DRUG

TISSUE

MECHANISM OF

EXPECTED TREND

ACTION
Noradrenaline

Vascular smooth muscle

1-adrenoceptor agonist

Surmountable Emax reached

with increasing NA

1-adrenoceptor binding,

concentration

causing conformation,
causes the activation of Gq

Plateau phase observed

linked proteins.
-

Increased PLC activity

Increase in IP3 production

IP3 Receptor binding

Stimulate release of
intracellular calcium from
sarcoplasmic reticulum.

Changes in membrane
potential

Influx of extracellular
calcium via VOLTAGE
GATED L-TYPE CALCIUM
CHANNELS

KCl

Vascular smooth muscle

- Smooth muscle contraction

Stimulant of smooth muscle

All or nothing effect

contraction
-

By pass the normal Ca2+

Increasing extracellular K+
concentration

Altering the electrochemical

gradient across the myocyte

Member potential became

more positive (closer to 0)

Causes opening of voltage

gated Ca2+ channels.

Serotonin (5-HT)

Vascular smooth muscle

- Smooth muscle contraction.

5-HT2A receptor agonist

Surmountable Emax reached

with increasing NA

5-HT2A receptor binding

Same mechanism as per

noradrenaline

Metoprolol

Cardiac Tissue

concentration
Plateau phase observed

- Smooth Muscle Contraction

1-adrenoceptor selective

Heart rate and peak expiratory

antagonist

respiratory flow rate increases

as the exercise intensifies.

Compete with catecholamine

binding to 1-adrenoceptor.

Reduction in heart rate and

Reduced cAMP formation

peak expiratory respiratory

Decreased sympathetic

flow rate, for all aspects after

nervous activity

metoprolol administration

Decrease in heart rate

Decreased Ca2+ sensitivity to

No difference, in PEFR between

contractile filaments

pre and post drug at rest

Decrease in forcefulness of
contraction

Increased

As the exercise intensifies,

bronchoconstriction

heart rate and

Decreased renin release

Increased vasodilation

2-adrenoceptor (lungs)

antagonist
-

No major changes

At Exercise
-

Increased sympathetic
nervous activity, increased
NA release and action on 1adrenoceptor, inducing
increased inotropy and

Prazosin

Vascular smooth muscle

chronotropy
1-adrenoceptor competitive

Parallel rightward shift

reversible antagonist
Surmountable Emax at higher
-

Reversible binding to

agonist concentrations

postsynaptic 1adrenoceptor.
-

Plateaued response

Inhibition of
vasoconstrictor effect of
locally released
catecholamines

Methysergide

Vascular smooth muscle

- Peripheral vasodilation
5-HT2A receptor partial

Rightward Parallel Shift

agonist
No change in Emax
At low [5-HT]
Cross over at low [5-HT]
-

5-HT2A receptor binding

Same mechanism as per 5-

At very low levels of 5-HT, it

HT

produced a surmountable Emax,

Smooth Muscle Contraction

a contractile response which

even surpassed that of 5-HT

At high [5-HT]

alone, this indicates agonism.

5-HT2A receptor binding

At high 5-HT concentration,

Inhibit 5-HT action of VSM.

the contractile response does

Reduction in contraction

not reach surmountable Emax,

indicating a form of antagonism
(competitive reversible)
Eventually reach the same Emax
as 5-HT, at higher agonist
concentration

Verapamil

Vascular smooth muscle

Non-competitive antagonist

(However it is cardioselective)
-

Reversible allosteric binding

to L-type voltage gated Ca2+
channel.

Reduced extracellular
calcium entry into the
myocyte.

Decreased phosphorylation
of contractile filaments

Carbachol/Acetylcholine

Vascular smooth muscle

- Smooth Muscle Relaxation

Physiological antagonist
The antagonist (in fact an
agonist) produces a response

which opposes the action of

the agonist
-

Bind to M3 muscarinic
receptors on the vascular
endothelium

Activation of Gq linked
proteins

Increased activity of iNOS

Increased NO synthesis in
endothelial cells

NO diffusion into smooth

muscle.

