Anxiety: Benzodiazepines and barbiturates Flashcards
1
Q
what are the historic uses of barbiturates ?
A
- anxiolytics
- anticonvulsants
- sedatives/hypnotic
- anaesthetics
2
Q
when are barbiturates used now ?
A
- only in extreme cases for anxiolytics and anticonvulsants
- for induction or short procedures as sedatives and anaesthetics
3
Q
why are barbiturates not used as often now ?
A
they are very dangerous in overdose as they cause respiratory depression
4
Q
what is the main mechanism of action of barbiturates ?
A
- as positive allosteric modulators of GABA-A receptor
- act by prolonging the opening time/ GABA works better
- at high concentrations can directly activate the receptor
5
Q
what can barbiturates do at high concentrations that makes them dangerous ?
A
- directly activate GABA-A receptor
- reason why more lethal in overdose
6
Q
apart from their actions at GABA-A receptors, what other receptors do barbiturates act at ?
A
- inhibit AMPA
- P/Q type voltage sensitive calcium channels (inhibits NT release)
7
Q
what is the role of barbiturates ?
A
they can increase inhibitory transmission and decrease excitatory transmission
8
Q
where do barbiturates bind on the GABA-A receptor and what does this mean?
A
- beta subunit (TMD 2&3)(maybe alpha too)
- would need to enter lipid bilayer to access binding site
9
Q
what is thiopental (a barbiturate) used for ?
A
- general anaesthesia
- status epilepticus (if not responding to benzodiasepines or anticonvulsants)
10
Q
what (human) medical use do barbiturates have ?
A
can be used to induce comas at higher doses
11
Q
what could barbiturates potentially be used for in vetinary practice ?
A
euthanasia
12
Q
In the US, lethal injection is a common form of capital punishment. What are 3 common drugs in the cocktail ?
A
- sodium thiopental
- pancuronim bromide
- potassium chloride
13
Q
What is the purpose of Sodium thiopental in the lethal injection cocktail?
A
To induce loss of consciousness and repress respiratory drive
14
Q
What is the role of Pancuronium bromide in lethal injections?
A
A long acting non-depolarizing neuromuscular blocker that paralyzes the respiratory muscles
15
Q
What does Potassium chloride do in the lethal injection process?
A
To depolarize the myocardium and bring about cardiac arrest
16
Q
What action did many pharmaceutical companies take regarding their drugs used in executions?
A
Objected to the use of their drugs
17
Q
What did the European Union issue in 2011 concerning lethal injection drugs?
A
An export ban on drugs used in lethal injections
18
Q
What was one consequence of the European Union’s export ban on lethal injection drugs?
A
A shortage in some of the ingredients of the lethal injection cocktail
19
Q
Which alternatives have some states used in response to the shortage of lethal injection drugs?
A
Pentobarbital or a benzodiazepine/opioid mixture
20
Q
what is the similarity between barbiturates and benzodiazepines ?
A
they are both positive allosteric modulators of the GABA-A receptors
21
Q
what can benzodiadepines be used as?
A
- anxiolytics
- sedatives/hypnotics
- anticonvulsants
- muscle relaxants
similar to barbiturates
22
Q
what is the difference between benzodiazepines and barbiturates ?
A
benzodiazepines are safer in overdose
replaced barbiturates in 70s
23
Q
what was the first benzodiazepine synthesised ?
A
chlordiazepoxide (librium)
24
Q
what is the trade name for diazepam, the second benzodiazepine on the market?
A
valium
25
what are some limitations of benzodiazepines ?
- found to have addiction and tolerance
- now used 'sparingly': low dose, short term
26
what subunits does the GABA-A receptor contain ?
- alpha 1 (x2)
- beta 2 (x2)
- gamma 2 (x1)
27
where are the GABA binding sites on the GABA-A receptor ?
at interfaces of beta and alpha subunits
28
where is the high affinity benzodiazepine binding site of the GABA-A receptor ?
interface between alpha(1) and gamma(2) subunits
29
what kind of receptors are GABA-A receptors ?
ligand gated chloride channels
30
what happens when GABA binds to the GABA-A receptor ?
