Caffeine

Question 1

What is the most widely-used psychoactive drug in the world?

Answer 1

Caffine

Question 2

When was caffeine first isolated from coffee? By whom? When? What was it called?

Answer 2

• First isolated from coffee in 1820 by Ferdinand Runge (German chemist), who called it ‘Kaffeebase’

Question 3

When was the term ‘caffeine’ first used in a medical dictionary?

Answer 3

1920

Question 4

Is caffeine a natural or synthetic compound?

Answer 4

Natural

Question 5

How many plant species produce caffeine? Name 4 of them and their country/region of origin.

Answer 5

around 60 plants are known to produce caffeine, including:
o Coffea arabica – grown in South America, mainly in Brazil
o Coffea robusta – grown in South-East Asia, mainly in Vietnam
o Camillia sinesis (tea plant) – grown mainly in China
o Theobroma cacao (cocoa tree) – grown mainly in South America

Question 6

How is caffeine typically administered?

Answer 6

• Caffeine is normally consumed through ingestion of caffeine-containing foods and drinks, where it is then absorbed via the gut

Question 7

Give the caffeine content of 5 different beverages.

Answer 7

Caffeine content varies dramatically across different foods and drinks:
o	Decaf coffee = 3mg 
o	Hot chocolate = 19mg 
o	Green tea = 20mg 
o	Espresso shot = 27mg
o	Can of cola = 40mg
o	Black tea = 45mg
o	Red bull = 80mg
o	Instant coffee = 82mg
o	Brewed coffee = 95mg

Question 8

What is the worldwide user average consumption of caffeine?

Answer 8

~170-300mg/day

Question 9

Chemically, what is caffeine? What properties does it have and what do they allow? Draw caffeine’s chemical structure.

Answer 9

• Caffeine is a member of a group of purine alkaloids called xanthines
• Caffeine is a methylxanthine – a xanthine molecule with 3 methyl groups
• It’s hydrophobic enough to readily pass through biological membranes, including the BBB

Question 10

How long does caffeine take to be absorbed?

Answer 10

• Gastrointestinal absorption is 99% within 45 minutes of consumption – fast
• Peak blood plasma levels occur within 120 minutes of ingestion

Question 11

What is the half-life of caffeine? What factor affects this and what are its medical implications?

Answer 11

• 3-7 hours (average is 5 hours) in adult humans
o This is doubled in women taking oral contraceptives – oestrogen inhibits the enzyme responsible for metabolising caffeine
o This increases progressively with ageing – the half-life is 30% larger when 70+ vs when 20-30 years old
• 80-100 hours in neonates – lack the enzymes to degrade caffeine so stays in the system for longer durations
o This is why pregnant women shouldn’t drink caffeine

Question 12

How is caffeine metabolised? Which enzymes catalyse this?

Answer 12

• Caffeine is metabolised in the liver by demethylation

* This is catalysed by the cytochrome P450 family of enzymes – specifically, CYP1A2 and CYP2E1

Question 13

What are the 3 primary metabolites of caffeine? Which enzymes are they produced by? Draw their structures.

Answer 13

o	Paraxanthine (~84% of caffeine becomes paraxanthine) - CYP1A2
o	Theobromine (~12% of caffeine becomes theobromine) - CYP1A2/2E1
o	Theophylline (~4% of caffeine becomes theophylline) - CYP1A2/2E1

Question 14

What is considered a low dose of caffeine and what are its physiological and psychological effects?

Answer 14

Low dose (50-300mg).

Physiological:
Elevated blood pressure due to vasoconstriction and increased heart rate
Relaxed bronchioles
Diuresis (increased urine production)

Psychological:
Increased alertness
Decreased drowsiness (different to alertness)
Improved attention
Improved motor skills

Question 15

What is considered a moderate dose of caffeine and what are its physiological and psychological effects?

Answer 15

Low health risk, similar effects to low dose; just more intense

Question 16

What is considered a high dose of caffeine? What are its physiological and psychological effects? Why do they occur?

Answer 16

High dose (>1000mg).

Physiological:
Tachycardia

Psychological:
Anxiety
Insomnia

These effects are the result of mass-receptor antagonism –> overexcitation and hyperactivity, amplifying the usual effects

Question 17

What is caffeinism?

What are its symptoms?

Answer 17

A state of intoxication wherein physical and mental control are diminished, caused by excessive caffeine consumption – typically >1000mg/day
•	Symptoms are divided into low-dose and high-lose symptoms:
Low dose (>100mg/day):
Restlessness
Nervousness
Insomnia
Flushed face
Diuresis
Gastrointestinal disturbance

High dose (>1000mg/day):
Rambling speech
Inexhaustibility
Psychomotor agitation (pacing around a room, tapping toes, or rapid talking – often occurs with anxiety)
Delusions
Muscle twitching
Tachycardia/cardiac arrythmia
Appetite loss (anorexia)

Question 18

What is considered a lethal dose of caffeine and what typically achieves this dose?

