Lecture 6 Mechanisms and Mediators of Neurovascular Coupling

Question 1

Briefly overview the first neurovascular ideas/research

Answer 1

Around 1880, Angelo Mosso Balance Board

1890 Roy and Sherrington conducted research on dogs from which they concluded vascular supply in brain can be varied locally in correspondence with functional activity

Question 2

describe what makes Murine models, good models in NV research

Answer 2

• we have a good understanding of anatomy (barrel cortex). With preserved anatomy from snout to barrelletes in the hindbrain, to barreloids in the VPN thalamus to barrels in the somatosensory cortex- similar to human trigeminal projections. With a good topographic map of murine model visible on cortex.
• easy to stimulate - whisking
• easy to access barrel cortex for measurements- thinning skull

Question 2

advantages of anaesthetised animals

Answer 2

• high levels of control
• able to be invasive
• a good anaesthetic can mimic quiet awake state in brain activity

Question 2

disadvantages of anaesthetised animals

Answer 2

• animals aren’t awake so difficult to compare to human fMRI
• Anaesthetic can alter NV coupling

Question 3

advantages of awake animals

Answer 3

• looking at more natural behaviour
• more comparable to human fMRI

Question 4

disadvantages of awake animals

Answer 4

• behaviour can confound results
• more resources required e.g. money and time to train
• welfare implications
• more high risk- if animals die, data is lost

Question 5

advantages of ex-vivo slice research

Answer 5

• good spatial precision (can isolate single cells)
• accurate pharmacology experiments- can see how drugs are acting
• have been vital in showing role of pericytes

Question 6

disadvantages of ex-vivo slice research

Answer 6

lack of active blood flow (major issues of performing NV research)
- time scale responses different- usually much slower
- questions about how healthy the slice is

Question 7

3 key cells of NV unit

Answer 7

astrocytes
pericytes
endothelial cells

Question 8

where are astrocytes found

Answer 8

between neurons and vessels- with feet on BVs

Question 9

where are pericytes found

Answer 9

around blood vessels

Question 10

where are endothelial cells found

Answer 10

lining blood vessels

Question 11

Zonta et al 2003 (astrocytes)

Answer 11

slice work with electrode stimulatiom, found that electrode stimulation to neurons, activated nearby astrocytes to increase Ca2+ , causing vasodilation of nearby vessels

Question 12

describe the prostaglandin pathway

Answer 12

astrocytes uptake glutamate from synapses, causing an increase in Ca2+ which activates the prostaglandin pathway where phospholipase transform phospholipids into prostanoids, via COX 1 and 2, prostanoids dilate blood vessels

Question 13

what do COX inhibitors do?

Answer 13

inhibit vasodilation

Question 14

how do we know that astrocytes are unlikely wholly responsible for NV coupling?

Answer 14

when COX is inhibited or metabotropic glutamate receptor antagonists are given, there’s 50% reduction in dilation- suggesting other mechanisms (for other 50%)

Question 15

Takano 2006 (astrocytes)

Answer 15

used in-vivo 2 photon calcium imaging in anaesthetised mice. Photolysis of caged calcium, led to vasodilation. Also showed how close to veins and arteries astrocytes were. They found COX inhibitors only reduced vasodilation 50%

Question 16

Nizar et al 2013 (astrocytes)

Answer 16

found astrocyte calcium increase was occurring after vasodilation- but dismissed fast responses as artefacts

Question 17

Lind et al. (astrocytes)

Answer 17

found that astrocyte Ca2+ does elevate before vasodilation- did not dismiss fast responses

Question 18

who has conducted much research on pericytes?

Answer 18

David Atwell

Question 19

discuss David Atwell’s Labs work on pericytes

Answer 19

found that pericyte activation caused vasocontriction and that pericytes seem to regulate blood flow in health and disease, however, this work was all done ex vivo slices and responses were very slow

Question 20

discuss work in conflict to David Atwell’s

Answer 20

Work at Yale suggested that it was actually arteriolar smooth muscle cell contractility causing these changes in blood flow- recorded in live animals

Question 21

what did Zlokovic find 2017 (pericytes)

Answer 21

that pericyte degeneration leads to NV uncoupling and limits O2 in brain- due to disease such as AD

Question 22

what is the issue with much pericyte research

Answer 22

generally done ex vivo, so lacking in vivo work

Question 23

what does Nitric Oxide (NO) do?

Answer 23

vasodilation

Question 24

what enzyme produces NO?

Answer 24

nitric oxide synthase

Question 25

what are the 3 isoforms of NOS?

Answer 25

iNOS (inducible)
eNOS (endothelial)
nNOS (neurons)

Question 26

draw the basic pathway for NO

Answer 26

check book

Question 27

what are the 2 types of inhibitors of NOS?

Answer 27

general and specific

Question 28

what are the 2 types of NOS general inhibitors?

Answer 28

L-NNA and L-NAME

Question 29

what do L-NNA and L-NAME do?

Answer 29

inhibit both eNOS and nNOS

Question 30

name the specific NOS inhibitor

Answer 30

7-NI

Question 31

what does 7-NI do?

Answer 31

inhibits nNOS only

Question 32

what is the issue with 7-NI

Answer 32

difficult to dissolve to cross BBB so has to be injected (neuroinflammation consequences)

Question 33

what’s the issue wit general inhibitors?

Answer 33

target both eNOS and nNOS so difficult to seperate when focussing on NV coupling

Question 34

Lindaver 1999 (NO)

Answer 34

found NO to be a modulator not mediator of NV coupling in rat cortex- when SNAP/cGMP were present to make NO available, blood flow was still increased with inhibitor- thought to be involved in basal floow flow

Question 35

Stefanovic 2017 (NO)

Answer 35

NOS inhibitors reduced BOLD response by 50% suggesting role of NO in NV coupling from nNOS interneurons

Question 36

discuss variation of nNOS throughout the brain

Answer 36

found at varying levels e.g. high in the cerebellum, this may mean different regions have differing mechanisms involving NV
each may be main modulator in cerebellum

Question 37

discuss research into the role of endothelium in NV coupling

Answer 37

research suggests that upstream dilation is propagated along vessel wall via gap junctions between smooth muscle cells .
Hillman found when light was used to disrupt vascular endothelium, backward dilation stopped suggesting separate mechanisms for capillaries and surface vessels