- Cholinergic receptors
- Adrenergic receptors
- Histamine receptors
- Dopamine receptors
- 5 Hydroxytryptamine (Serotonin) receptors
- Opioid receptors
- GABA receptors
Welcome to ePharmacology! Today we will discuss the following topics:
Life cycle of receptors
Receptor synthesis occurs in the endoplasmic reticulum and then pass through the Golgi system.
Golgi vesicles containing receptors fuse with the plasma membrane, thereby inserting the receptor into the mobile lipid domain of the cell surface.
So receptor synthesis is just like any other protein synthesis!
The fate of a receptor after exposure to agonist
The fate of a receptor after exposure to agonist is only known for some of the receptors like transferrin, peptide hormones, insulin.
There is receptor-mediated endocytosis. When an agonist binds to the receptor, the receptor becomes clustered in specialized depressions in the cell membrane called coated pits.
The inner membrane surface of the coated pit has an electron-dense coating made up of a protein called clathrin that is detectable by electron microscope. These coated pits (half-life of about 1 minute) then pinches off from the cell surface to form coated vesicles.
Coated vesicles then get uncoated by shedding off their clathrin coats which is due to the effect of clathrin-depolymerizing enzyme. Then they fuse with one another to form larger vesicles called endosomes (receptosomes).
Endosomes are uncoated vesicles (average diameter of 0.5 μm) and maintains a pH of 5.5 within it by an ATP-dependent proton pump. Some endosomes are recycled for cell surface receptor after fusion with the Golgi system and others are degraded in lysosome. For example, LDL receptor can go through 150 such cycles without losing its function. Transferrin receptor and iron are recycled. On the contrary, both epidermal growth factor and its receptor are degraded within the acidic endosome.
The loss in response has been most commonly termed desensitization. When this loss in response is very rapid, then it is termed tachyphylaxis. Nicotinic cholinergic receptor undergoes very rapid desensitization (within 1 second), whereas G protein or tyrosine coupled receptor desensitizes over many seconds to minutes or even hours.
A number of receptors in the plasma membrane regulate distinct effector proteins through mediation of a group of GTP (guanosine triphosphate) binding proteins known as G proteins.
The activity of this regulatory protein depends on the presence of GTP and magnesium.
G protein acts as an intermediate between receptor and enzyme (an effector).
G protein is located on the inner surface of the plasma membrane. It has subunits designated as α, β and γ.
The G proteins have three domains: guanine nucleotide binding domain, domain for interaction with receptor and effector.
VARIETIES OF G α (ΑLPHA) PROTEIN
There are several types of Gα proteins- Gαs , Gαi, Gαo , Gαq , Gαt and more.
Gαs(s for stimulatory) stimulates adenylyl cyclase after being activated by an agonist. The same G protein also activates calcium channel.
Gαi (i for inhibitory) inhibits the adenylyl cyclase activities and activates potassium channel.
Gαo inhibits calcium channel whereas Gαq activates phospholipase C.
Gαt designating transducin, mediates rhodopsin activation of cyclic GMP phosphodiesterase.
Several G proteins may be present in a single cell. Each of these may respond to several different receptors and regulate several different effectors. One receptor can also regulate more than one G protein.
Function of G alpha proteins
|G alpha protein||Function|
Inhibits adenylyl cyclase, Activates potassium channel
Inhibits calcium channel
Activates phospholipase C
Stimulates adenylyl cyclase present in eye
Activates adenylyl cyclase, Activates calcium channel
ACTIVATION / INACTIVATION CYCLE OF G PROTEIN
In resting (inactive) state of G protein, GDP (guanosine diphosphate) is tightly bound to the α subunit. But when an agonist is bound to G protein-coupled receptor then the GDP bound to the α subunit gets replaced by GTP.
This α-GTP is then dissociated from the β and γ subunits and subsequently interact with the membrane bound effector(such as adenylyl cyclase). The GTPase activity of the α subunit increases on binding, leading to hydrolysis of the bound GTP to GDP which allows the α subunit to recombine with the βγ complex.
Second messenger system
When first messenger (ligand) binds with its specific receptor, the drug-receptor complex is formed which subsequently causes the synthesis and release of another intracellular regulatory molecule called second messenger. These are:
- Adenosine 3’-5' monophosphate (cyclic AMP; cAMP)
- Guanosine 3‘-5' monophosphate (cyclic GMP; cGMP)
- Inositol 1,4,5-triphosphate (IP3)
- Diacylglycerol (DAG)
- Calmodulin (CaM).
cAMP AS SECOND MESSENGER
The first recognized second messenger cAMP is synthesized by the plasma membrane attached enzyme adenylyl cyclase in response to activation of many receptors such as β-adrenergic receptors. The function of activated adenylyl cyclase is to convert ATP into cAMP.
