5. Receptors

Question 1

What are ligands and their TWO subtypes?

Answer 1

Ligands are small molecules which BIND SPECIFICALLY to a RECEPTOR site.

Two ligand subtypes:

a) AGONISTS - activate receptor
b) ANTAGONISTS - don’t activate receptor

Question 2

What are partial agonists and how may they affect full agonists?

Answer 2

Partial agonists are ligands which activate receptors WITHOUT causing MAXIMUM CELL RESPONSE.

These can act as ANTAGONISTS to full agonists (preventing them from binding and causing a greater response).

Question 3

What are acceptors?

Answer 3

Acceptors are molecules whose function is MODIFIED (‘regulated’) by binding of small chemicals but, unlike receptors, their basic function can be carried out without this interaction.

i.e. Acceptors are NOT functionally silent when unbound

Question 4

How are receptors classified?

Answer 4

1. CLASSIFICATION by AGONIST
   (e. g. nicotinic or muscarinic)
2. SUBCLASSIFICATION by STRENGTH OF ANTAGONIST BINDING
   (e. g. M1, M2, M3)

Question 5

Give TWO differences between receptor binding sites and enzyme active sites?

Answer 5

1. HIGHER AFFINITY of ligand binding at receptors than enzyme substrates (or regulatory molecules).
   - Dissociation Constant (Kd) for receptors is nano/micromolar
   - Michaelis Constant (Km) for enzymes is micro/millimolar
2. NO CHEMICAL MODIFICATION of ligands at receptor sites whereas enzyme substrates are modified (note: enzyme regulatory molecules are not modified)

Question 6

Give SIX uses of receptors in cell physiology.

Answer 6

1. SIGNALLING (hormones or local mediators)
2. NEUROTRANSMISSION (synaptic)
3. CELLULAR DELIVERY (LDLs, transferrin)
4. Control of GENE EXPRESSION (thyroid or steroid hormones)
5. Release of CALCIUM FROM STORES (IP3)
6. IMMUNE RESPONSES

Question 7

What are the FOUR types of signal transduction?

Answer 7

1. Receptors are INTEGRAL ION CHANNELS
2. Receptors have INTEGRAL ENZYME ACTIVITY
3. G PROTEIN COUPLED RECEPTORS
4. INTRACELLULAR RECEPTORS

Question 8

Insulin and growth factor receptors exhibit integral enzyme activity. How do they work?

Answer 8

Insulin and growth factor receptors are TYROSINE KINASE LINKED.

Binding of the ligand extracellularly causes:

1. AUTOPHOSPHORYLATION - each subunit phosphorylates the other.
2. PHOSPHOTYROSINE RECOGNITION by either:
   a) Transducing protein (e.g. insulin receptor substrate 1)
   b) Src Homology 2 domains directly
3. Either way, enzymes with SH2 are activated.

Question 9

How do intracellular receptors work?

Answer 9

Intracellular receptors respond to hydrophobic ligands. These receptors are stabilised at rest by chaperones called an INHIBITORY PROTEIN COMPLEX which dissociates upon ligand binding.

This allows its DNA BINDING SITE to attach to control regions of DNA.

Question 10

What are receptors?

Answer 10

Receptors are molecules which SPECIFICALLY recognise a LIGAND or family of ligands and in response to BINDING a ligand, bring about REGULATION of a cellular process.

Receptors must be FUNCTIONALLY SILENT when unbound.

Question 11

What are G-Proteins?

Answer 11

G-Proteins are GUANINE NUCLEOTIDE BINDING PROTEINS. These interact with a superfamily of GPCRs or seven transmembrane receptors and facilitate signal transduction.

G-Proteins are heterotrimeric:

1. Alpha (a) subunit
2. Beta (B) subunit
3. Gamma (y) subunit

Alpha (a) subunit binds GTP and hydrolyses it to GDP.
It connects the other subunits (By) with the GPCR

Beta (B) and Gamma (y) subunits function as a single unit.

