calcium

Question 1

L type voltage gated calcium channel

Answer 1

open for long periods of time, found in skeletal muscle and cortex

Question 2

T type voltage gated calcium channel

Answer 2

open for short periods of time, found in the heart and thalamus

Question 3

N type voltage gated calcium channel

Answer 3

open for long periods of time, high conductance, found throughout the CNS and PNS

Question 4

P type voltage gated calcium channel

Answer 4

high voltage activated, found in Purkinje cells in cerebellum

Question 5

Ca2+ imaging

Answer 5

measures how much Ca2+ enters the cell

Question 6

Ca2+ indicators

Answer 6

fluoresce when bound to Ca2+, more fluorescence = more intracellular Ca2+ = more neural activity
-can be chemical or GECI’s

Question 7

chemical calcium indicators

Answer 7

small molecules that selectively chelate Ca2+, low specificity and short period of action
-dyes injected into a target region and diffuse into every cell type in the area before their fluorescence is recorded during intracellular calcium waves

Question 8

genetically encoded calcium indicators (GECIs)

Answer 8

fluorescent proteins derived from GFP fused with calmodulin or troponin C, genes encoding these proteins are transfected into cell lines

• can be selective for cellular subtypes
• long term in vivo imaging

Question 9

aequorin

Answer 9

the first GECI, requires coelenterazine, a photoprotein found in jellyfish, fluoresces when bound to free calcium

• wasn’t specific to calcium
• unstable in pH environments
• requires consumption of coelenterazine which is only found in marine animals (doesn’t work in mammals)

Question 10

probes using fluorescence resonance energy transfer (FRET)

Answer 10

a more versatile category of GECI

• genetic calcium indicators derived from GFP
• utilized two fluorescent molecules that when bound to calcium, moved closer together and emitted a specific wavelength of light
• power of technique is low

Question 11

probes that utilize conformational changes of GFP

Answer 11

• GECI category, have limited signal intensity (bad spatial and temporal resolution)
• protein only fluoresced after calcium binding induced a conformational change in the GFP protein itself
• signal to noise ratio was too high

Question 12

G-CaMP

Answer 12

-GECI - calcium probe based on a single GFP molecule with high calcium affinity and intensity, fluoresces when calcium binds to calmodulin inducing a conformational change in cpEGFP (tightens up)

Question 13

limitations of a confocal/two photon microscope (for in vivo microscopy of Ca2+ imaging)

Answer 13

• a two photon microscope can only penetrate 1mm of brain tissue before light is scattered and signal is lost (can only measure cortical activity)
• need to head fix the animal (restriction of movement)

Question 14

fibre miniscope

Answer 14

measures Ca2+ wave recordings in freely moving animals

• heavy, movements were limited
• very little depth of light penetration
• cannot measure single cell activity

Question 15

fiber optics

Answer 15

-implanted single multimode fibre

Question 16

GCaMP Fibre photometry

Answer 16

measures fluorescent change relative to baseline, allows measuring cell population calcium transients in freely moving animals, measures fluorescence given off by GCaMP when bound to calcium