TECHNIQUES This course outline is designed to provide you with a study guide. Use the lecture, book, and other resources to expand on its contents.

Learning Objectives

1. Describe the steps in tissue preparation and discuss the importance of each.

2. Discuss the basic differences between a cell body and a myelin stain.

3. Explain how neural connections are traced.

4. Discuss how autoradiography is used to measure metabolic activity.

5. Discuss the similarities and differences between CT, MRI, PET, and SPECT scans.

6. Describe ablation procedures and explain the advantages and disadvantages of each.

7. Explain the differences between EEG and evoked potential recording.

8. Discuss the important limitations of ablation and recording in determining brain-behavior relationships.

9. Discuss reversible lesions and describe the advantages and disadvantages as compared with RF lesions.

Histological Procedures / Tissue Preparing Techniques

• Perfusion

• Fixation : Destroy autolytic enzymes

• Sectioning
  • Microtome - 10-80 um thick
  • freezing - quick
  • embedding - paraffin sectioning

• Mounting
  • albumin

• Staining
  • Cell Body - Nissl substance
    • RNA and DNA
    • Cresyl violet - most common

  • Myelin Stains - axons

  • Membrane Stains
    • Golgi-Cox stain (silver)

  • Autoradiography
    • Inject radioactive substance.
    • Labeled amino acids taken up by cell.
    • Brain sections coated with sensitive emulsion develop like photograph.

Tracing Neural Connections

Axoplastic Transport - transport of material along neurotubles.

Tracing Efferent Axons

• Anterograde labeling - use radioactive amino acit then autoradiography to monitor flow down axon along neurotubles.

• Lectins - stain cells of immune system

• Immunocytochemical methods - take advantage of immune reaction

Tracing Afferent Axons

• Retrograde labeling methods - taken up by terminal endings & carried to cell body.
• Horseradish perioxidase - Chemical injected into region
• Taken up by terminal boutons
• Transported to cell body

• example; Flurogold

Horseradish peroxidase

• Enzyme
• Picked up at ends of neuron & transported back to cell.

Tracing Brain Connections

• After person dies, the brain is fixed
• Dil crystals placed on fiber tract
• Travel by diffusion to ends of fibers
• Under UV light, it appears flourescent

Neurochemical Procedures

Three basic ways of locating of neurons that produce chemicals:

• Localizing the chemicals
• Localizing the enzymes that produce
• Localizing the messenger RNA for synthesis

Peptides are localized directly

Autoradiography

Measuring Metabolic Activity

Radioactive 2-deoxyglucose
• Form of glucose - taken up by cell
• High rate activity takes up most
Genetic activity
• Neuronal activity - genes turned on
• Fos protein staining

Neuroradiological Procedures (Brain Imaging)

Structural Measures

• Skull X-ray
• Cerebral angiography - dye in vessels & X-ray
• Pneumoencephalograph - air in ventricles & X-ray

Computerized Axial Tomography (CT Scan)

• X-ray emitter & detector
• Brain scanned from all angles - circular

Magnetic Resonance Imaging (MRI)

• Doesn't use X-rays
• Strong magnetic field
• Nuclei emit radio waves
• Detects radiation from H molecules

Functional Imaging

Regional Cerebral Blood Flow

• Inhale radioactive isotope & use sensors on scalp to detect distribution.

Positron Emission Tomography (PET)

• Inject radioactive 2-DG
• Decay of radioactive isotopes emit positrons
• Detected by PET scanner
• Requires onsite cyclotron

Single Photon Emission Computerized Tomography (SPECT) Scan

• Less expensive but less spatial resolution

Functional Magnetic Resonance Imaging (fMRI)

• MRI sensitive to chemical environment of neural tissue
• Active neurons - increased blood flow
• Doesn't require isotope injection
• Higher resolution than PET

MENU

Problems with Functional Imaging

• Time course of events that these methods measure is slow (min) but neural events fast (ms)
• Measures metabolism not cognitive functioning.

• Doesn't tell function, rather what rest of brain can do in its absence.
• Brain areas highly interconnected
• Nuclei vs fibers of passage
• Bilateral organization
• Aspiration

• Electrical Lesions
  • DC current
  • RF lesions

• Neurotoxin Lesions
  • Kanic acid, Ibotenic acid
  • Destroy cell bodies, not fibers of passage
  • 6-hydroxy-dopamine

• Reversible Lesions
  • Cryogenic Lesions
  • Other Chemicals

Stereotaxic Surgery

• Stereotaxis - Solid Arrangement
• Stereotaxic Atlas (Bregma)
• Stereotaxic Apparatus

Electrical Recording

Electrodes

• Microelectrodes - single cell recording
• Gross electrodes - average electrical events from a specific area
• Surface electrodes - gross averaging of spontaneous activity from large brain regions

EEG Electroencephalogram - measure of spontaneous electical potential from large brain regions

Evoked Potentials use macroelectrodes - measure neural response to stimuli using computer averaging

MEG - magnetoencephalography: measures magnetic fields from brain

Stimulation

• Electrical
• Chemical
• Electrocorticography (ECo) - stimulation of exposed cortex (Penfield).

Terms to Know