Via cGMP and PKG activation

which leads to a reduction
of intracellular calcium

Oxytocin

Uterine smooth muscle

- Smooth muscle relaxation

Oxytocin receptor agonist

Surmountable Emax reached

with increasing NA

Contraction needed to prevent

Binding to oxytocin receptor

bleeding

Activation of Gq linked
proteins

Activation of PIP2

IP3 activation.

Release of Ca2+ ions from

concentration
Plateau phase observed

sarcoplasmic reticulum
-

Influx of extracellular
calcium via voltage gated
calcium channels.

Terbutaline

Uterine smooth muscle

- Smooth muscle contraction

2-adrenoceptor agonist (also

Differing potency

physiological antagonist with

regards to oxytocin)
-

2-adrenoceptor binding

Increased activity of adenyl

cyclase

Increased cAMP production

Increased Protein kinase A

activity

Inhibition of myosin light

chain kinase activity

Decreased phosphorylation
of myosin light chains

Increased activity of MLCK

phosphatase activity

Salbutamol

Uterine smooth muscle

Smooth muscle relaxation

Inhibition of uterine

contraction
2-adrenoceptor agonist (also
physiological antagonist with
regards to oxytocin)
-

2-adrenoceptor binding

Increased activity of adenyl

cyclase

Increased cAMP production

Increased Protein kinase A

activity

Differing potency

Inhibition of myosin light

chain kinase activity

Decreased phosphorylation
of myosin light chains

Increased activity of MLC

phosphatase activity

Diltiazem

Uterine smooth muscle

Smooth muscle relaxation

Inhibition of uterine

contraction
L-type calcium channel Blocker
-

Differing potency

Binding to L-type voltage

sensitive calcium channels

Reduction in the influx of

extracellular calcium into
the cytosol

Reduction in the calcium

binding to calmodulin

Decreased formation of
calcium-calmodulin
complexes

Decreased activation of
MLCK.

Nifepidine

Uterine smooth muscle

Smooth muscle relaxation

Inhibition of uterine

contraction
L-type calcium channel Blocker
-

Differing potency

Binding to L-type voltage

sensitive calcium channels

Reduction in the influx of

extracellular calcium into
the cytosol

Reduction in the calcium

binding to calmodulin

Decreased formation of
calcium-calmodulin
complexes

Decreased activation of
MLCK.

Apomorphine

Central Nervous System

Smooth muscle relaxation

Inhibition of uterine

contraction
Non-selective Dopamine

Contralateral turns to the side

receptor agonist

of 6-OHDA lesion

Nigro-striatal dopaminergic
pathway

Direct stimulation on D1/D2

receptors.

Retrograde destruction of the

Super-sensitization of

pathway from SN to striatum,

Dopamine receptors in the

causes parkinsonism like motor

lesioned hemisphere

symptoms.

Increased dopamine
production

6-OHDA

Central Nervous System

Lesion at one side (right) of

hemisphere

Nigro-striatal dopaminergic
Amphetamine

pathway
Central Nervous System

Dopamine receptor agonist

Ipsilateral turns to the side of

6-OHDA lesion
Nigro-striatal dopaminergic

Act on pre-synaptic DA
receptors

Left side have higher

Increased release of stored

dopamine activity, since 6-

Drug induced rotations, since 6-

dopamine from the DA

OHDA lesions all the

OHDA induced rotations are

terminals

dopaminergic pathways on the

Inhibition of dopamine re-

right side. Thus the rat want

uptake pathways

to avoid the side with highest

Increased endogenous

dopamine activity. And rotate

dopamine levels in the

towards the right.

pathway
-

not uniform

synaptic cleft
At the right where there is
less DA pathways, the
receptors are supersensitized.
And at the administration of
apomorphine, which acts
similarly like dopamine at this
supersensitized site, which
means the right has higher
activity, the rat do not like
this thus turns towards the
Chlorpromazine

Central Nervous System

Dopamine Receptor Antagonist

left.
Low Dose

Dopaminergic Pathways

Equal affinity binding to D2

Alert

and D4 receptors

Un-coordination

Decreased cAMP levels

Reduced respiration rate &

Decreased dopamine

temperature

synthesis
-

Reduced dopamine levels in

Foot Pad Pinch

the synaptic cleft

Corneal Reflex

A reduction in parkinsonism

Righting Reflex

relate symptom

Catalepsy Test

A reduction in
extrapyramidal activity.

