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Techniques In Neuroscience > Lecture Content > Flashcards

Lecture Content Flashcards

1
Q

What is a CT scan?

A

CT scan is a structural scanner that takes images of the human brain by taking multiple X-ray images that then are combined with a computerized algorithm in order to makeup a complete brain image. Non-invasive.

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2
Q

In vivo vs In vitro

A

In the first, studies are performed within the organism, whereas in the latter, tissue/sample has been extracted to be studied in a Petri dish (in glass).

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3
Q

Evolutionary conservation

A

The presence of similar genes and phenotypes, chromosomes, proteins in different species, pointing to a commonality of origin.

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4
Q

What is homology? Types?

A

Homology is similar characteristics between different species. It could be molecular, structural, or functional (processes).

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5
Q

Phylogenetic Tree

A

First drawn by Darwin, prior to his Origin of Species. It represents the evolutionary relationships and patterns between different biological species: their similarities and differences.

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6
Q

What is “Glucose on”?

A

A gene has been turned off in experiments.

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7
Q

What is “Galactose”?

A

A gene has been turned on in experiments.

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8
Q

Forward genetics

A

Molecular genetics approach that aims to determine the genetic origin of a phenotype.

  1. Identify a mutant strain based on a phenotype
  2. Recover a wild-type gene by complimentation

—> using a mutant phenotype to identify (clone) the gene

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9
Q

Reverse genetics

A
  1. Start with a cloned gene
  2. Make directed mutations in the cloned gene
  3. Reintroduce into the organism and investigate the phenotype

—> using the gene to identify the mutant phenotype

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10
Q

What is mitochondria responsible for?

A

Metabolizing energy from food, through an oxidation process, mainly within the muscles, brain, liver, heart, and gastrointestinal tract.

*Switching off the mitochondria in dopaminergic neurons has been linked to Parkinson’s.

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11
Q

Neural circuit

A

A population of neurons interconnected by synapses that carry out a specific function when activated.

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12
Q

Electrophysiology

A

Branch of neuroscience that explores the electrical activity of living neurons and investigates the molecular and cellular processes that govern their signaling. A dynamic, functional, SI unit-based, real-time, hi-fidelity and high temporal resolution approach. Many electrophysiological approaches can be used vivo. Electrophysiology can be used simultaneously or in conjunction with optical, molecular, biochemical and pharmacological techniques

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13
Q

How do neurons communicate? (Types of signals)

A

Electrically and chemically

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14
Q

Extracellular recording

A

The electrode is placed just outside the neuron of interest.

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15
Q

Intracellular recording

A

The electrode is inserted into the neuron of interest.

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16
Q

Patch clamp recording

A

The electrode is closely apposed to the neuronal membrane, forming a tight seal with a patch of the membrane.

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17
Q

Concentration gradient

A

The difference of ion concentration inside the cell compared to outside the cell.

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18
Q

What is chemical gradient? (See image below)

A

The ion separate which occurs right at the membrane of the neuron: more K+ (potassium) ions on the inside of the membrane and more Na+ (sodium) ions on the outside.

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19
Q

What is electrical gradient? (See image below)

A

The difference in charge across the cell membrane. Ions have an electrical charge. At rest, there are more positive ions on the outside than the inside of the cell membrane.

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20
Q

Electrochemical gradient

A

The ionic imbalance represented by the electrical and chemical gradients at the membrane of the neural cell.

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21
Q

Measuring unit for the electrical potential of the neural cell?

A

Voltage

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22
Q

What’s the voltage of the Resting Potential of the membrane?

A

-70mV

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23
Q

How do we call the movement of charge over time?

A

Current (I)

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24
Q

What limits the movement of ions across the membrane?

A

The membrane Resistance (R)