Question 12

Describe the sequence of events which occurs as a ligand interacts with a GPCR.

Answer 12

1. BASAL CONDITIONS
   (G Protein with associated GDP, is on the inner face of the membrane)
2. ACTIVATED GPCR - agonist bound
   (GPCR has high affinity for GDP-G Protein)
3. GDP-GTP EXCHANGE - as G Protein interacts with GPCR
4. DISSOCIATION of Ga-GTP and GBy from GPCR
   (once GTP binds the Ga has low affinity for GBy and the receptor)
5. INTERACTION WITH EFFECTORS
6. TERMINATION of effector interaction - intrinsic GTPase activity
   (GDP-Ga and GBy reform heterotrimer assuming basal position)

Question 13

What are TWO characteristics of GPCRs?

Answer 13

1. ON/OFF SWITCH - facilitated by GDP-GTP exchange
2. TIMER FUNCTION - facilitated by GTPase activity of Ga (RGS proteins increase GTPase activity thus shorten the ‘timer’.

Question 14

Which G Proteins are different cholinergic and noradrenergic GPCRs coupled to?

Answer 14

(a) Noradrenergic ‘QISS’

Q: Gq - Alpha1
I: Gi - Alpha2
S: Gs - Beta1
S: Gs - Beta2

b) Cholinergic (muscarinic) ‘QIQ’

Q: Gq - M1
I: Gi - M2
Q: Gq - M3

Question 15

How does visual excitation occurs in rod cells of the retina?

Answer 15

1. LIGHT acts on RHODOPSIN (a receptor linked to Gt)
2. TRANSDUCIN (Gt) activates CYCLIC GMP PHOSPHODIESTERASE
3. Cyclic GMP Phosphodiesterase DEACTIVATES CYCLIC GMP (second messenger) by converting it to 5’GMP
4. MEMBRANE HYPERPOLARISATION as cGMP operated in channel (Na+ and Ca2+) is closed.
5. Hence VISUAL EXCITATION

Question 16

Which TWO well-known toxins which exhibit ADP-Ribosyltransferase activity and what does this do?

Answer 16

ADP-Ribosyltransferase activity refers to the ability to covalently modify a G protein (alpha subunit) with an ADP-RIBOSYL group which affects its function.

a) CHOLERA TOXIN - modifies Gs to become ALWAYS ACTIVE
(inhibits GTPase activity)

b) PERTUSSIS TOXIN (whooping cough) - modifies Gi and INHIBITS
(prevents GDP-GTP exchange)

Question 17

What is adenylyl cyclase?

Answer 17

Adenylyl Cyclase is an integral plasma membrane protein which acts as a SECOND MESSENGER GENERATOR.

It is activated by Gs and inhibited by Gi.

When activated, adenylyl cyclase generates CYCLIC AMP. This is a second messenger which activates PROTEIN KINASE A.

Question 18

What is phospholipase C?

Answer 18

Phospholipase C is an integral plasma membrane protein which acts as a SECOND MESSENGER GENERATOR.

It is activated by Gq and upon activation, phospholipase C converts the minor membrane phospholipid, PIP2, into two second messengers:

a) Inositol 1,4,5 Triphosphate (IP3)
- Ca2+ release from ER
- Thus activation of Ca2+ SENSITIVE PROTEIN KINASES

b) Diacylglycerol (DAG)
- Activation of PROTEIN KINASE C family

Question 19

After agonist binding and activation of a GPCR pathway, give FIVE ways deactivation occurs.

Answer 19

1. AGONIST RECEPTOR DISSOCIATION - G Protein interaction weakens agonist binding.
2. RECEPTOR DESENSITISATION - activated GPCRs susceptible to phosphorylation by GPCR KINASES (GRKs) which prevents G protein interaction.
3. ALPHA GTP LIFETIME - GTPase activity (affected by RGS proteins)
4. SECOND MESSENGER METABOLISM
   a) PHOSPHODIESTERASES - degrades cAMP to 5’AMP
   b) 5’ PHOSPHATASE - degrades IP3 to IP2
5. OPPOSITION TO CASCADE - proteins downstream of second messenger or protein kinase oppose their effects.
   (e. g. protein phosphatase)

Question 20

With reference to GPCRs, how is chronotropy in the heart regulated?