High Dose
Heavily tranquilized
Tail curling

Pentobarbitone

Central Nervous System

Anti-Psychotic Properties
GABAA Receptor Agonist

Foot Pad Pinch

Corneal Reflex

Righting Reflex (dull)

Catalepsy Test

Low Dose

GABAergic pathways

Reduction in GABA-sensitive

Animal very alert

neuronal calcium

Reduced body temperature

conductance.
-

Hypersensitive to external
-

Binding at the Cl ionopore

of the GABAA receptor.

stimuli

Increase the duration of

-

time for Cl ionopore to open

-

Foot Pad Pinch

Corneal Reflex

Increased Cl ion influx

Righting Reflex

Prolonged post-synaptic

Catalepsy Test

inhibitory of GABAA in
thalamus.
-

High Dose

Acute potentiation of
inhibitory GABAergic tone

Nerve twitching
Anaesthetized

Anaesthetic Effects

Reduction in body
temperature.

Methylphenidate

Central Nervous System

Dopamine Transporter Blocker

Foot Pad Pinch

Corneal Reflex

Righting Reflex

Catalepsy Test

More excitable
Elevated body temperature

Dopaminergic Pathways

Binding and inhibition of

and respiration rate

dopamine transporter

Extensive piloerection

protein in central adrenergic

Hyperexcitable

neurons
-

Inhibition of dopamine

Low Dose

uptake from synaptic cleft

Increased dopamine levels

Foot Pad Pinch

in the synaptic cleft

Corneal Reflex

Hyperactivity

Righting Reflex

Catalepsy Test

Stimulatory Effects
High Dose

Diazepam

Foot Pad Pinch

Corneal Reflex

Righting Reflex

Catalepsy Test

GABAA Receptor Agonist

Low Dose

Alert

Diazepam bind nonspecifically to benzodiazepine

binding site of GABAA
receptor

Increase GABA affinity to

GABAA receptor

Increase the frequency of

chloride ionopore opening.

Increased Cl- influx

Hyperpolarized cell membrane

Prevention of further
excitation of cell.

Increased inhibitory effect

Uncoordinated movement

Foot Pad Pinch

Corneal Reflex (dulled)

Righting Reflex

Catalepsy Test

Medium Dose
Heavily sedated
Reduced body temperature

of GABA

Lowered respiration rate

Tranquilizing Effect

Foot Pad Pinch

Corneal Reflex (dulled)

Righting Reflex (sluggish)

Catalepsy Test

High Dose

 Heavily tranquilized
Reduced body temperature
and respiration rate

Foot Pad Pinch (slight)

Corneal Reflex (very dull

and slow)

Histamine

Skin

Righting Reflex

Catalepsy Test

H1 receptor agonist

Increase in weal size

Histamine Binding to H1

Weal size = increased

receptor

permeability of post-venule

Activation of Gq linked

capillaries, local oedema, cell

proteins

tight junctions are becoming

Activation of PLC and PIP2

loosened and thus allowing

pathways

things to flow through

Vasodilation
Flare = Local vasodilation

Fexofenadine

Skin

H1 receptor competitive

Reduction in weal size

antagonist
-

Compete with free histamine

at the H1 receptor binding
site.

Reduced activation of Gq
linked proteins

Reduced activation of PLC

and PIP2 pathways

Vasoconstriction

Laboratory 1
Cumulative Addition (i.e. into a 20ml organ bath)
Example: Add acetylcholine cumulatively into a 20ml organ bath to achieve the final concentrations 1x10 -10M, 3x10-10M and 1x10-9M. The
stock you have been given is 1x10-6M.
Note: Start from the lowest concentration and work your way up. Ideally you should not add less than 10 L or more than 40 L into
the organ bath in any one addition, however in certain circumstances it may be necessary to add volumes outside this range (e.g. when a
stock solution is saturated so cannot be made more concentrated).
Step 1: Calculate your first addition. C2 = 1x10-10M, V2 = 20mls, C1 = 1x10-6M.
V1 = C2 x V2 / C1
V1 = 2L
The problem with this though is that the volume is too small. Best practice suggests no volume less than 10 L. Thus we need to dilute
our starting stock ten-fold, ie 20L of stock + 180L of water to make 200L of 1x10-7M. This is our new working stock solution.
Step 2: Recalculate the first addition. C2 = 1x10-10M, V2 = 20mls, C1 = 1x10-7M.
V1 = C2 x V2 / C1
V1 = 20L
The first addition into the organ bath is 20L of 1x10 -7M.

