L10 & 11 - Metabotropic Receptors

Question 1

Two types of synaptic transimission

Answer 1

1) Ionotropic - leads to EPSP (Na+ inflow) or IPSP (Cl- inflow
2) Metabotropic - GPCR, slower than Ionotropic receptors, plays a modulator role - alters ion-channel permeability

Question 2

GPCR has _ TM segments

Answer 2

7 TM alpha helical segments

• aka “serpentine” receptors
• They have no intrinsic activity

Question 3

G protein is a _ protein, with _ subunits

Answer 3

Trimeric protein, alpha, beta, gamma

Question 4

Which parts of the G protein have covalently attached lipid tails to the cell membrane?

Answer 4

Alpha and Gamma

Question 5

GPCR binding process

Answer 5

1) GPCR is bound weakly to G-protein. Upon receptor activation via ligand binding, the affinity for receptor to protein is high
2) Alpha subunit which is bound to GDP (inactive) swaps it for GTP (abundant in cytosol).
3) GTP binding leads to conformational change - dissociationg of a from b, g subunits OR sometimes they also stay together and no change is seen.
4) Leads to TWO effectors. Where Alpha activates target protein, relaying signals to other targets of signalling cascade and Beta-gamma complex does something else to another effector target.
5) GTP hydrolyses to GDP - often accelerated by RGS (Regulator of G protein Signalling)
6) GDP and alpha subunit joins back to b, g compelx

Question 6

Upon prolonged stimulation of GPCR, what occurs?

Answer 6

Receptor kinase activates cytosolic components of GPCR, phosphorylating it. Next, arrestin protein binds to it - preventing GPCR binding to G-protein. Arrestin act as adpator proteins, recruiting the phosphorylated receptor to clathrin coated pits leading to endocytosis via degradation using lysosomes.

Question 7

There are _ different a subunits. There are _ different B subunits. There are __ different gamma subunits. ____ possible G-proteins. How many different effector possibilities can you get?

Answer 7

20, 5, 12

1200

20 alpha + 60 (5x12) beta-gamma different effector possibilites - alpha acts as one effectors and beta-gamma acts as another.

Question 8

3 Ways Metabotropic Receptors Regulate Ion channels

Answer 8

1. Coupling by G-protein directly to an ion channel (E.g. muscarinic receptors)
2. Coupling by G-protein to a second messenger system, where the 2nd messenger (eg cAMP, cGMP) directly regulates an ion channel (E.g. cAMP gated ion channels)
3. Coupling by G-protein to a second messenger system, leading to ion channel phosphorylation (e.g. activating PKA which phosphorylates ion channels)

Question 9

_ types of muscarinic receptors

Answer 9

5

Question 10

Gαs, Gαi, Gαq, Gαt

What do they stimulate or inhibit? Qαq in detail?

Answer 10

• Gαs stimulates Adenylyl Cyclase -> cAMP -> Protein Kinases -> Phosphorylation of Target Protein
• Gαi inhibits Adenylyl Cyclase (sensitive to pertussis toxin)
• Gαq stimulates Phospholipase C -> cleaves PIP2 into DAG and IP3 -> both activate PKC -> Phosphorylation of Target protein -> Cellular response

a) DAG binds to PKC
b) IP3 binds to Ca2+ ion channel allows Ca2+ flow into cytosol from ER -> Ca2+ binds to PKC

• Gαt stimulates cGMP Phosphodiesterase, reducing cGMP levels

Question 11

Why are there 1200 different G-proteins?

Answer 11

Due to Signalsomes: preformed

scaffoldings in which GPCR, G-protein, effector and regulatory molecules are locked into position

Question 12

How are GPCR receptor signals switched off?

Answer 12

Detachment of ligand -> hydrolysis of GTP -> removal of 2nd messengers (may be labile (changed), removed, destroyed) -> activation of phosphotases (undo the work of whatever kinases were activated, so that protein activity, channel conductance etc all return to their pre-stimulus level)

*Turning off signals is more complex

Question 13

PKA - does not only phosphorylate ion channels but also?

Answer 13

GPCR kinase - phosphorylating GPCR so arrestin will bind to it

Question 14

Peptide NT are transported along _____ to ____ via ____

Answer 14

Transported along microtubules to axon terminals via vesicles.

*ALL peptide NT are metabotropic

Question 15

List 4 peptide NT

Answer 15

• Substance P (pain transmission)
• Neuropeptide Y (blood pressure, appetite stimulation)
• Angiotensin II (thirst and salt appetite)
• Endorphins (pleasure, reward, inhibition of pain)

Question 16

Are peptide NT recycled?

Answer 16

They are not recycled at the nerve terminal. The fused vesicular membrane is endocytocised and transported back to the cell body. There is no re-up take of peptide NT - they undergo proteolytic cleavage after release.

Question 17

Peptide NT - larger or smaller than A.acid? Light or dark stain under electron microscopy?

Answer 17

Larger than A.acid NT, dark-stained

Question 18

Autoreceptors – where is it found and how does it work?

Answer 18

-Presynaptic receptor
e.g. Dapamagernic terminal but also found in Ach, glutamatergic and etc.
Form of negative feedback that reduces amount of dopamine released
Dopamine binds to autoreceptor -> Inhibits VACC (Voltage Activated Ca2+ Channel) -> decreases amount of Ca2+ flowing into the terminal -> Inhibit release of NT

Question 19

Non-synaptic Receptors

Answer 19

An example of neuromodulatory action, but located outside of the synapse altogether.

Question 20

Neuromodulators

Answer 20

They alter the excitability of the post-synaptic neuron, but don’t regulate the membrane voltage directly

Question 21

Another name for vesicles containing peptide transmitters? Size? Dense or not under electron microscope? What else do they contain?