Answer 20

POSITIVE CHRONOTROPY

1. Noradrenaline acts on B1 adrenoceptors at the SAN
2. Gs linked therefore increase in adenylyl cyclase activity
3. Increased cAMP produced
4. Increased activity of HCN channels (‘funny current’ channels)
5. Increased slope of pacemaker potential so raised heart rate

NEGATIVE CHRONOTROPY

1. Acetylcholine acts on M2 receptors at SAN
2. Gi linked: a-GTP and By act on K+ channels
3. Increased opening of K+ channels causes hyperpolarisation
4. Lowers heart rate (lowers slope of pacemaker potential)

Question 21

With reference to GPCRs, how is inotropy in the heart regulated?

Answer 21

Cardiac ventricles only have B1 adrenoceptors in abundance.

POSITIVE INOTROPY

1. Noradrenaline binds to B1 adrenoceptors at cardiac ventricles
2. Gs linked therefore increased activity of adenylyl cyclase
3. Increased cAMP generated
4. cAMP activated Protein Kinase A
5. PKA and a-GTP both interact with and activate VOCCs
6. Increased magnitude of Ca2+ entry, increases force of contraction

Question 22

With reference to GPCRs, how does arteriolar vasoconstriction occur?

Answer 22

ARTERIOLAR VASOCONSTRICTION

1. Noradrenaline acts on a1 adrenoceptors in arterioles
2. Gq linked therefore increased activity of phospholipase C
3. Increased production of second messengers: IP3 and DAG
4. IP3 increases Ca2+ release from SR (increases contraction)
5. DAG activates Protein Kinase C which phosphorylates key proteins required for contraction.

Question 23

u-Opioid receptors are presynaptic GPCRs which modulate neurotransmitter release. How do such receptors work?

Answer 23

Neurotransmitter release is governed by Ca2+ entry at the presynaptic knob due to depolarisation. Therefore to inhibit neurotransmitter release, Ca2+ entry can be reduced.

Presynaptic (u-opioid) receptors bind endogenous opioids or analgesics which result in liberation of GBy subunits. These interact with VOCCs and reduce Ca2+ entry thus inhibit neurotransmitter release.

Question 24

What are TWO possible consequences of excessive agonist binding?

Answer 24

Excessive agonist binding leads to cells becoming LESS RESPONSIVE (TACHYPHYLAXIS) although this can occur in one of two ways:

a) DESENSITISATION and UPREGULATION - receptors are less responsive but there are more of them thus more agonist is required for the same response.

There is a change in RECEPTOR PROPERTIES:

• Conformational change to desensitised form
• Phosphorylation by GRKs
• Inhibitor Protein Binding e.g. Arrestin

b) DOWN REGULATION - internalisation by RME either into an intracellular pool (reversible) or degradation (irreversible).

Question 25

What is the difference between homologous and heterologous desensitisation?

Answer 25

HOMOLOGOUS desensitisation refers to desensitisation or down regulation of the receptor which has been stimulated by excessive agonist.

HETEROLOGOUS desensitisation refers to desentisation or down regulation of other receptor i.e. receptors which haven’t been stimulated by excessive agonist.

• It requires NEGATIVE FEEDBACK of a desensitised or down regulated receptor onto a signalling pathway COMMON to other GPCRs.

Question 26

What happens to receptors which have been deprived of agonist or supplied with excess antagonist?

Answer 26

Excess antagonist or agonist deprivation causes SUPRASENSITIVITY.

This involves UPREGULATION of a receptor and is an adaptive response to overcome effects of antagonism thus there are more receptors available to the natural agonist.