Receptor theory Flashcards
Examples of what the ligand can be radioligand binding assays
Neurotransmitters, hormones, growth factors, cytokines, drugs, toxins
Radioligand binding assays
Prepare cells e.g. with detergent treatment and centrifugation, aliquot out membranes onto filters, add radiojlabel at different concentrations to each tube, add an excess of non-radiolabelled to one tube
How can you reduce non-specific binding?
Anti-absorbents
What must the ligand be in a radioligand binding assay?
Biologically active
Advantages of using I125 in a radioligand binding assay
If the compound has an aromatic hydroxyl group it can e incorporated at very high specific activities, ionisation easy in most labs and cheap
Disadvantages of using I125
More readily degraded, short half life (67 days), function of compound can be altered
What can the tissue be in a radioligand binding assay?
Isolated membrane, slices, synaptosomes, cultured cells or solubilised/ purified receptors from total brain or specific brain region
What concentrations of protein and assay volumes do you need in incubation?
Protein conc- 0.1-1mg membrane protein/ ml. Assay volumes - 0.25-1ml
Scatchard equation
B/F=Bmax-B/ Kd Bmax= max number of binding sites, B- specifically bound F-free Kd-affinity
Kd
The drug concentration when 50% of the receptors are occupied by drug
Langmuir equation
Bound= Bmax+Xa/Xa+Kd
Units of Bmax and KD
Bmax- pmole/mg protein. Kd=mol/litre
Potency depends on
Affinity, efficacy and spare receptors
Equation for response
Response= max x [Xa]n / [Xa]n = [EC50]n n= slope factor (number of agonists that need to bind to each receptor)
What are partial agonists useful for?
Less overdose, less desensitisation of the receptors, opiates as pain killers, alleviate addicts
Example of chemical antagonism
Inactivation of heavy metals whose activity is reduced with the addition of a chelating agent
Example of pharmacokinetic antagonism
Patients taking Warfarin have to be careful when taking antibiotics as they may stimulate the metabolism of warfarin so reducing its effective concentration in the blood
Example of non-competitive antagonist
Smooth muscle histamine receptor, activates GPCR, generates IP3, causes calcium release from store, activates CaM, phosphorylation of myosin, Cav L type open which contributes to calcium signalling and causes contraction. Dihydoperidines block Cav L type for people with high blood pressure so reduces calcium influx therefore contraction
What is dose ratio?
How much more time agonist is needed in the presence of an antagonist
How do you find pA2 from a schild plot?
Negative of the value where the line crosses the x axis
Why might the effect of a drug decrease over time?
Enocytosis of receptors, change in receptor itself, exhaustion of mediators, increased metabolic degradation of extrusion of the drug, physiological adaptation
Tachyphlaxis
Rapid and short term onset of drug tolerance