Step 3: Calculate the second addition. C2 = 3x10-10M, V2 = 20mls, C1 = 1x10-7M.

V1 = C2 x V2 / C1
V1 = 60L
Note: We already have 20L of drug in the organ bath which is still there. Thus we don't need to add the whole 60L. We only need
to add another 40L of 1x10-7M stock to get a final concentration of 3x10-10M. You might also see that there is a shortcut here. The
second concentration you require in the organ bath is three times that of the first concentration. Thus to achieve it you need 3X the
amount of drug, ie 3 x 20L = 60l in total.
Step 4: Calculate the third addition: C2 = 1x10-9M, V2 = 20mls, C1 = 1x10-7M.
V 1 = C2 x V2 / C1
V 1 = 200L
The problem with this answer is that the volume is too big. Even if we subtract the 60L already in the bath we would still have to add
140L to achieve the desired concentration. So what do we do now??
Hint: 200L of 1x10-7M is equivalent to 20L of 1x10-6M, thus if we have already added 60L of 1x10 -7M it is the same as saying we
have added 6L of 1x10-6M. Therefore to get a concentration of 1x10 -9M we need to add another 14L of our original stock of 1x10 -6M.
The lab has two different models of specs, one will read automatically once the door is closed (black specs), whilst the other model
requires you to press the 'read' button (beige specs).

Laboratory 4 Beta Blockers

Figure 1: The changes in (A) heart rate and (B) peak expiratory flow rate following metoprolol administration, at rest and during low and high
intensity exercise. A repeated measure design was conducted; where data for all 3 conditions at baseline (no drug) and post-drug (1 hour after 50mg
metoprolol administration) were obtained from the same participant. Across all 3 conditions, HR was taken manually from the carotid or radial artery
and PEFR was recorded using the PEFR meter. Data represents means calculated by Microsoft Excel 2010, n = 37

1. Compare and contrast the pharmacological concepts of selectivity versus specificity of a drug.
Selectivity = preferential binding to a specific type of receptor but starts binding to other receptors at high concentrations e.g.
metoprolol preferentially binds to B1 at therapeutic doses but binds to B2 at high doses
Specificity = can only bind to one type of receptor no matter how high the concentration (pretty sure no current drug on the
market is completely specific)
2. Understand the concept of repeated measures design.
Measurements will be repeated 1 hour following oral intake of the beta-blocker metoprolol.

Laboratory 6 Drugs and Blood Vessels

Competitive Reversible Antagonism (CRA): the antagonist binds to the receptor to prevent the agonist binding. A competitive
antagonist binds to the receptor and prevents the binding of the agonist. In competitive reversible antagonism the effect of the
antagonist can be overcome by increasing the concentration of the agonist.

Competitive Irreversible Antagonism: As above the competitive antagonist binds to the receptor and prevents agonist binding.
However if the antagonist binds irreversibly increasing the agonist concentration will not overcome the effect of the antagonist.
Non Competitive (NC) Antagonism: the antagonist interferes with the transduction process between agonist binding and response.
Physiological Antagonism: the antagonist (in fact an agonist) produces a response which opposes the action of the agonist.
Pharmacokinetic antagonism: The antagonist reduces the concentration of the agonist at its site of action due either to reduced drug
absorption or an enhanced elimination rate.
Chemical antagonism: the antagonist combines with the agonist (very uncommon).

Laboratory 8 Drugs and the Uterus

Figure 2. The impact of anti-tocolytic drugs on the contraction of rat uterine muscle.
Blue = terbutaline
Red = salbutamol
Green = diltiazem
Purple = nifepidine

1. Understand different concepts (potency AND efficacy) for determining which tocolytic drugs would be best in a
clinical setting and why.
Expected results: Salbutamol > Nifepidine > Diltiazem > Verapamil
2. Determine whether any of the studied tocolytics change frequency of contraction instead or as well as magnitude.
Explain the possible mechanism behind this.
3. The reason for one drug being more potent that another is due to the affinity of the drug for its receptor and
the resulting response that is elicited when it binds.

