Intro to CNS Pharmacology

Question 1

How have proteins evolved in function to allow ionic passage across the membrane?

Answer 1

1. ATPase driven pumps
2. Transporters
3. Ion channels

Question 2

Ion channels: some generalities

Answer 2

1. Integral membrane proteins
2. Multiple membrane-spanning domains
3. Form a hydrophobic channel in the center
4. Selective for ions and regulated by changes in the cellular environment
5. Multiple gene products; multiple subunits
6. Glycosylated on the extracellular side
7. Consensus sequences for kinases
8. Exhibit specificity for the ion(s) that permeates the channel
9. Ionic movement is driven by its electrochemical gradient

Question 3

Functional classification of ion channels based upon the gating mechanism:

Answer 3

1. Passive: non-gated, always open
2. Active: are gated
   
   • i.e. the closed and open states of the channel are regulated

Question 4

Some types of gating include:

Answer 4

• membrane potential difference (voltage gated)
• small extracellular molecules (i.e. neurotransmitters)
• other membrane proteins
  
  • e.g. beta-gamma subunits of G proteins
• Intracellular molecules
  
  • e.g. ions, ATP

Question 5

What is a leak channel?

Answer 5

channel is open at resting membrane potential

• Can be either active or passive
• All passive channels are leak channels

Question 6

What is the resting membrane potential in neurons?

Answer 6

Em (or Vm) = -60 mV

Question 7

What factors that give rise to the resting membrane potential?

Answer 7

• Intracellular proteins are predominantly anions
• Leak channels are present in the plasma membranes that allow for potassium and chloride movement across the membrane
• Conductance (g) of the membrane to K is 20 times greater than the conductance to Na
• As a result, there is an unequal distribution of Cl, K and Na across the membrane

Question 8

Describe the distribution of Na⁺, K⁺ and Cl^- across the membrane:

Answer 8

1. K⁺:
   
   • high inside and low outside
2. Na⁺ and Cl^-:
   
   • high outside and low inside

Question 9

Nernst potentials:

Answer 9

membrane potentials at which the ion is in
electrochemical equilibrium across the membrane

1. E_K = -75 mV
2. E_Na = +55 mV
3. E_Cl = -69 mV

Question 10

What can oppose the leak channels?

Answer 10

Na-K ATPase pump that moves Na ions
out of the cell and K ions into the cell

Question 11

What causes action potentials in CNS neurons?

Answer 11

voltage operated sodium channels open in the membrane in response to localized depolarization

Question 12

Action Potentials:

Properties

Answer 12

• Since V=IR, increased sodium current results in change in V
• Voltage gated potassium channels also open
  
  • opening is more gradual
  • inactivation is slower than sodium channels
• Action potentials are all or none
• Amplitude of about 100 mV
• 1-10 msec in duration
• Propagated through cycles of depolarization and repolarization

Question 13

Synaptic potentials:

Properties

Answer 13

1. Small, graded potentials that can lead to the initial depolarization that causes an action potential
2. Local
3. Can summate in time and space
4. Only a few mV in size and a few msec in duration

Question 14

Synaptic potentials:

Two Types

Answer 14

1. Excitatory, postsynaptic potential (EPSP)
   
   • membrane potential becomes more positive
   • if it increases enough, threshold will be reached
2. Inhibitory postsynaptic potential (IPSP)
   
   • Membrane potential moves to more negative values
   • Impacts a summating EPSP which will now not reach threshold

Question 15

Two mechanisms by which an EPSP can occur:

Answer 15

1. Increased conductance
   
   • Open a ligand gated ion channel for sodium or calcium
     
     • Nicotinic cholinergic receptor
     • Glutamate receptor
2. Decreased conductance
   
   • Close a leak channel for potassium
     
     • Usually due to changes in the phosphorylation of the channel
     • regulated by second messenger cascades; GPCRs

Question 16

Mechanisms for the production of IPSPs:

Answer 16

1. Increased conductance of the membrane to either potassium or chloride
   
   • Ligand gated chloride channel
     
     • e.g. GABA receptor
2. G protein coupled receptor activation can result in the opening of K channels
   
   • Via direct interactions between the channel protein and G protein
   • As a result of changes in phosphorylation state of closed K channels
     
     • mediated by second messenger cascades

Question 17

Norepinephrine:

CNS Distribution & Physiological Roles

Answer 17

• Noradrenergic neurons are located in the medulla oblongata, pons and midbrain
  
  • Called the reticular activating system
• Very important in arousal (wakefulness) and in regulation of autonomic functions like breathing and blood pressure

Question 18

How is norepinephrine synthesized?

