Measurement Of Drug Action At Receptors

Front 1

What is KD?

Back 1

The molar concentration of drug require to occupy 50% of receptors at equilibrium

Front 2

How can we measure the affinity of a new drug e.g relaxin at cannabinoid receptors in the brain?

Back 2

Label the drug with a radioactive isotope

We can gain information on the number of specific binding sites (cannabinoid receptors) in our tissue (brain) and the affinity of the drug (relaxin) for these receptors

Front 3

How can we do a radioactive binding assay?

Back 3

Front 4

What is "hot relaxin"?

Back 4

Radioactive relaxin

Front 5

What will occupy specific receptor sites and what will occupy non specific receptor sites?

Back 5

Specific receptor sites occupied by "cold" relaxin

Non specific binding sites occupied by "hot" relaxin

Front 6

For the first set of tubes, the amount of binding is plotting against the concentration of labelled relaxin. What will this give a plot of?

Back 6

"Total binding" and will include both "specific" (to receptors) and "non specific" (elsewhere) binding

Front 7

How can we tell from this curve that the drug isn't just binding to receptors?

Back 7

If drug is just binding to receptors, line should eventually level off as there are a limited number of receptors.

Front 8

For the second set oftubes plot the amount ofbinding against theconcentration of thelabelled relaxin. What will this give a plot of?

Back 8

Non-specific binding since all the receptor sites willhave been occupied by the“cold” relaxin and thus nothave been free to bind theradioactive drug

Front 9

How can we get a measure of specific relaxin binding?

Back 9

Subtract non-specific binding from the total binding

Front 10

What kind of plot is this called and why?

Back 10

A saturation plot as the specific binding saturates as the receptor sites become full

Front 11

What can we calculate from data such as this?

Back 11

The maximum number of receptors per tissue (B max)

The affinity of (KD) of the radioactively labelled drug

Front 12

The extent to which the agonist curve is shifted by the antagonist is a measure of what?

Back 12

The affinity of the antagonist for the receptor

Front 13

What is meant by the "dose ratio"?

Back 13

The ratio of concentration of agonist producing the same response in the presence and absence of the antagonist

Front 14

Under control conditions, you may need 1mM of agonist to get a response, but then you use a reversible competitive antagonist and you now need 20mM to get the same response. What is the dose ratio?

Back 14

20

You have had to increase the concentration of drug by 20 times to get the same response

Front 15

The extent to which you have to increase the concentration of agonist is a measure of what?

Back 15

The antagonists affinity to the receptor

Front 16

For a reversible competitive antagonist, how should the does ratio increase with the concentration of agonist?

Back 16

Linearly

Front 17

How is the affinity of a reversible competitive antagonist measured?

Back 17

Using its pA2.

Front 18

What is

Back 18

The negative logarithm of the concentration of antagonist that necessitates that you double the agonist concentration to produce the same response (i.e dose ratio =2)

Front 19

The presence of 6.3 x10-9 M propranololnecessitates that you need to double theconcentration of noradrenaline to producethe same increase in heart rate. What is the pA2 for propranolol?

Back 19

Front 20

A lower drug concentration gives a higher pA2. Do you want a drug to have a high or low pA2?

Back 20

High

This means it is effective in low concentrations so less of the drug needs to be present to produce the blocking action

Front 21

The pA2 can be obtained by experiment. A log [concentration] vs response curve for the agonist is plotted in the presence of several concentrations of antagonist. This allows the pA2 to be calculated by "Schild analysis". What else does it allow?

Back 21

Allows the experimenter to determine whether the antagonism is competitive.

Schild analysis cannot be carried out on non competitive antagonists.

Front 22

What is the Schild equation?

Back 22

Front 23

What is KB?

Back 23

The dissociation equilibrium constant (similar to KD) for the antagonist

Front 24

For a reversible competitive antagonist a plot of log(dose ratio-1) vs log [antagonist] (aSchild plot) should have a slope of what and where does it intercept the x axis?

Back 24

Slope of 1

Intercept x axis at (log KB)

Front 25

For a reversible competitive antagonist, what is pA2 equal to?

Back 25

Front 26

What is the first step in producing a Schild plot?

Back 26

Construct (log) [agonist] vs-response curves to the agonist in the presence of several concentrations of antagonist

From the curves read the log EC 50 values for the agonist in the absence of presence of different concentrations of antagonist.

Calculate the EC50 values for the agonist in the absence and presence of the different concentrations of antagonist

Front 27

What is the second step in producing a Schild plot?

Back 27

Calculate the dose ratio for the concentration of each antagonist

Plot Log (dose ratio -1) vs log [agonist]

Front 28

The X-axis intercept occurs when Y=0 so what is the does ratio here?

Back 28

Front 29

If the slope of the Schild plot is not equal to 1.0 ( or at least fall in the range 0.8-1.2) them way does this suggest?

Back 29

The antagonism is either NOT competitive or NOT reversible

Front 30

Is the pA2 obtained for the antagonist dependent or independent of the agonist used?

Back 30

Independent as it is a measure of the antagonists affinity for the receptor

Front 31

2 different experiments with different agonists but with same antagonist should have different pA2s. True or False.

Back 31

False.

Should have same pA2 because pA2 is nothing to do with agonist.

Front 32

What does it suggest if the same drug has a different PA2 in the heart compared to the lungs?

Back 32

Receptors must be different.

Front 33

What is B max?

Back 33

The maximum number of receptors per tissue