neuro methods

Question 1

main methods used in neuroscience

Answer 1

• Psychophysical methods (e.g. illusions) and behaviour
• Lesions and other ways to silence neurons or parts of the brain
• Anatomical studies and morphology
• fMRI, electrophysiological recordings and imaging
• Modeling and theoretical simulations
• Brain connectomics

Question 2

model organisms

Answer 2

humans
primates
lower mammals - cats/rodents
lower vertebrates - zebrafish
invertebrates - drosophila

Question 3

how are sharp electrode recordings used

Answer 3

• very thin electrodes placed inside neurones
• compare potential between 2 sides of membrane
• predict action potentials

Question 4

disadvantages of sharp electrode recordings

Answer 4

• No solution change inside or outside the cell (some electroinjection is
  still possible)
• Limited possibility for controlling membrane potential
• Cannot measure single channels

Question 5

on cell patch clamp

Answer 5

electrode placed on membrane of cell

Question 6

inside out patch clamp

Answer 6

pull on on-cell patch clamp

Question 7

whole cell patch clamp

Answer 7

suck on on-cell patch clamp
- breaks part of membrane to record currents through rest of membrane
- can use to inhibit cell by inserting inhibitors

Question 8

outside out patch clamp

Answer 8

outer part of membrane exposed and change solution in outer memb/record activity

Question 9

how can you visualise neurone morphology? Any problems?

Answer 9

dye in pipette = diffuses into cell and labels neurone
problems:
- cant label many cells
- limited ability to label specific cell type
- limited to label cellular compartments
- limited for live labelling

Question 10

how to do fluorescence microscopy

Answer 10

excitation filter = makes wavelength more precise
emission filter = passes one type of wavelength
dichroic mirror = reflects light of certain wavelength and allows certain wavelengths through

Question 11

Using GFP to understand neurone function

Answer 11

• Fusion of GFP and two calcium binding proteins
• In presence of calcium these two proteins interact
• GFP becomes much brighter
• increased Ca conc in cytoplasm when neurones are active
  
  • neurones become brighter

Question 12

adv of WC patch clamp over sharp electrode

Answer 12

larger opening at the tip of the patch clamp electrode provides lower resistance and thus better electrical access to the inside of the cell.

Question 13

general principle of patch clamp recordings

Answer 13

• glass pipette containing electrolyte solution is tightly sealed onto the cell membrane and isolates a membrane patch electrically.
• Currents fluxing through the channels in this patch flow into the pipette and can be recorded by an electrode that is connected to a highly sensitive differential amplifier.
• In the voltage-clamp configuration, a current is injected into the cell via a negative feedback loop to compensate changes in membrane potential. Recording this current allows conclusions about the membrane conductance.

Question 14

principles of GCAMP imagins

Answer 14

GCaMP5G is genetically expressed in Gal4-labeled cells and fluoresces with low intensity prior to neuronal stimulation. Application of a stimulus triggers an action potential in the neuron and causes Ca 2+ influx

Question 15

benefits of using confocal microscopes

Answer 15

Confocal microscopy provides the ability to collect clear images from a thin section of a thick sample with low background and minimal out-of-focus interference.
= high spatial resolution

Question 16

how does channel rhodopsin respond to light

Answer 16

blue light = depolarisation (activation)

Question 17

how does halorhodopsin respond to light

Answer 17

yellow light = hyperpolarisation = less likely to fire