Answer 18

Lethal dose = ~10g:
o Quite difficult to do by drinking coffee – it’s around 100 cups
o But, it’s possible if you’re mixing high-caffeine sources (coffee + energy drinks + caffeine tablets), or taking powdered caffeine (workout supplements)
o Death is caused by convulsions and respiratory failure

Question 19

What dosage of caffeine is typically required to develop dependence?

Answer 19

• Consumers of caffeine readily become dependent:
o The median dose of individuals exhibiting dependence is 360mg/day
 But – 40% of individuals exhibiting caffeine dependence consume <300mg/day

Question 20

List 5 of the symptoms typically encountered when withdrawing from caffeine dependency. When do they occur? Are symptoms related to the levels of caffeine normally consumed?

Answer 20

•	Withdrawal symptoms include:
o	Headache
o	Tiredness
o	Anxiety
o	Lack of concentration
o	Increased muscle tone
o	Nausea

Initial withdrawal symptoms begin 12-24 hours after stopping caffeine intake
o Peak withdrawal symptoms occur between 20-48 hours after stopping intake
o Expression of withdrawal symptoms is not related to levels of daily caffeine

Question 21

What are the mechanisms of action of caffeine?

Answer 21

• Caffeine has several biochemical targets in the body, but when at non-toxic doses its effects are mainly mediated by its action on adenosine receptors (ARs)
o At higher (toxic) doses (0.1-10mM) (or 50 cups of coffee), caffeine with block GABAARs, inhibit phosphodiesterases, and agonise ryanodine receptors (causing Ca2+ release)
• Caffeine mainly mediates its effects at A1Rs and A2ARs

Question 22

What is adenosine?

Answer 22

• It’s a product of cellular metabolism – breakdown of ATP –> adenosine production

Question 23

Which properties of a neurotransmitter does adenosine possess?

Answer 23

• Adenosine is not stored in presynaptic vesicles, but it does share other fundamental properties of a neurotransmitter:

1. Adenosine-producing enzymes are present at the synapse
2. It’s released from presynaptic terminals
3. It binds to receptors to elicit a response in post-synaptic neurons; also elicits a response from presynaptic autoreceptors, which regulate neurotransmitter release
4. It’s cleared from the synaptic cleft by reuptake or metabolism
5. Its effects can be blocked by selective antagonists

Question 24

How does caffeine achieve its effects?

Answer 24

• Caffeine antagonises adenosine receptors
o By affecting adenosine signalling, caffeine mediates its effects
o Caffeine is a competitive antagonist of adenosine receptors (aka P1 receptors)

Question 25

What sort of receptor are adenosine receptors? How many subtypes are there?

