Lectures Flashcards
3 components of Communication Systems in body
- Nervous system
- Endocrine system
- Immune system
three most important communication systems in body
these symptoms are interconnected
forms of communication between nervous system and endocrine system
- hormones
- neurotransmitters
forms of communication between endocrine system and immune system
- hormones
- cytokines
forms of communication between nervous system and immune system
- cytokines
- neurotransmitters
nervous system splits into…
central nervous system and peripheral nervous system
central nervous system
division of the nervous system
structures encased in bone
- brain (cranium)
- spinal cord
peripheral nervous system
division of the nervous system
- autonomic system
- somatic system
autonomic nervous system
responsible for INVOLUNTARY control of the body
somatic nervous system
responsible for CONSCIOUS PERCEPTION and VOLUNTARY MOTOR RESPONSES
autonomic system splits into…
- sympathetic
- fight or flight - parasympathetic
- rest and digest
what kind of neurons for parasympathetic system?
cholinergic
what kind of neurotransmitter for sympathetic system?
norepinephrine
somatic nervous system
subdivision of peripheral nervous system
allows you to move and control muscles
feeds info from 4 senses to brain
2 pathways
four senses whose info are carried by the somatic NS
smell
sound
taste
touch
somatic NS: two pathways of info
- afferent
- efferent
afferent pathway
carries info from SENSORY receptors to the CNS
efferent pathway
carries MOTOR INFO away from the CNS to the MUSCLES/GLANDS
classical definition of behaviour
behaviour results from 3 interacting components
hormones can effect which of the components of behaviour?
all three (input, CNS or output)
3 interacting components which result in behaviour
- input system/sensorial stimulus (internal or external)
- central NS
- output system/peripheral NS (effectors)
superior
above
inferior
below
rostral
front
caudal
back
anterior
in front
posterior
behind
dorsal
back
ventral
front/belly
3 axes we use to slice brain
- coronal
- sagittal
- horizontal
coronal slice
divides front and back of brain
sagittal slice
splits brain into the two hemispheres (along the longitudinal fissure)
horizontal slice
divides top and bottom of brain
entire central nervous system comes from what in the fetus?
neural tube
entire brain comes from this tube
neural tube develops when
3 weeks
3 vesicles of the neural tube
forebrain
midbrain
hindbrain
later on in development, the forebrain divides into…
telencephalon (cerebrum)
diencephalon (lower structures)
telencephalon is composed of…
- cortex
- limbic system
- basal ganglia
diencephalon
- thalamus
- hypothalamus
most of the brain is composed of the…
cerebral cortex
cerebral cortex functions
consciousness, thought, emotion, reasoning, language, memory
cerebral cortex is subdivided into…
4 lobes
frontal, temporal, parietal, occipital
corpus callosum
connects hemispheres of brain
cortex is how thick
3 mm
covers both hemispheres
cortex is composed of…
grey matter
bodies of the neural cells
white matter
axons
means that all the grey matter (cell bodies) are connected all over the brain by the white matter (axons)
DTI
uses water to show the neuron connections and their activity
frontal lobe 4 main functions
- reasoning/executive functioning (PFC)
- motor control (motor cortex)
- emotion
- language (Broca’s area)
where is the motor cortex?
PFC
precentral gyrus
Broca’s area
essential for language production
in frontal lobe
early evidence of importance of frontal lobe in behaviour regulation
case of Phineas Gage
Phineas Gage
railroad construction accident survivor
large iron rod was driven completely through his head
where were the lesions in Phineas Gage’s brain?
widespread lesion of LEFT FRONTAL LOBE
reported effects of Gage’s lesions
affected personality and behaviour
parietal lobe
processes sensory info of the body
contains SOMATOSENSORY CORTEX
somatosensory cortex alternative name
postcentral gyrus
somatosensory cortex is organized in what manner?
