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Biomembranes > ACh Release > Flashcards

ACh Release Flashcards

1
Q

Describe how activation of the nAChR leads to opening of the associated cation-conducting ion channel

A
  • agonist binding to the receptor induces a conformational change to open associated ion channel
  • nAChR channel is a cation channel primarily conducting Na into the cell and K out of the cell
  • signalling is extremely rapid
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2
Q

How can we study the functional properties of the receptor-channels in real time

A

patch-clamp technique

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3
Q

Describe the patch-clamp technique

A
  • sharp electrodes penetrate into muscle fibre
  • patch electrodes polish end and make it smooth
  • patch electrode touches surface of neuron
  • suck on syringe and hope that in membrane there is a receptor or two
  • seal is mechanically tight and electrically tight
  • ACh binds to receptor
  • can see current that flows with sodium going through receptors
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4
Q

How can we analyse results from the patch-clamp technique to study the activity of single receptors

A
  • jump down on graph is channel opening -> sodium fluxes through
  • can be used to explore effects of drugs on singular receptors
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5
Q

Describe the inside-out formation of the receptor arising from the patch-clamp technique

A

receptor is located facing inside the glass electrode and is immediately pulled from the cell

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6
Q

Describe the outside-out formation of the receptor arising from the patch-clamp technique

A

membrane snaps back in such a way that there is a piece of membrane at each end of patch electrode that fold to have receptor pointing out towards the back (facing the whole cell)

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7
Q

Advantages of the patch-clamp technique

A
  • can record the activity of single receptors
  • the intracellular milieu is maintained
  • control of the extracellular solution
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8
Q

Disadvantages of the cell-attached patch-clamp technique

A
  • investigating different concentrations of drugs e.g. agonists is complex
  • the agonist is always present, meaning receptors can become desensitized and inactive
  • no control of the intracellular content
  • the resting membrane potential is not known
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9
Q

Advantages of the whole-cell voltage clamp patch-clamp technique

A
  • control of the intracellular and extracellular milieu
  • can record from a population of receptors/ion channels
  • can introduce biochemical modulators (intracellularly) via the recording pipette
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10
Q

Disadvantages of the whole-cell voltage-clamp patch-clamp technique

A
  • biochemical “washout” of intracellular milieu
  • can rarely observe single receptor activity
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11
Q

Advantages of the outside-out patch patch-clamp technique

A
  • control of the intracellular and extracellular milieu
  • can change/control extracellular drug (agonist) concentrations
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12
Q

Disadvantages of the outside-out patch patch-clamp technique

A
  • biochemical “washout” of the intracellular milieu
  • cannot easily change intracellular messengers e.g. cAMP
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13
Q

Advantage of the inside-out patch patch-clamp technique

A
  • control of the intracellular and extracellular milieu
  • can introduce biochemical modulators e.g. cAMP to the intracellular face
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14
Q

Disadvantages of the inside-out patch patch-clamp technique

A

cannot easily change concentration of agonist

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15
Q

What is the reversal potential

A

the potential at which there is no net current flow

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16
Q

How can the reversal potential be used

A
  • can be used as a fingerprint to identify which ions are being conducted through the channel
  • can be coupled with molecular biology techniques to determine the part of the protein that controls the conduction of cations/anions
17
Q

High net current indicates outward flow of which ion

A

Potassium

18
Q

Low net current indicates inward flow of which ion

A

Sodium

19
Q

Where net current = 0 (reversal potential)

A

equal inward and outward flow of sodium and potassium (respectively)

20
Q

Therapeutic prospects of the neuronal nicotinic receptors

A
  • Alzhimers - cognitive enhancement
  • Parkinson’s
  • Schizophrenia
  • ADD
  • Pain - analgesia
  • Anxiety
  • Smoking cessation
21
Q

How can we study functional properties of recombinant wild type and mutant receptors

A
  • Xenopus oocyte expression system with voltage-clamp
  • epithelial cell transfection system with whole-cell voltage-clamp and patch-clamp
22
Q

Describe the oocyte expression system

A

oocytes derived from xenopus laevis are injected with DNA/RNA and placed into a voltage-clamp recording and the dunction is studied using the two-electrode voltage-clamp technique

23
Q

Application of neurotransmitter to oocyte

A

receptor opens, allowing chloride ions to leave cell, resulting in transient inward current that is recorded and analysed by computer software

Biomembranes (12 decks)

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