- increased chloride permeability
- E Cl close to RMP so stabilises MP close to E Cl
- (can lead to hyperpolarisation sometimes but mostly) inhibition
31
what is the difference between GABA-A receptors in adult or human tissues ?
adult is always inhibitory, but in foetal can be excitatory
32
what are properties of compounds that decrease GABA-A function ?
convulsants
## Footnote
(less inhibition)
33
what can ligands at the BDZ binding site do to the GABA-A receptor ?
can modulate function in a...
- positive (enhance function)
or
- negative (decrease function)
or
- neutral (no effect)
...way
34
what is another name for the drugs that bind to the BDZ site on GABA-A receptor and enhance function?
benzodiazepine agonists
35
what are 4 examples of benzodiazepine agonists ?
- flunitrazepam (rohypnol)
- diazepam (valium)
- temazepam
- nitrazepam (mogadon)
36
what desirable properties can BDZ agonsits have ?
anxiolytic, sedative, hypnotic, amnesia inducing
37
what are the undesierable effects of BDZ agonists ?
- high addictive liability
- can develop high tolerance
38
what is the mechanism of action/ function of BDZ agonists ?
increases GABA-A receptor channels opening frequency (increases affinity of GABA)
## Footnote
do not directly activate the receptor
39
how are BDZ agonists different to barbiturates ?
BDZ increase channel opening FREQUENCY, barbiturates increase opening TIME
40
what is an example of a benzodiazepine antagonist ?
flumazenil
41
what is the mechanism of action for BDZ antagonists (e.g. flumazenil) ?
compete with agonists for same binding site
42
what does flumazenil (BDZ anatgonist) do to GABA-A receptor function ?
pharmacologically fairly inactive when applied alone
- little effect on receptor activation
- blocks potentiation by flunitrazepam + (sometimes) other BDZ agonists
43
even though flumazenil has little effect on GABA-A receptors, how can it be used in clinical setting and why ?
- to reverse BDZ sedation (overdose)
- due to blocking potentiation of some BDZ agonists
## Footnote
but tricky to manage flumazenil
44
what are the two classes of BDZ ligands ?
- BDZ agonists
- BDZ antagonists
- BDZ inverse agonists
45
why is flumazenil not a perfect antagonist ?
- at some doses can show inverse agonist activity
- at others it can show agonist actvity
| but mostly neutral antagonist
## Footnote
can be tricky to use in clinical setting
46
what are the two compounds that are classified as BDZ inverse agonists ?
- beta-carbolines
- DMCM
47
what are the characteristic properties of BDZ inverse agonists ?
- proconvulsants
- anxiogenic (inc. anxiety)
48
what is the mechanism of action for BDZ inverse agonists ?
- act at same site as flunitrazepam
- competitive (in binding assay)
49
what kind of ligand is flumazenil ?
neutral ligand
50
how can we block the effects of BDZ inverse agonists ?
by use of neutral ligand flumazenil
51
what is the role/ mechanism outcome of BDZ inverse agonists ?
decrease channel opening frequency
52
summary diagram for spectrum of action for BDZ ligands
53
what are the 3 main BDZ ligands to remember and what are their effects?
## Footnote
summary
- Diazepam (BDZ agonist): potentiates GABA, sedation, anxiolytic
- Flumazenil (BDZ antagonist): little effect of GABA, block agonist and inverse agonist
- DMCM (BDZ inverse agonist): decreases GABA activation, causes convulsions
## Footnote
GABA activation : at receptor
54
what is the BDZ binding site on the GABA-A receptor ?
an allosteric site
## Footnote
antagonist/agonist/inverse agonist not completley correct as doesn't actually have a receptor
55
what are the alternative names for the 3 main BDZ ligands ?
- Diazepam: PAM (positive allosteric modulator)
- Flumazenil: SAM (silent allosteric modulators)(or neutral allosteric ligands)
- DMCM: NAM/NCA (negative allosteric modulator/ non-competitive antagonist)
56
what is special about BDZ binding sites ?
they are hetrogenous
## Footnote
done by looking at native brain tissue and pharmacological tools
57
what compound was used to help our understading of BDZ binding sites, and what did this reveal ?
