Viva C(iv)b

This viva is relevant to the objectives of Section C(iv) from the 2017 CICM Primary Syllabus, which expects the exam candidate to "explain receptor activity with regard to... second messengers and G proteins". 

What is a "second messenger"? Define the term.

A second messenger as an intermediate molecule for an intracellular signal transduction cascade, which is used to transmit and amplify the signal between an extracellular stimulus and an intracellular effector.

Possible valid answers include:

" intermediary molecule that is generated as a consequence of hormone receptor interaction"

"...a chemical substance inside a cell that carries information farther along the signal pathway from the internal part of a membrane-spanning receptor embedded in the cell membrane"

"...any of various intracellular chemical substances...that transmit and amplify the messages delivered by a first messenger to specific receptors on the cell surface"

Explain how a second messenger system works. How does drug-receptor binding produce a response via a second messenger system?
  • he drug-receptor or receptor-ligand interaction often does not result in the direct activation of the intracellular effector
  • Instead, often an intermediate molecule is used as a signal to the effector.
  • This intermediate molecule is synthesised or released in response to the receptor-ligand interaction, and then degraded afterwards. 
  • The rate of synthesis and degradation of this molecule is tightly regulated to control the magnitude of response to receptor activation, and this regulation can be used to amplify or dampen the response.
  • The second messenger molecule can act locally, or can diffuse distally to convey the signal to a multitude of targets; and multiple second messenger systems can interact to produce complex responses to receptor-ligand binding.
What are some examples of second messenger systems?

It is possible to classify the second messenger systems into several broad types:

  • Hydrophobic molecules, such as DAG and phosphatidylinositols which do most of their work from the intermembrane space
  • Hydrophilic molecules such as  cAMP, cGMP and IP3 - which diffuse freely in the cytosol
  • Ions such as ionised calcium, potassium and sodium
  • Gases, such as nitric oxide (NO) and carbon monoxide (CO) which diffuse easily through lipid and water alike.
  • Soluble proteins such as Jak/STAT, NF-kB, etc
What is cyclic AMP?

cAMP is a cyclic nucleotide. Nucleotides are composed of three main components:

  • a phosphate group (or two, or three),
  • a sugar (classically a pentose sugar such as ribose), and
  • a nitrogenous nucleobase (in the case of cAMP, adenine).

The "cyclicness" is conveyed upon straight AMP by the bonds created between the phosphate group and the hydroxyl groups on the sugar, creating a little ring. 

cAMP has a very short half-life, about 1 minute.

How is cyclic AMP synthesised?
  • cAMP is generated out of ATP by adenylyl cyclase; it converts ATP to cyclic AMP
  • It is a transmembrane protein modulated by G-protein coupled receptors (i.e. they can increase or decrease its activity).
  • It is activated by Gs proteins, and deactivated by Gi proteins.
How is cyclic AMP degraded and regulated?
  • Phosphodiesterase converts cAMP back into "straight" AMP.
  • There are multiple species of phosphodiesterase.
  • PDE 1, 2, 3, 10 and 11 hydrolyse both cAMP and cGMP, whereas PDE 5 6 and 9 specifically hydrolyse cGMP only. Thus by targeting only PDE 5, sildenafil only inhibits the degradation of cGMP.
How does cyclic AMP produce intracellular effects?

Its effectors are:

  • PKA (which phosphorylates numerous metabolic enzymes)
  • EPAC (a guanine-nucleotide-exchange factor) which is involved in all sorts of things like cell adhesion, exocytosis, apoptosis, and gene expression
  • cyclic-nucleotide-gated ion channels (usually these are unselective cation channels; one example is present in human rod and cone cells)
What are the effects of increased intracellular cyclic AMP?