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25
How is Resistance generated?
By properties of the membrane, such as how many channels are open or closed.
26
What is Ohm’s law?
V=IxR. It represents the relationship among the membrane potential (V), the current flow (I), and the membrane resistance (R).
27
Resting membrane potential
The voltage difference across the membrane of a neuron when it is at rest (non-signalling). In a neuron, the inside of the cell is about 70 mVs (millivolts) less than the outside, so the resting membrane potential is said to be -70 mVs.
28
Depolarisation
A change in a neurons membrane potential that make it more positive (less negative).
29
Hyperpolarization
A change in a neurons membrane potential that makes it more negative. It is the opposite of depolarization.
30
Ionotropic receptors
Transmembrane proteins that form a channel allowing ions to travel in or out of a cell. These channels are opened when the receptor binds a ligand, like a neurotransmitter. Glutamate receptors and GABAA receptors are examples of ionotropic receptors.
31
Voltage-gated ion channels
Transmembrane proteins that form ion channels whose opening and closing is regulated by the membrane potential near the channel.
32
Action potential
The process by which a neuron sends information down its axon. Also referred to as a spike, is an all-or-none, rapid, tran- sient depolarization of the neuron’s membrane
33
At what V do these occur?
34
What is graded potential?
Local voltage change. Small charge.
35
How do we call a membrane potential that is more positive?
Excitatory postsynaptic potential (EPSP)
36
How do we call a membrane potential that is more negative?
Inhibitory postsynaptic potential (IPSP)
37
Field potential
This is the electric potential in the extracellular space around neurons.
38
What is the nerve comprised of?
A bundle of axons.
39
Compound axon potential
The sum of the activity in a number of nerve fibers (or axons).
40
Ways in which Extracellular recording (ER) is applied:
.field potentials • whole nerve activity • multi-unit activity • single unit activity • multi-electrode arrays (MEAs) *in each case: • the electrode is outside but close to the neurons • the electrodes pick up only field potentials and low frequency filtered action potentials • it is not possible to record Vm rest or post-synaptic potentials
41
Ways in which Intracellular recording (IR)is applied:
• activity within single cells • sharp electrodes • patch suction electrodes
42
Ways in which Single channel recording (SCR)is applied:
• recording activity of single ion channels • patch clamp-type electrodes
43
Protein tertiary structure
The three- dimensional structure of a protein.
44
Cross linking fixatives:
*formaldehyde and glutaraldehyde: create covalent bonds between proteins in the tissue *aldehydes: anchor proteins
45
Precipitating fixatives:
* ethanol and methanol: disrupt hydrophobic bonds between proteins causing them to irreversibly precipitate
46
Hydrophobic bonds:
These arise from the interaction of their hydrophobic (water-disliking) amino acids with water.
47
Immersion fixation:
Fresh tissue is placed in fixing fluid and gently agitated.
48
Perfusion fixation:
Injection of fixing fluid into the circulatory system.
49
Factors affecting quality of fixation:
50
Cryopreservation
Preservation of tissue structure and components by freezing them rapidly without fixation.
51
Processing fixed tissue to paraffin wax (steps):
52
What is sectioning in histology?
The process of cutting thin slices from the sample, which are required for microscopic examination.
53
What is microtome used for?
Used for sectioning fixed specimens samples.
54
Types of microtomes:
* benchtop rotary microtome (paraffin fix) * sledge microtome (paraffin fix) * vibratome (agarose or gelatine fix) * cryostat (snap-frozen tissues) *sliding (snap-frozen tissues)- cheaper than cryostat
55
What type of dye is Luxol Fast Blue?
Acidic dye in solution that is used to visualise central nervous system myelin sheaths in paraffin wax sections.
56
Myelin sheaths
Formed by oligodendrocytes in the central nervous system. Composed of cholesterol and glycoproteins, myelin serves to insulate the axons of neurons, which significantly increases the conduction velocity of action potential.
57
Counterstain
Either produces a contrasting background colour to the main stain or provides complementary information.
58
Golgi stain
A metal precipitation, developed by Camillo Golgi, and modified by Santiago Ramon y Cajal. *procedure: small piece of formalin fixed tissue is immersed in potassium chromate, then silver nitrate. *purpose: to stain small, random subsets of neurons in exquisite detail. To illustrate neuronal network. (In studies of schizophrenia)
59
Antibodies
Y-shaped proteins used by the immune system to identify and protect against unwanted foreign substances such as viruses and bacteria.
60
Antigen
Molecule (toxin or another foreign substance) which induces an immune response in our bodies, in particular, the production of antibodies.
61
Epitope
Small sequence or part of the antigen recognised by the antibody.
62
Immunohistochemistry
A method of identifying specific antigens in tissues, by immunostaining techniques, using the fact that antibodies bind specifically to antigens.
63
Immunofluorescence
A technique for determining the location of an antigen (or antibody) in tissues by reaction with an antibody (or antigen) labeled with a fluorescent dye.
64
Antigen unmasking/retrieval
Technique where the masking of an epitope is reversed so that the antibody can again bind to it.
65
Immersion (histology)
Placing small brains, or even entire animals, in fixative solutions.
66
Perfusion (histology)
Delivering a fixative through an animal’s cardiovascular system.
67
Neuroanatomical planes (sectioning):
*coronal (A) *sagittal (B) *horizontal/transverse (C)
68
Enzyme
Biomolecule capable of catalyzing a biological reaction.
69
Reporter gene
Nonendogenous gene encoding an enzyme or fluorescent protein whose expression is controlled by a promoter for a separate gene of interest.
70
Neuronal tracer
Chemical probe that labels axon paths to illuminate connec- tivity in the nervous system. Tracers are described by the direction they travel in a neuron.
71
Types of neuronal tracers