Answer 25

``` • Adenosine receptors are GPCRs with 4 subtypes: o A1 o A2A o A2B o A3 ```

Question 26

List the 4 adenosine receptor subtypes, their signalling mechanisms, the potency of adenosine on these receptors (EC50 (nm)) and their effects in neurons.

Answer 26

A1 Gi/o-->inhibition of adenylyl cyclase --> decreased [cAMP] 310 Inhibition of N-, P-, Q-type Ca2+ channels Activation of GIRK channels Inhibition of NMDARs A2A GS-->activation of adenylyl cyclase -->increased [cAMP] 700 Increased GABAergic (inhibitory) neurotransmission in the basal ganglia A2B GS-->activation of adenylyl cyclase --> increased [cAMP] 24000 Mainly expressed in non-CNS tissues Activated by high [adenosine] under pathological conditions (e.g. stroke) A3 Gi/o-->inhibition of adenylyl cyclase -->decreased [cAMP] 290 Neuroprotection and neurodegeneration

Question 27

How does adenosine mainly exert its effects in the CNS?

Answer 27

Adenosine mainly exerts inhibitory (depressant) effects in the CNS, through activity at the A1 and A2A receptors

Question 28

With the aid of a diagram, explain the release of adenosine.

Answer 28

• ATP is released through nucleotide transporters/vesicles (sometimes alongside glutamate) • ATP converted to AMP by ectonucleotidases (found in the cell membrane of the presynaptic neuron); AMP converted to adenosine by 5’-nucleotidase • Adenosine can then: o Be reabsorbed into the presynaptic neuron via a nucleoside transporter o Bind to the A1 receptor of the postsynaptic neuron o Be degraded to inosine by adenosine deaminase (extracellularly) diagram on lecture slides (25/11/19)

Question 29

With the aid of a diagram, explain the result of adenosine binding to its effectors

Answer 29

• When released, adenosine can bind to: o A1 receptors  Presynaptically: • Inhibits adenylyl cyclase -->decreased [cAMP] • Inhibits N/P/Q-type Ca2+ channels --> reduced Ca2+ influx and reduced neurotransmitter release  Postsynaptically: • Activates GIRKs (G-protein linked inwardly-rectifying K+ channels) --> hyperpolarisation of cell • Inhibits NMDARs --> reduced Ca2+ influx --> less chance of action potential being generated • Inhibits adenylyl cyclase -->reduced [cAMP] o A2A receptors:  Presynaptically: • Activates L-type Ca2+ channels -->increased Ca2+ influx -->increased probability of neurotransmitter release • Activates adenylyl cyclase --> increased [cAMP] • Inhibits A1R activity  Postsynaptically: • Activates adenylyl cyclase --> increased [cAMP]  On blood vessels: • Causes vasodilation diagram in lecture notes (25/11/19)

Question 30

With the aid of a diagram, explain how caffeine's antagonism of A1Rs and A2ARs causes the effects associated with caffeine usage

Answer 30

• Antagonises A1Rs and A2ARs both pre- and post-synaptically, as well as on blood vessels diagram in lecture notes (25/11/19)

Question 31

What is caffeine's affinity at each of the adenosine receptor subtypes?

Answer 31

High affinity at A1,2A,2BR subtypes: KD(μm) = 12, 2.4, 13 (respectively) Remember: A2BRs aren’t activated at normal physiological [adenosine] (~300nm) There’s little A2BR expression in the CNS Low affinity at A3Rs Therefore: Caffeine mainly exerts its effects on CNS function by antagonising adenosine signalling at A1Rs and A2ARs

Question 32

What is the typical effect of A1R agonism?

Answer 32

A1Rs are found almost all over the brain. The probability of neurotransmitter release from presynaptic terminals is decreased by activation of presynaptic A1 autoreceptors (occurs when adenosine is released). When thalamocortical cells are under electrophysiology (whole-cell patch clamp): EPSC amplitude is decreased by application of 100μM adenosine (probability of neurotransmitter release is inhibited); this is reversed by co-application of 1μM of the adenosine receptor antagonist CPT. Caffeine produces the same effects as CPT: It antagonises presynaptic A1Rs -->increased probability of neurotransmitter release --> increased amplitude of postsynaptic currents

Question 33

What does caffeine suppress?

Answer 33

Inhibition of neurotransmitter release typically achieved through adenosine binding to its receptors --> neurotransmitter release

Question 34

Where are A2ARs most concentrated?

Answer 34

In the basal ganglia (a dopamine-rich brain region)

Question 35

Where are A2Rs typically located?

Answer 35

• A2ARs are mainly located on GABAergic interneurons that: o Originate in the striatum o Project to the globus pallidus o Express D2 dopamine receptors (Gi/o-coupled) o A2ARs and D2Rs signal via adenylyl cyclase – but they have opposing effects; activation of A2ARs inhibits signalling by D2Rs, and vice versa

Question 36

What effect does adenosine and caffeine binding to A2ARs have on the neuron?

Answer 36

• Adenosine activation of A2ARs (Gs-coupled) increases the release of GABA from striatal interneurons projecting into the globus pallidus o Caffeine decreases GABA release – this is causes decreased inhibition --> increased neuronal activity in the globus pallidus Caffeine blocks the effects of the A2AR; blocking the downstream effects of adenylyl cyclase and having the same effect as dopamine binding a D2R – inhibiting the inhibitory activity of GABA release  a net increase in activity within the globus pallidus

Question 37

Explain, with the aid of a diagram, typical A2AR function in the basal ganglia movement circuit and how this function is affected by caffeine administration.

Answer 37

* Normally, adenosine acting on A2ARs increases the release of GABA from striatal interneurons projecting to the external globus pallidus * Caffeine blocks this action by antagonising the A2ARs; thereby decreasing the release of GABA from striatal interneurons projecting to the external globus pallidus, hence increasing glutamate release (and therefore activity) within the basal ganglia movement circuit as a whole diagram in lecture notes (25/11/19)

Question 38

What effect does caffeine have when administered to the striatopallidal area?

Answer 38

Caffeine-mediate suppression of striatopallidal inhibition promotes movement. This is thought to be how caffeine improves performance in motor tasks – by improving motor coordination.

Question 39

What are the effects of caffeine on sleep-wakefulness?

Answer 39

* Extracellular [adenosine] increases in the basal forebrain and neocortex throughout wakefulness – this is due to an elevated cellular metabolism when awake * Adenosine inhibits activity in a neuronal circuit found in the basal forebrain, which promotes wakefulness by acting on an A1R – increasing the pressure to sleep * Caffeine suppresses the inhibitory effect of adenosine on this circuit, promoting wakefulness