- CL218-872
- 2 distinct CNS populations of BDZ binding sites: Type I and Type II
58
what is the difference between the two types of binding sites found when using the compound CL218-872 ?
Type I - high affinity (for CL compound)
Type II - low affinity (for CL compound)
- Type I and Type II sites had different regional distributions
59
what does Type I and Type II sites differing in regional distribution indicate ?
receptor heterogeneity
60
what does the pharmacology of the BDZ binding site depend on ?
the type of alpha subunit present
61
where do we find the BDZ binding site on the GABA-A receptor ?
interface of alpha and gamma-2 subunit
62
what are some examples of classic BDZs ?
Diazepam, fluniztrazepam
63
how do we get a high affinity sight for classic benzodiazepines ?
- gamma-2 subunit and alpha-1
- Type I receptor
64
what type of receptor does a gamma-2 and alpha-1 subunit give rise to ?
type I
## Footnote
also had high affinity for CL compound
65
how do we get a high affinity sight for classic benzodiazepines (give examples of 'classic' BDZs)?
- gamma-2 and alpha-2/3/5
- Type II receptor
66
what happens if we have a receptor with (gamma-2 and) alpha 4 or alpha 6 ?
- doesn't have high affinity for classic BDZs
- does have high affinity for Ro 15-4513
67
What does the compound Ro 15-4513 also bind to ?
- receptors that have delta instead of gamma which also have alpha -4,6
68
what is Ro 15-4513 ?
synthesised alcohol antagonist
## Footnote
ethical issues around marketing this
69
historically, what were BDZs first line treatments for ?
- anxiety
- sleep disorders
- anticonvulsants
70
why are BDZs used less frequently now ?
due to the fact they can cause dependence and that tolerance can develop quickly
71
what are BDZs used for now ?
- short term treatment of anxiety and/or sleep disorders
- sedatives for minor surgical prodecures
- to aid with alcohol withdrawls
- epilepsy
- muscle spasms
72
what BDZ is typically used as a sedative in minor surgical procedures ?
midazolam
73
what BDZ is typically used to aid alcohol withdrawl ?
chlordiazepoxide
74
what 2 BDZs are typically used to help with epilepsy ?
- clobazam
- clonazepam
75
what BDZ can be used to help treat muscle spasms ?
DIazepam
76
what are 'Z drugs' ?
non-benzodiazepine structures
77
what are 3 examples of Z Drugs ?
- zaleplon
- zopiclone
- zolpidem
78
when are z drugs used and why ?
- to treat sleep disorders
- they are hypnotics or sleeping tablets
- due to short plasma half life (useful to people who have trouble falling asleep)
79
what is a benefit of Z drugs compared to normal BDZs in terms of side effects?
- short plasma half-lives (zapelon: 1 hour, zolpidem: 2.5 hours, zopiclone: 6 hours)
- rapid elimination helps avoid 'hangover' effects seen with older drug like diazepam (48 hour half-life)
80
what is a benefit of Z drugs that is similar to BDZs?
tolerance and dependence developes, but more slowly than (real) BDZs
81
how are Z drugs prescribed ?
at low doses for short perioids of time
e.g. 14-28 days
82
83
what is oppositional tolerance ?
in response to the drug, the brain will 'adjust' its neurochemistry to try to maintain the status quo
84
what happens in oppositional tolerance with BDZs ?
GABAergic transmission is potentiated by BDZs so the reaction of the brain is to down-regulate GABA-ergic transmission
## Footnote
causes tolerance to drug to develop and results in withdrawl symptoms if the person stops taking it abruptly
85
why are BDZs only recommened for short term use ?
tolerance can develope rapidly
## Footnote
normally use for 28 days max. for sleep and anxiety disorders
86
when can withdrawl effects develop especially rapidly ?
BDZs with short half-lives
## Footnote
e.g. with xanax
87
what is the cheical name of xanax ?
alprazolam
88
what are the side effects of withdrawl from BDZs ?
