Animal Ethics in Research & Animal Models (Zebrafish, frogs, mice)

Question 1

what are the 3Rs of animal ethics?

Answer 1

1. replacement - using non-animal methods (e.g. cell culture) whenever possible
2. reduction - minimising the number of animals needed for valid results
3. refinement - minimising pain/stress/discomfort for animals during treatment (e.g. local anaesthetic)

Question 2

beyond the 3Rs, what other ethical considerations must be made?

Answer 2

1. justification of using animals = must be scientifically valid (related to replacement principle)
2. ethical reviews = via animal welfare & ethical review bodies, some experiments need ethical approval
3. regulated under legislation - e.g. Animals (Scientific Procedures) Act 1986

Question 3

What makes C. elegans ideal for cell lineage studies?

Answer 3

has an invariant lineage - every worm has 558 cells with the same developmental pattern; each cell’s fate is known

Question 4

advantages of C. elegans (roundworm)?

Answer 4

transparent
easy to culture
short embryogenesis (approx. 15hrs)
invariant cell lineage - easy to trace
fully sequenced compact genome

Question 5

disadvantages of C. elegans (roundworm)?

Answer 5

simple body plan & not a vertebrae - limited applicability to human systems

Question 6

advantages of Drosophila (fruit fly) for genetic studies?

Answer 6

1. powerful genetic tools available for fruit flies
2. short life cycle (approx. 14 days)
3. many conserved developmental genes - homologues between fruit flies & humans
4. easy maintenance & cost-effective
5. high reproduction rate - approx. 50 eggs at a time, can generate multiple generations in a month
6. no/little ethical considerations - not a vertebrate
7. good human disease models - can understand disease mechanisms through them, identify involved genes/pathways

Question 7

disadvantages of Drosophila as a model?

Answer 7

1. not transparent
2. not a vertebrate - dissimilarity to humans with physiology/ body structure/ organ development

Question 8

why are zebrafish embryos excellent for live imaging techniques like WMISH?

Answer 8

transparent embryos - undergo external development

Question 9

advantages of zebrafish as a model?

Answer 9

1. ex utero development
2. transparent embryos
3. rapid embryo development (approx. 72 hours)
4. large numbers of embryos/ clutches produced at once - large sample size, easy to generate multiple generations within days
5. easy maintenance, cost-effective & small
6. genetic similarity to humans - many homologous genes
7. easy genetic manipulation - add chemicals/substances into medium, or genetic manipulation at early stages

Question 10

disadvantages of using zebrafish models in genetic studies?

Answer 10

1. less accessible genetic manipulation than mice - more tools available for them
2. species-specific differences in physiology & metabolism between zebrafish & humans
3. ethical considerations
4. many zebrafish genes are duplicated - hard to isolate the function of specific genes homologous to a human gene

Question 11

advantages of frogs?

Answer 11

1. large embryos - easy to manipulate & can observe consequent development
2. rapid external development
3. easy to maintain
4. high genetic similarity to humans
5. easy to inject & genetically manipulate

Question 12

advantages of frogs/Xenophus?

Answer 12

1. poor genetic tools available for further genetic manipulation - less available compared to zebrafish and mice
   2.has 4 sets of chromosomes - complicates genetic studies
2. takes a long time to reach sexual maturity (1-2 years) = long generational time

Question 13

advantages of chick embryos?

Answer 13

1. large, accessible embryos
2. mammal-like development - share many developmental pathways, good human model
3. embryos are inexpensive, readily available, cost-effective
4. live in-vivo observations possible = live imaging techniques & real-time observation

Question 14

limitations of chick embryos?

Answer 14

1. hard to create transgenics - limited genetic manipulation possible
2. longer development (21 days)
3. harder to access chick embryo through egg shell
4. ethical considerations
5. sensitive to environmental factors - need to be kept in an incubator/ under ideal developmental conditions

Question 15

advantages of mice models?

Answer 15

1. many powerful genetic tools available for genetic manipulation - e.g. knockouts, CRISPR for gene editing…
2. fully sequenced genome
3. genetic similarity to humans (approx. 85% of genes) - can study disease & genetic traits, finding applicable to humans
4. vertebrate development
5. small, cost-effective, easy to keep and breed
6. short lifespan, short gestational period (approx. 21 days), rapid reproduction
7. humanised mice models possible - transplanting human cells/organs into mice to study disease process

Question 16

disadvantages of mice (embryo) models?

Answer 16

1. no transparent development or ex-utero - need to cull the mother to see embryo, less accessible embryo
2. ethical considerations - esp. considering culling the mother
3. genetic background of mouse can influence observed phenotype in genetically engineered mice - hard to isolate effects of a specific gene

Question 17

Zebrafish are advantageous for developmental studies because…
a) Their embryos are opaque
b) They have poor genetics
c) They develop externally and are transparent
d) They lack vertebrate features
e) They require in utero development

Answer 17

c) They develop externally and are transparent

Question 18

Which animal model offers the best invariant cell lineage tracing?
a) Mouse
b) Drosophila
c) Chick
d) C. elegans
e) Zebrafish

Answer 18

d) C. elegans

Question 19

Which model allows high-throughput genetic screening of segmentation defects?
a) Zebrafish
b) Frog
c) Drosophila melanogaster
d) Mouse
e) Chick

Answer 19

c) Drosophila melanogaster ( fruit fly) - cuticle segmentation allows clear readouts of AP patterning defects

Question 19

Which model is most suitable for manipulative experiments on large accessible embryos but has poor genetics?
a) Frog
b) Mouse
c) Chick
d) Zebrafish
e) Drosophila

Answer 19

a) Frog - frog embryos great for manipulation but genetics are limited (few transgenics tools, large duplicated genome makes it harder to edit - physical manipulation is easier over gene editing)

Question 19

Which feature makes chick embryos especially valuable for embryonic manipulations?
a) Transparency
b) Fast genetic transformation
c) External development in large eggs
d) Short gestation
e) Live birth

Answer 19

c) External development in large eggs - 21 days to hatch, fast developmental events (e.g. body axis formation)

Question 20

how is chick embryo development monitored in an egg?

Answer 20

• small hole/ window in egg shell
• inject dyes, DNA or beads through it
• use a glass cover to watch development under a microscope (time lapse for real-time imaging)

Question 21

You want to track the lineage of every cell during embryogenesis using invariant cell divisions. What model organism is best?
a) Zebrafish
b) Mouse
c) C. elegans
d) Frog

Answer 21

c) C. elegans - has invariant lineage (558 cells every time) and full cell fate maps

Question 22

You plan to knock out a gene suspected to be involved in mammalian heart development. Which model is best?
a) Frog
b) Chick
c) Zebrafish
d) Mouse

Answer 22

d) Mouse - best genetic tools for knockouts, CRISPR, and mammalian relevance

Question 23

You need rapid, transparent embryos and want to test the effect of a drug on early organogenesis in real time. a) Mouse b) Chick c) Zebrafish d) Fruit fly

Answer 23

c) Zebrafish - transparent, grows in water, easy chemical manipulation, fast development

Question 24

You want to study anterior-posterior patterning using mutants with segmental body defects. What would you choose? a) Fruit fly b) Frog c) Mouse d) Chick

Answer 24

a) Fruit fly - classic AP patterning via cuticle pattern analysis, Hox genes...

Question 25

You need large embryos for microinjection and want to visualize axis formation with grafting experiments. a) Mouse b) Frog c) Roundworm d) Zebrafish

Answer 25

b) Frog - large, easy-to-manipulate embryos, perfect for grafting and injection