The observable effects include:

  • mobilization of stored energy eg. β-adrenergic lipolysis / glycogenolysis
  • vasopressin-mediated water retention
  • parathyroid-hormone mediated calcium homeostasis
  • response to catecholamines (β-adrenergic)
 What is cyclic GMP? How is it synthesised?
  • Cyclic guanosine monophosphate is similar to cyclic AMP, with the exception that instead of adenine cGMP has a guanine nucleobase.
  • It is derived from GTP (guanosine triphosphate).
  • There are two major pathways of its synthesis, one via a membrane-bound guanylyl cyclase bound to a natriuretic peptide receptor, and the other a soluble guanylyl cyclase which is activated by nitric oxide. 
What are the effects of increased cyclic GMP?

The relevance of cGMP to cardiovascular physiology and pharmacology is mainly related to its activation by nitric oxide, and  the relevance of this is mainly to vascular smooth muscle.

  • Something (anything) activates nitric oxide synthase
  • Nitric oxide (NO) is synthesised
  • NO activates the soluble form of guanylyl cyclase at nanomolar concentrations
  • Guanylyl cyclase produces cGMP
  • cGMP activates Protein Kinase G (PKG)
  • The 1β isoform of PKG phosphorylates the IP3 receptor, inhibiting IP3-mediated release of calcium
  • PKG also phosphorylates and thus inactivates voltage-gated calcium channels.
  • PKG also phosphorylates phospholamban, leading to increased calcium ion uptake into sarcoplasmic reticulum

The net effect of these changes is a decrease in the availability of intracellular calcium, and therefore smooth muscle relaxation.

cGMP also has several other effects:

From Protein Kinase G activation

  • Negative inotropic effect, by reduction of myofilament calcium responsiveness
  • Promotion of angiogenesis

From cGNP-gated ion channels

  • mainly unselective cation channels, relevant to the movement of sodium and calcium ions (Kaupp et al, 2002) - these are mainly expressed in retinal and olfactory neuroepithelium and in nephrons (i.e. these channels have no relevance to the cardiovascular system).

From cGMP-modulated phosphodiesterase 

  • cGMP can bind to phosphodiesterases which increases their activity against both cGMP and cAMP, resulting in the inhibition of both secondary messenger systems. 
How does calcium act as a second messenger?
  • Intracellular calcium concentration is kept at a nanomolar level- generaly some decimal fractions of a nanomole.
  • Calcium is actively pumped out of the cell, and actively sequestered in intracellular stores.
  • The entry of calcium into the cell is usually mediated by the actions of phospholipase C. 
  • Phospholipase C (β) is activated by G-protein-coupled receptors, whereas phospholipase C (γ) is activated by the tyrosine kinase pathway.
  • Either way, the result is the hydrolysis of a membrane phospholipid PIP2, phosphatidylinositol 4,5-bisphosphate. 
  • The results of the hydrolysis is production of inositol triphosphate (IP3) and diacylglycerol (DAG), i.e. both molecules are produced by this reaction.
  • IP3 and DAG then go on to have distal effects. 
What are the downstream effects of inositol triphosphate release?

IP3 opens an IP3-gated calcium channel, allowing calcium into the cell.

Calcium release causes activation of calmodulin-sensitive enzymes:

  • Phosphodiesterase (yes, the one that degrades cAMP and cGMP)
  • Myosin light chain Kinase which allows muscle contraction to begin by enabling the formation of the actin-myosin crossbridge
  • Calmodulin-dependent protein kinase (I and II) involved in calcium homeostasis, long term memory formation, CD-8 T-cell activation, chloride transport in epithelia, neuronal transmission, cilia motility, apoptosis, regulation of various enzymes, etc etc
What are the downstream effects of diacylglycerol release? Give examples.

It in turn activates Protein Kinase C. This thing has multitudes of effects.

  • α-1 adrenoreceptor mediated contraction of smooth muscle
  • α-1 adrenoceptor mediated ejaculation
  • Prostaglandin and thromboxane mediated contraction of smooth muscle
  • Muscarinic M3 receptor mediated contraction of smooth muscle 


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