- hallucinations
- increased anxiety
- insomnia
- photophobia
## Footnote
said to be worse than heroin
89
what were some FIRST WAVE BDZ ligands developed and studied to target the anxiety but avoid sedation ?
## Footnote
1990s
- Bretazenil
- Alpidem
- Ocinaplon
90
what did Bretazenil show in the hunt for a BDZ that targeted anxiety and avoided sedation ?
- animal studies suggested anxiolysis but little sedation
however
- human phase I trials: profound sedation and stronger interaction with ethanol than diazepam
91
although BDZs are relatively safe in overdose, when does their risk of death become closer to those of barbiturates ?
when combined with alcohol e.g. ethanol
92
what did Alpidem and Ocinaplon show in the hunt for a BDZ that targeted anxiety and avoided sedation ?
- worked well in animals: anxiolyss with reduced sedation
- maintained in human trials (less sedative tha diazepam)
- never made it to the market: Ocin. failed phase III trials, Alpidem withdrawn after license
BOTH due to liver toxicity
93
how are sedative effects measured in animal models ?
a reduction in motor activity
94
what are 2 behavioural tests used in animal models for measuring anxiety ?
- elevated plus maze
- vogel water-lick conflict test
95
what is a behavioural test used in animal models used to measure motor activity ?
rotorod test
96
how can we abolish the anxiolytic effects of ocinaplon and what does this tell us ?
- with use of flumazenil
- does use BDZ binding site
97
98
What is a hypothetical ancestor of the GABA-A receptor described as?
A homo-pentamer consisting of a single type of subunit arranged around a central ion channel
## Footnote
This ancestral protein is part of the family of ligand-gated ion channels.
99
What are the two faces of subunits in the ancestral GABA-A receptor?
Plus and minus faces
## Footnote
The plus faces are on the clockwise side, and the minus faces are on the anti-clockwise side.
100
How many agonist binding sites did the ancestral GABA-A receptor likely have?
Five agonist binding sites
## Footnote
These sites were located in the extracellular domains.
101
What evolutionary process led to the diversity of subunits in ligand-gated ion channels?
Duplication events
## Footnote
This process allowed for mutations to accumulate without deleterious effects due to redundant copies.
102
In a typical GABA-A receptor, how many copies of beta and alpha subunits are present?
Two copies of beta and two copies of alpha
## Footnote
This configuration contributes to the receptor's function.
103
What do the mutations in the interfaces of GABA-A receptor subunits indicate?
Different evolutionary paths taken by each subunit after duplication
## Footnote
This led to the accumulation of mutations in what were once agonist binding sites.
104
What is required for GABA to bind to the GABA-A receptor?
Amino acid residues on the beta subunit's minus face and the alpha subunit's plus face
## Footnote
There are only two interfaces in the receptor complex with the correct amino acid combination.
105
How many redundant subunit interfaces are present in the GABA-A receptor?
Three redundant subunit interfaces
## Footnote
Originally these would have been agonist binding sites but have accumulated mutations preventing GABA binding.
106
What allows benzodiazepines to bind to the GABA-A receptor?
They fit into a redundant agonist binding site
## Footnote
This site retains some original machinery from the ancestral protein despite being unable to bind GABA.
107
True or False: The GABA-A receptor requires all five binding sites to function properly.
False
## Footnote
The receptor operates well with just two GABA binding sites.
108
109
what are knockout studies used for ?
investigating how a specific protein contributes to a phenotypical feature
## Footnote
investigating the role of the protein
110
what can knockout studies be used to test in terms of BDZ and how is this done?
- to investigate the role of alpha subunits in the sedative and anxiolytic properties of BDZs
- done by eliminating individual subunits from genome
111
what is a common problem of knockout studies ?
compensatory expression of other proteins
## Footnote
shows limited utility of knockout studies as can cause problems
112
what happened when the alpha-6 subunit was knocked out of the GABA-A receptor in animals?
compensatory expression of other GABA-A subunits and K channels
## Footnote
K channels are also inhibitory
113
what happened when the Beta-3 and Gamma-2 subunits were knocked out of the GABA-A receptor in animals ?
animals die; knockout of these subunits was perinatally lethal
114
how can we understand the pharmacological difference of the affinity for BDZ binding in GABA-A subunits ?
examine sequence difference in the subunits
115
what did examining the different sequences of subunits reveal for the GABA-A receptor ?