For working out which had the biggest effect I would take the raw grams data and compare the absolute value of
relaxation, ie how many grams of contraction did each tocolytic achieve compared to its oxytocin maximum. eg if
oxytoxin gave 3.37gm of contraction and the maximum change we got from 1 x 10-6M terbulatine was -1.32gms then the
amount of relaxation seen was 3.37 - -1.32gms which is 4.69gm.

Laboratory 9 Effects of Drugs on Behavioural Function 1

Freedom from hunger and thirst. In order to achieve this all animals must be provided with water and a nutritious diet that has not
been contaminated by other animals, dietary pests or micro-organisms such as mould.

Freedom from discomfort. To provide comfort to animals includes providing clean fresh caging regularly, fresh air, fresh nesting
materials and environmental enrichment where ever possible.
Freedom from pain and injury or disease. Ensuring that all animals are inspected regularly, apparent ill health or injury is reported to
the senior staff to enable treatment to be instigated or alternative regimes adopted and prevention of disease by adopting strict
husbandry procedures can fulfil this freedom.
Freedom to express normal behaviour. This can be fulfilled by providing animals with group or social housing wherever possible and
by providing sufficient space in which animals can express their normal behaviours.
Freedom from fear and distress. Fear and distress can be avoided in many ways, these include prevention of sudden and loud noise,
reduction in changes to routines, providing cages of adequate sizes for the groups of animals held, not overcrowding animals and caring
husbandry techniques.
Mouse-Handling
Getting them out of their container: Take off the lid and tilt the container. Gently jiggle the container over your towel so the mouse
slides out on to the towel. Grab the base of the tail gently and then rest the mouse on a flat surface such as your lab coat arm,
restraint cage, towel or lab bench. DO NOT DANGLE THE MOUSE IN THE AIR BY THE TAIL !
Keeping mice subdued: Whilst the mouse is in its cylindrical plastic white boring container it is relatively subdued. As soon as you open
the lid and let it out, it will be excited and in the mood for exploring. This keeps the mouse occupied for about a minute, thus is the
perfect time to restrain the mouse ready for injection.
Restraining before injection: An ideal place for the mouse is on a soft towel on a flat surface. Whilst holding the tail with one hand,
approach the head of the mouse with your forefinger bent and your thumb splayed (forming a 'V' shape that can grip the scruff).
Slowly but firmly, grab the mouse by the scruff using your thumb and forefinger. Take a large chunk of skin and fur for a tight hold
that will immobilize the mouse and make it unable to turn its head. Aim for the area under the ears and above the shoulder blades. You
may need to practise this a few times to get a good hold. Once you feel comfortable about the hold, orientate the mouse so that the
belly is facing you. You may like to wedge the mouse's tail between your two last fingers to help keep the abdomen taught.
Injection volume (mls) = Dose (mg/kg) x Weight of mouse (kg) / Stock concentration (mg/ml)

Laboratory 10 Effects of Drugs on Behavioural Function 2

1. Understand and describe the difference between qualitative and quantitative data.
Quantitative data are numerical presentations of data.
Qualitative data are descriptive/narrative focused.

Laboratory 11 Histamine and Anti-Histamines

Figure 3. Green = fenofoxedine, red = histamine

Verapamil is a cardioselective blocker of voltage gated L-type channels. Its effect is causing muscle relaxation and thus reduction in
cardiac output. Agonist have utilized 2 sources of calcium, intracellular SR as well as extracellular calcium. Blocking the receptor
directly reduce calcium entry. Verapamil stops one of the extracellular calcium, the binding to the channel is reversible, but it is at a
different binding site, a type of non-competitive reversible antagonism.
Focus on absolute relaxation, what is the most effective drug, terbutaline because it produced the highest relaxation, however
secondary thoughts, the potency must be considered, if there is not an adequate potency, then there will be a reduction in selectivity
and the side effects must be considered.
Corneal reflex, allow us to distinguish between tranquilized mice and anaesthetic mice
Anaesthetic will have analgesic properties (pain relief)
Tranquilized, just means really sleeping, corneal reflex should still work.