Binding of the agonist to α2-adrenergic receptors, M2 receptors leads to inhibition of cAMP formation within the cell.
Normally cAMP is hydrolysed within the cell by an enzyme phosphodiesterase which is inhibited by drugs like caffeine, theophylline. So, there is increased intracellular concentration of cAMP following ingestion of drugs containing caffeine and theophylline and thus more activity.
Adenylyl cyclase can also be activated directly (by-passing the receptor) by some drugs like forskolin and fluoride ions.
cAMP acts exclusively through cAMP-dependent protein kinase (A-kinase) to phosphorylate enzymes and proteins involved in cell function.
A-kinase is composed of two regulatory (R) and two catalytic (C) subunits. When cAMP binds to the regulatory subunits, there is dissociation of the regulatory subunits with resultant activation of the catalytic subunits. There is transfer of phosphate (phosphorylation) from ATP to various cellular proteins.
cAMP mediates the responses such as the rate and contraction force of heart muscle, the relaxation of smooth muscle, the breakdown of carbohydrates in liver, the breakdown of triglycerides in fat cells, calcium homeostasis and many other endocrine and neural processes.
CALCIUM as second messenger
Intracellular calcium plays an important role in the function of most of the cells.
Intracellular calcium is present in both free and bound forms. It is the free form that is responsible for the cell function. There is a great variation in the concentration of free calcium in extra and intracellular compartment. Recently, great achievement has been obtained to measure intracellular free calcium concentration before, during and after stimulation by a drug in intact cell using the fluorescent dye Quin 2 or Fura 2.
The concentration of free calcium outside the cell is in millimolar range whereas in resting state the intracellular free calcium concentration is around 100 nanomolar, i.e. its concentration is 10,000 times less in the intracellular compartment. When the cells are stimulated by a full agonist, intracellular free calcium concentration increases rapidly, i.e. the concentration of free calcium will be increased from 100 nanomolar to about 500 nanomolar. This increased free calcium concentration is responsible for the effect. The bound form of calcium is present in millimolar concentration in the inner face of the plasma membrane, endoplasmic reticulum, mitochondria, and secretory granules.
cGMP as second messenger
The second messenger cGMP is produced from the GTP by an enzyme guanylyl cyclase which is present in the inner phase of the plasma membrane.
The guanylyl cyclase is activated when muscarinic receptor is occupied by its agonist. This cGMP then activates the intracellular cGMP-dependent protein kinase (G-kinase). The subsequent G-kinase mediated effect is not yet known.
INOSITOL 1,4,5 PHOSPHATE (IP3) as second messenger
Recently IP3 has been well accepted as a second messenger. It is the hydrolytic product of phosphatidylinositol (PI). PI is the minor phospholipid of the cell membrane. Phosphorylation of PI causes the formation of phosphatidylinositol monophosphate (PIP) which is later converted to phosphatidylinositol 4,5-biphosphate (PIP2). The activation of enzyme phospholipase C (a membrane-bound enzyme) causes the hydrolysis of phosphatidylinositol 4,5-biphosphate (PIP2) and there is formation of water soluble Inositol 1,4,5 phosphate (IP3) and Diacylglycerol (DAG). This IP3 is released into the cytoplasm and stimulates the release of calcium from endoplasmic reticulum. So, IP3 increases the intracellular free calcium concentration which ultimately involves in producing the effect. This IP3 is then dephosphorylated to inositol 1,4 phosphate (IP2), inositol 1 phosphate (IP), inositol and finally to phosphatidylinositol.
The antipsychotic drug lithium causes depletion of membrane PI and accumulation of intracellular IP by inhibiting the hydrolysis of IP to inositol.
DIACYLGLYCEROL as second messenger
Another second messenger diacylglycerol (DAG) is produced in the cell membrane from the metabolic product of PIP2. This DAG activates directly the intracellularly located protein kinase C (C- kinase). DAG is phosphorylated to form phosphatidic acid coupled with IP to form PI.
CALMODULIN as second messenger
Calmodulin (CaM) is a single peptide chain containing 148 amino acid residues and is considered as the receptor for intracellular free calcium. It has four binding sites. Three or four of these need to be occupied by calcium before CaM will activate myosin light chain kinase (MLCK). One molecule of calcium-CaM interact with one of MLCK and without this interaction MLCK is inactive. While phosphorylated, myosin forms cross bridges with actin and sliding of actin over myosin filaments occur. This sliding effect produces contraction of muscle.
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