Answer 18

1. Precursor: tyrosine
2. Tyrosine ⇒ 3,4-dihydroxyphenylalanine (DOPA) by tyrosine hydroxylase
3. DOPA is converted into dopamine by DOPA decarboxylase
   
   • low substrate specificity
4. Dopamine ⇒ norepinephrine by dopamine beta hydroxylase
   
   • will oxidize almost any phenyl-ethylamine to the corresponding phenylethanolamine

Question 19

Norepinephrine:

Regulation of synthesis

Answer 19

Primary regulation of norepinephrine synthesis occurs via tyrosine hydroxylase (TH)

1. TH is normally saturated with tyrosine so its activity is the rate limiting step for DOPA synthesis under basal conditions
2. TH has an essential co-factor tetrahydrobiopterine, BH₄
3. Short-term regulation of tyrosine hydroxylase activity occurs via:
   
   • phosphorylation at four different serine residues
   • end-product (i.e. norepinephrine) inhibition of BH4 binding to the enzyme
     
     • detects over-filled vesicles
4. Long-term regulation occurs via new protein synthesis
   
   • primarily increased amounts of tyrosine hydroxylase

Question 20

Norepinephrine:

Regulation of storage and release

Answer 20

1. Norepinephrine is found in vesicles along with the enzyme dopamine betahydroxylase
   
   • Dopamine is taken into the vesicle, the last step of synthesis occurs there
2. Vesicular monoamine transporters are called VMAT; VMAT2 is found in the brain
3. Three mechanisms of norepinephrine release
4. Release is regulated by presynaptic receptors (autoreceptors)

Question 21

Three mechanisms of norepinephrine release:

Answer 21

1. calcium-dependent exocytosis of vesicles
2. reversal of plasma membrane transporters
3. dendritic release that is not calcium-dependent

Question 22

What are the autoreceptors that regulate norepinephrine release?

Answer 22

1. alpha-2 receptor inhibits release
2. beta receptor increases release

Question 23

Norepinephrine:

Regulation of Inactivation

Answer 23

1. All transmitters can be inactivated by diffusion
2. Reuptake by the presynaptic neuron is the most important: neuronal not astrocytic
3. Enzymatic inactivation

Question 24

Describe the reuptake of norepinephrine in the presynaptic neuron:

Answer 24

• High affinity carrier proteins move norepinephrine from extracellular to intracellular compartments
• Energy requiring ⇒ sodium co-transporter
• Binding site for norepinephrine that is the site of action of inhibitors
• Once norepinephrine is intracellular, it can be re-packaged

Question 25

Describe the enzymatic inactivation of norepinephrine:

Answer 25

1. Monoamine oxidase (MAO)
   
   • deaminates norepinephrine
   • two forms: MAOA and MAOB
   • MAOB is found in the brain
2. Catechol-O-methyl-transferase (COMT)
   
   • transfers a methyl group from S-adenosylmethionine to the meta OH of norepinephrine
3. These two can act on the same molecule, in either order

Question 26

Norepinephrine:

Receptors

Answer 26

• All of the receptors for norepinephrine are G protein coupled
• **alpha and beta designation **

Question 27

Norepinephrine:

alpha receptors

Answer 27

bind norepinephrine slightly better than epinephrine

1. alpha 1 subtypes:
   
   • predominately post-synaptic in the periphery
   • tend to function as excitatory receptors:
     
     • increase calcium​
2. alpha 2 subtypes:
   
   • presynaptic, release modulating alpha receptor is alpha2
   • inhibit adenylyl cyclase

Question 28

Norepinephrine:

beta receptors

Answer 28

bind epinephrine better than norepinephrine

1. unlike the alpha receptor subtypes, the beta subtypes are all very similar to each other with regard to function and signal transduction mechanisms
2. All beta receptors couple to an increase in adenylyl cyclase activity
3. What does differ is the tissue distribution of these receptor subtypes

Question 29

What is an example of a neuroactive peptide?

Answer 29

neurotensin

Question 30

Neurotensin:

Nervous system distribution & physiological roles

Answer 30

1. Neurotensin is a 13 amino acid peptide
2. Found in the prefrontal cortex (limbic cortex) and in the hypothalamus and midbrain
   
   • Always is co-localized with the dopamine
3. Peptides in general are co-localized with other neurotransmitters and affect and are affected by these NTs:
   
   1. Therefore, it is hypothesized that the neuropeptides add complexity, (“color”) to the simple signaling that could occur with a single transmitter

Question 31

Neurotensin:

Regulation of synthesis

Answer 31

1. Gene encoding a precursor to neurotensin is transcribed, spliced and translated by ribosomes
   
   • Precursor: neuromedin N
2. Packaged at the Golgi into secretory granules
3. Granules also contain peptidases that process the precursor protein into neurotensin
   
   • Granules are dense core

Question 32

Neurotensin:

Regulation of storage and release

Answer 32

1. Stored in different type of vesicles: dense core
   
   • Transported to the terminals via axonal transport
   • Concentrations at the terminal are low and can be variable
2. Release is calcium dependent, but require high intensity, rapid firing of neurons to be released
3. Release is not necessarily at the active zone
   
   • ​​can be released anywhere along the membrane

Question 33

How is neurotensin inactivated?

Answer 33

1. Nonselective peptidases cleave the peptide
2. Diffusion
3. No reuptake by transporters
   
   • However, peptide bound to receptor can be internalized along with the receptor
4. As a result, peptides tend to “last longer”
   
   • more persistent at receptors than norepinephrine

Question 34

Neurotensin:

Receptors

Answer 34

G protein coupled

• Neuropeptide is large compared to glutamate, its binding results in many bonds being formed between the receptor and ligand
• Results in a higher affinity binding
  
  • (i.e. receptor activation occurs at lower concentrations of neurotensin compared to glutamate or NE
• This could compensate for the lower amounts released and the distance needed to travel