Answer 21

LDCV - Large dense core vesicles

• 200-400 nm in diameter
• dense under the electron microscope
• they contain proteins which act as a scaffolding
• they contain peptidases which may act after release

Question 22

Some metabotropic synapses have different morphology

Answer 22

• More spread out vs. ionotropic receptors which are clustered tightly with the PSD
• PSDs are less dense
• Some receptors lie outside the PSD

Question 23

T/F: Kinases phosphorylate GPCR without a ligand bound to it

Answer 23

False – they only phosphorylate ligand bound GPCRs

Question 24

Tachyphylaxis

Answer 24

Acute rapid decrease in response to a drug after its administration. This concept is present in GPCRs when GPCRs are internalized and removed after prolonged stimulation and hence drug won’t be as effective anymore.

Question 25

Is it the receptor or transmitter that determines the nature of transmission?

Answer 25

The receptor – you can have transmitters that activate both ionotropic and metabotropic receptors

Question 26

Metabotropic transmitter – GABA A or GABA B?

Answer 26

GABA B

Question 27

2 different responses from dopamine

Answer 27

Initiating movement or feeling pleasure

Question 28

2 different areas of the brain Ach stimulates

Answer 28

Pons – sleep and wakefulness

Forebrain – Learning, memory, attention

Question 29

Which area of the brain is Serotonin found?

Answer 29

Raphe nuclei (in medulla, pons and midbrain)
*Appetite, libido, sleep, mood, pain

Question 30

Which area of the brain is NA system found in?

Answer 30

Locus coreleus (pons)
*Mood, vigilance, pain

Question 31

The major neuromodulatory systems – dopamine, ach, serotonin, NA. Most is found in brainstem but one is found in the basal forebrain?

Answer 31

One type of Ach

Question 32

What does the ascending reticular activation system consist of? What does it control?

Answer 32

1) Serotoninergic raphe nuclei (of which there are 9)
2) Cholinergic system of the pons
3) Noradrenergic locus coeruleus

This system controls sleep wake cycles (including REM), arousal and vigilance

Question 33

The Cholinergic Neuromodulatory System in the Basal Forebrain – What structures are present? Where do these structures innervate?

Answer 33

It consists of several contiguous groups (nuclei) of magnocellular cholinergic neurons. From rostral to caudal, these are

1) Septum – innervates hippocampus aka Septo Hippocampus system (traverse via a bundle of axons aka fornix to innervate it)
2) Diagonal Band (of Broca) – innervates Olfactory cortex
3) Basal Nucleus (of Meynert) aka Meynert Band – innervates entire neocortex (from frontal, occipital to parietal cortex)

Septum and hippocampus used to be neighbours but grew apart with the development of the cerebral cortex. Fornix traces evolutionary separation between the two structures.

Question 34

Where are the only cells responsive to NGF (Nerve Growth Factor) found? This are is also the most severely and early affected area in Alzheimer’s Disease

Answer 34

In the cholinergic neuromodulatory system

Question 35

One early symptom of Alzheimer’s disease?

Answer 35

Loss of smell

Question 36

Most important signalling cells of hippocampus

Answer 36

CA3, CA2, CA1 pyramidal neurons – have Ach muscarinic R

-Other neurons are usually inhibitory interneurons

Question 37

What types of neurons and receptors are present on the hippocampus?

Answer 37

• Pyramidal neurons (glutamatergic)
- m1 receptors (excitatory)
• Interneurons (GABAergic)
- m2 receptors (inhibitory)
• Autoreceptors
- m4 (inhibit release)

Question 38

M1-5, which ones are excitatory and which ones are inhibitory?

Answer 38

M1, 3, 5 – excitatory, activates Phospholipase C -> IP3 -> Ca2+ -> Excitation
M2, 4 – inhibitory, inhibit Adenylyl Cyclase -> Decrease cAMP -> decrease excitability (post-synaptic) -> decrease release (pre-synaptic)

Question 39

Activation of M1ACh receptor in hippocampus – what is its effect on M type K+ channels? Effect on Na+ channels?

Answer 39

• Slight depol
• Inhibits M type K+ channels (Beta-gamma) which allows pyramidal neuron to discharge at high frequency, maximising effect of hippocampus
• Activates pacemaker sodium channels (increases oscillatory behaviour of membrane potential) -> rhythmic action potential discharges

Question 40

90% of cholinergic release sites in the neocortex and hippocampus are not ____ but Ach is released from ______ into the ___ Cholinergic axons form many branches in the target tissue, and each branch has varicosities spaced about __ microns apart – ranging from _ to _ release sites

Answer 40

Synaptic, varicosities, ECF, 2.5, 200 K, 500 K

Question 41

What happens when hippocampus loses cholinergic input?

Answer 41

Hippocampal pyramidal neurons become hypo-excitable – impairing memory function (especially spatial and anterograde memory e.g. formation of new long term memories)

Question 42

What happens to Cholinergic Neurons after Fornix axotomy? What happens to cholinergic synapses in hippocampus? i.e. cutting the connections between forebrain cholinergic neurons and their target cells in the hippocampus

Answer 42

• Neurons shrink
• Synthesis of Ach decreases -> synthesis of proteins such as choline acetyltransferase and choline transporters also decrease
• Neurons lose their cholinergic phenotype, neuronal phenotype, and even undergoing apoptosis
• IN HIPPOCAMPUS: Cholinergic synapses retract and die

Question 43

Describe the two-way signal flow of the CA1 Pyramidal neurons in the Hippocampus

Answer 43

Hippocamus depends on cholinergic neurons to function

Cholinergic neurons have NGF receptors TrkA and p75, and need NGF from pyramidal neurons to make Ach.