- for alpha1,2,3,5 (high affinity of classic BDZs) had a conserved histadine residue at position (alpha)105
- alpha4,6 (high affinity for Ro) had an arginine present at position (alpha)105
116
where is the conserved histadine residue ?
in the binding site for BDZs (alpha subunits1,2,3,5)
117
what does the presence of the conserved histadine residue in the binding site for BDZ determine ?
determines that there is high affinity of classic BDZs e.g. diazepam
118
what does the presence of argenine in the alpha subunit determine ?
low affinity for classic BDZs e.g. diazepam
119
what happens if we mutate the conserved histadine residue to an arginine for alpha1,2,3,5 in vitro?
- you get a binding site with low affinity for classic BDZs (e.g. diazepam)
- GABA-A receptor functions normally, no change in GABA sensitivity, no change in receptor assembly
- but complete loss of classic BDZ binding
## Footnote
similar to alpha4,6
120
what is a knock-in study ?
in vivo replacement of single amino acid in a defined gene
121
in a kock in study for GABA-A subunits, how were the mice mutated and what did this result in ?
- mice had histadine to arginine mutation in alpha1,2,3 subunits
- normal phenotypes
- receptors respond same to GABA
## Footnote
same as in vivo
122
what did the knock-in studies reveal about the role of the individual alpha subunits of the GABA-A receptor in the various properties of the BDZ ligands ?
## Footnote
when alpha subunits1,2,3 mutated from histadine -> arginine
- alpha 1: no amnesia or sedation, retained anxiolysis: role in sedation and amnesia
- alpha 2: retained amnesia and sedation, no anxiolysis: role in anxiolysis
- alpha 3: retained all three properties
## Footnote
wt = wildtype
123
what BDZ properties do the alpha 1 subunits in the GABA-A receptors seem to contribute to and when would this be useful?
## Footnote
as a result of knock-in studies
sedative, amnesic actions
- potentially sleeping tablets ? (but anxiolytic effects would be useful)
124
what BDZ properties do the alpha 2 subunits in the GABA-A receptors seem to contribute to and when would this be useful?
## Footnote
as a result of knock-in studies
anxiolytic actions
- anti-anxiety meds without sedation
125
what is a complexity that has arisen through alpha 2 knock in mice ?
## Footnote
knock-in (histadine-arginine)
- shouldn't show anxiolytic effects with any classic BDZ ligands
- but alpha 3 selective compounds did show anxiolytic effects in the alpha 2 knock-in mice
- suggests alpha 3 might be important
126
what is TPA023B ?
## Footnote
attempt of a selective non-BDZ compound
- zero efficacy at alpha 1 but moderate at alpha 2,3
- anxiolytic in rodents and primates but not sedating
- but pahse I human trials found it highly sedating
127
what were the reasons for failure of developing selective (for subunits) BDZ compounds ?
- species differences between rodens/primates and man
- we have only proposed homogeneity of alpha subunits in a receptor (e.g. 2x alpha 2), when it could be heterogenous (e.g. alpha 1 + alpha 3) in humans in a single receptor
## Footnote
therefore we've had little developement for BDZs that are selective for anxiolytic properties rather than sedative properties
128
how does the vogel test work ?
- measures the number of licks of the bottle
- on third day mild shock given every 20 licks
- decreased rate of licking
- but if given anxiolytic drugs there will be no decrease as fear will be diminished
129
what does the Vogel water-lick conflict test measure ?
anxiety
130
what does the elevated plus maze measure ?
anxiety
131
which subunit fo GABA-A receptor contains the primary binding site for barbiturates and how is this shown?
located in the transmembrane domains of the beta subunits, with possibly a smaller contribution from the alpha subunits. Homomeric GABAA receptors formed from just beta subunits (which can't be activated by GABA) can be activated by high concentrations of barbiturates, showing that the beta subunit is the major determinant of barbiturate binding.