*Anterograde: transport from the cell body through the axon to the presynaptic terminal *Retrograde: the opposite direction * Transsynaptic: cross synapses and label multiple neurons in a circuit.
72
What is tissue culture:
Cultivation of eukaryotic tissues outside of the organism, in a growth media with the necessary nutrients, inorganic salts and pH required to function in a physiologically normal manner.
73
What is cell culture:
Culturing of dissociated cells rather than pieces of tissue.
74
Clinical applications of tissue culture study:
*diagnosis of chromosomal disorders from the culture of blood or amniotic fluid samples *generation of monoclonal antibodies for the production of vaccines as a result of the development of hybridoma cell lines by Kohler & Milstein, 1975. * in vitro fertilisation, through techniques developed for the culture of the early embryo, and first achieved by Patrick Steptoe & Robert Edwards (1977).
75
What is Wilhelm Roux responsible for?
Maintained the neural folds from early chick embryos in a saline solution (1885).
76
What is Ross Granville Harrison responsible for?
Removed small section of frog embryos and embedded them in blood clots on the underside of coverslips to allow microscopic evaluation (1907).
77
What are Alexis Carrel & Montrose Thomas Burrows responsible for?
Generated the first ‘cell line’ from embryonic chicken heart (1911).
78
What are Margaret Reed Lewis & Warren H. Lewis responsible for?
Made first defined liquid media to try and overcome this problem (1911): * grew embryonic chick tissue in a relatively simple defined liquid media *nowadays, cell types are still typically grown with media containing serum
79
What are Francis Peyton Rous & F.S. Jones responsible for?
First demonstrated the use of proteolytic enzyme trypsin to dissociate tissues into individual cells for culture (1916).
80
Specifics of extracellular matrix (ECM):
*Most cell types, with the exception of blood cells, grow attached to an extracellular matrix (ECM). * The extracellular matrix is composed of a complex mixture of polysaccharides and proteins such as collagens and laminin. * Tissue culture vessels coated with purified of unpurified components of the ECM help to support attachment and normal functioning of many types of adherent cells. * Cell adhesion molecules on the surface of many cells bind strongly to components of the ECM. * The use of trypsin also allowed for the re-plating of cells grown attached to a substrate.
81
What substrates are being used to dissociate tissues into single cells?
*Trypsin * Accutase: gentler enzymes that result in reduced cell death *EDTA: non-enzyme that result in reduced cell death
82
What are Alexis Carrel & Lillian E. Baker responsible for?
83
What makes the carrel flask particularly useful?
84
Contact inhibition
Stops the cell from dividing further when there’s no longer space, and can alter the characteristics of the cells under investigation.
85
What is mycoplasma?
The smallest known bacterial cells, making them very difficult to observe under conventional light microscopes. * can be a significant problem in long-term culture *some antibiotics are effective against mycoplasmas *best practice is to prevent contamination by employing an aseptic technique
86
Characteristics of Class I cabinets:
87
Characteristics of Class II cabinets:
88
Characteristics of Class III cabinets:
89
Characteristics of Tissue culture incubators:
90
What are Leonard Hayflick & Paul Moorhead responsible for?
Derived the first strains of human fibroblasts (WI-38) (1961). Made the distinction between primary cells, cell lines and cell strains.
91
What’s the distinction between primary cells, cell lines and cell strains?
92
What is “karyotype”?
The size, shape, and number of chromosomes in a cell.
93
What is George Otto Gey responsible for?
Immortal human cell lines (1951). Cultured cells from an individual called Henrietta Lacks who had cervical cancer, and discovered that cells derived from the cervical tumour could grow and divide indefinitely.
94
What are HeLa cells?
Named after Henrietta Lacks: cervical cancer “immortal human cells”. * aided the development of the first Polio vaccine *still being used for research today
95
What is Martin Evans responsible for?
Isolated embryonic stem cell from mouse blastocysts (1982): • can in principle generate any cell type of the body in a cell culture dish
96
What is Jamie Thomson responsible for?
First generated human embryonic stem cells from human blastocysts (1998): * allowed for the generation of inaccessible cell types, such as neurons, in large numbers for the first time *ethical issues need to be considered
97
What is Shinya Yamanaka responsible for?
Directly generated embryonic stem cells by directly manipulating fibroblasts in a process called reprogramming (2006, 2007) *produced the so-called induced pluripotent stem cells (iPSCs) *used to study genetic diseases and inaccessible cell types *no ethical issues as those surrounding embryonic stem cells
98
What is a plasmid?
A circular piece of DNA; a vector that we choose to express gene in cells: the plasmid contains the gene for the protein you are interested in, as well as promoter for that gene.
99
What is a Promoter:
Where transcription of a gene is initiated.
100
What is Transcription:
When a section of DNA is copied to make RNA. RNA may then be used as a template for making a protein.
101
What is cDNA (complementary DNA)?
DNA that is synthesised by using RNA as a template. cDNA is often used to clone eukaryotic genes in prokaryotes.
102
Prokaryote:
A unicellular organism that lacks a membrane bound nucleus, like a bacterium.
103
Eukaryotic cells:
Have a membrane bound nucleus that houses the genetic material. The cells that make up our body are eukaryotic cells.
104
What are Introns?
Sections of DNA or RNA that do not code for proteins.
105
Up/downstream sequences:
A gene has two ends, a 5′ end and a 3′ end. Upstream refers to sequences that are towards the 5′ end, and downstream refers to sequence that are towards the 3′ end. These sequences are involved in the control of transcription.
106
Gene isoforms:
Different versions of RNA transcripts made from the same gene.

Techniques In Neuroscience (1 decks)

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