, CTB-HRP)
Antibodies e.g. CTB
Streptavidin-HRP conjugate for biotinylated tracers e.g. BDA, biocytin
Lectins and Toxins
WGA-HRP
Cholera Toxin beta subunit (CTB)
Retrograde, anterograde and transganglionic
Detection: antibody, HRP conjugate, conjugated to fluorophor
Application: 1 % aqueous solution, iontophoresis or pressure injection
Different efficiency in labeling among different neuronal populatioins and species
Transganglionic tracing of sensory axons with CTB
PHA-L
Anterograde tracing with PHA-L
Gerfen et al. in:
Neuroanatomical tract tracing
FITC/RITC
Fluoresceine isothiocyanate (FITC): green Rhodamine isothiocyanate (RITC): emission >590 nm (red)
Anterograde and retrograde transport
Pressure injection of 1-3% aqueous solution
Lipophilic Carbocyanine Dyes
Lipophilic Carbocyanine Dyes
Labeling of radial glia
Thanos et al. 2000
Dextran amines
Biotinylated dextran amine (BDA)
BDA
Reiner et al. 2000
Anterograde tracing of corticospinal axons
Biocytin/Neurobiotin
Application: 5% solution, pressure injection or iontophoresis
Fast degradation-short survival time 2-3 days
Mostly anterograde transport
Requires glutaraldehyde fixation
Retrograde tracers
All anterograde tracers are partially transported retrogradely
Purely retrograde tracers:
Fast Blue (FB)
Diamidino Yellow (DiY)
Microspheres
Edmund Hollis, UCSD
Scale bar 100 µm
Fluorogold
Fluorogold
Naumann et al. 2000
Ling Wang, UCSD
Cell filling
Viruses
Choosing the Right Tracer
Transgenic “Golgi” stains

GENSAT
 Objective: generate BAC-transgenic mice expressing GFP or CRE under the control of a gene specific promoter
 In situ Hybridization
 Emulsion Autoradiograpy
 Double labeling

Blurton-Jones et al
Blurton-Jones et al

Multiplex mRNA detection
Dave Kosman (Ethan Bier and Bill McGinnis labs, UC San Diego)
http://superfly.ucsd.edu/%7Edavek/images/quad.html
Immunohistochemistry
 Detection Methods
 TSA

Neuroanatomical Techniques.PPT

Read more...

The Athlete’s Knee

The Athlete’s Knee
Presentation by:
John R. (Trey) Green, III, MD
University of Washington
Sports Medicine Clinic
206-543-1552

Anterior Knee Anatomy
* Superficial fascia
* Quadriceps muscle group
* Patella
* Synovium / plicae
* Bursae

Anterior Knee Anatomy
* Quadriceps muscle group
o Rectus femoris
o Vastus medialis
o Vastus lateralis
o Vastus intermedius

Quadriceps Muscle Group
* Vastus medialis
o Larger, and more distal insertion than vastus lateralis
o Oblique distal fibers (VMO)

Patella Articular Surface

* Thickest articular cartilage in the body (5mm)
* 25% non articular (inferior pole)

Anterior Knee Anatomy

* Bursae
o Occur to assist tissue gliding
o Variable location

Patellofemoral Biomechanics

* Patella function
o Act as a fulcrum to increase the lever arm of the quadriceps muscle

Resultant Force on the Patella

* Compression
* 2-3 x body weight
* Maximum force at 70-80 degrees of flexion

Q Angle

* Angle between the quadriceps tendon and patella tendon in full extension

Anterior Knee Pain History
* Pain
* Instability
* Catching
* Crepitation
* Weakness
* Swelling

Physical Examination

* Gait

Physical Examination

* Limb length

Physical Examination
* Compartment assessment (crepitation)
* Observation
Effusion
Prepatellar Bursal Fluid
* Range of Motion (ROM)
* Anterior knee
* Patellar position
* Patellar tracking
* Muscle tone/bulk
* Thigh circumference
* Sitting
* Lying
* Hip abductor strength
* Prone quadriceps tightness

Radiologic Examination
Rosenberg View
Merchant View
Measuring Merchant’sView
Anterior Knee Pain Treatment
* Rehab with therapist
Anterior Knee Pain Treatment
* Refer to Orthopaedist

Assessment of Meniscus, Ligament and Articular Cartilage Injuries
* Most require orthopaedic consultation
History
Pain Location
Meniscus Tears
* Meniscus tears are common
* Rapid flexion with rotation is most common mechanism
* A history of mechanical symptoms and joint line pain and tenderness suggests meniscal tear
Meniscus Tear Types
Meniscus Tears
* Arthroscopy is the gold standard for diagnosis and treatment
* Basic principles of meniscus surgery are:
o Conserve meniscal tissue
o Remove abnormal tissue
o Prevent further tear propagation
o Repair when possible
Articular Cartilage Injury
* Limited intrinsic repair capability
* Likely to eventually progress to arthrosis
* History of pain with recurrent effusions may indicate cartilage lesion
Articular CartilageTreatment Options
Medial Collateral Ligament
Beware Co-existing ACL or PCL Injury
Lateral Sided Ligament Injury
ACL Tear
PCL Tears
Knee Physical Exam
Physical Examination
Collateral Ligament Testing

The Athlete’s Knee.ppt

Read more...

Blood Brain Barrier: Structure, Function and Bypass by Microorganisms

Discovery
What is the Blood Brain Barrier?
Structure of Blood Brain Barrier
Integrity of BBB
Astrocyte end feet
Tight Junctions between BMEC
Claudin
Occludin
Barrier Function of Occludin and Claudin
Junction Adhesion Molecules:
BMEC intercellular space
Barrier function of JAM
Cytoplasmic accessory proteins
Membrane associated guanylate kinase-like proteins (MAGUKS)
Adherens Junction
Pericytes
Astrocyte end feet
Circumventricular organs
Circumventricular organ functions:
Normal BBB transport
Factors produced by astrocytes
Glutamate,
Aspartate
Taurine
ATP
Endothelin-1
NO
MIP-2
Tumor necrosis factor alpha TNF-α
Interleukin beta IL-β
Bradykin
5HT
Histamine
Thrombin
UTP
UMP
Substance P
Qionolonic acid
Platelet activating factor
Free radicals
E. Coli model
Physical damage of BBB
Ligand receptor interactions followed by host cell actin cytoskeletal rearrangements
Transcellular transport while maintaining integrity of BMEC
Physcial damage of BBB
Conclusion
References



Blood Brain Barrier: Structure, Function and Bypass by Microorganisms Download

Read more...