topographically
ie. cortical homunculus
temporal lobe main function, and secondary ones
main: processing auditory signals
secondary: learning, memory, language, integrates info from other brain regions
occipital lobe
at back of head
interprets VISUAL info
also involved in balance - cerebellum
boxing anecdote occipital lobe
illegal in boxing to punch at the back of the head
because it can cause loss of vision
subcortical regions
regions other than the frontal, parietal, temporal, and occipital lobes are SUBCORTICAL
4 subcortical regions
- amygdala (fear/anxiety)
- hippocampus (learning/memory)
- hypothalamus (homeostasis - connection with endocrine system)
- thalamus (relay centre of brain)
limbic system is composed of…
amygdala, hippocampus, hypothalamus, thalamus
amygdala has lots of receptors for…
cortisol
reacts to stress
limbic system is where our ____ originate
initial emotional responses
limbic system is located where
between brainstem and neocortex
acts like a bridge between these two areas
has a consensus been reached on what parts of the brain compose the limbic system?
not really
but it’s agreed upon that the amygdala, hippocampus, thalamus and hypothalamus are part of it
what is the structure behind our body FEELING the emotions that we experience?
hypothalamus
limbic system in reaction to scary movie
watching movie
- hypothalamus receives a signal from the amygdala that something frightening is being presented
- hypothalamus triggers the fear/fight or flight response
“the master regulator of the endocrine system”
the hypothalamus
if hippocampus is damaged…
can’t convert STM to LTM (anterograde amnesia)
people feel less fear, hyper-sexuality and mellowness
SM’s amygdala was damaged…
and she couldn’t feel fear as a result
case of HM
suffered from seizures
removed hippocampus in attempt to treat it
function of hippocampus was unknown at the time
seizures stopped, IQ improved
ONE PROBLEM: his memory was gone
- lost most memories of past
- short term mem couldn’t be consolidated
couldn’t form new memories
but could keep it in working memory
before HM, memory was thought to be…
monolithic and stored in the same place in the brain
HM paved way to distinction between STM and LTM
and idea that diff mems are stored in diff areas
hippocampus main function
memory consolidation
STM to LTM
procedural memory relies more on which two structures?
- basal ganglia
- cerebellum
^these two structures were both intact in HM’s brain
2 main parts of the midbrain
substantia nigra
ventral tegmental area
(located deep within the brain)
midbrain is responsible for what three main things?
mood, reward and addiction
hindbrain 3 parts
- medulla
- pons
- cerebellum
medulla
controls automatic processes of ANS
blood, breathing, heart rate
pons
connects hindbrain to rest of brain
means “bridge”
cerebellum
receives msgs from joints and ear structures
controls balance
some procedural memory
two types of neuronal cells
- neurons
- glial cells
glial cells comes from what word
Greek word for “glue”
glial cells were initially thought to only…
- hold neurons in place
- act as supportive cells
now we know that glial cells have a variety of functions, such as…
- providing support to neurons
- repairing damage
- fighting infections
- influencing neurotransmission
3 types of glial cells
- microglia
- oligodendrocyte
- astrocyte
microglia
immune response
homeostasis
supporting neuronal function
oligodendrocyte
produce myelin sheaths that wrap around axons
nurturing and sustaining the environment around axons
astrocyte
neural support
repair damage
regulate neuronal communication
in PNS, oligodendrocytes are called…
schwann cells
in PNS, astrocytes are called…
satellite cells
neurons are _______ cells
excitable
can be excited in certain situations
dendrite
serve as input sites where SIGNALS ARE RECEIVED from other neurons
are different at diff ages
old people have less, and shorter, dendritic spines
stress and dendrites
stress can reduce number of dendritic spines
each dendritic spine is postsynaptic to…
one or two axon terminals
intellectual disability and dendritic spines
spines have diff (abnormal) formations in intellectual disabilities
axons
longest projections from the soma
end at multiple terminal buttons
terminal buttons
where the signal (action potential) finishes
where the neuron communicates with another neuron/other cells
soma
central part of neuron
soma contains…
watery fluid called CYTOSOL
cytosol
watery fluid inside the cell
salty, potassium rich solution
cytosol is separated from the outside of the cell by the…
neuronal membrane
membrane-enclosed structures within the soma
organelles (ribosomes, proteins)
cell body of neuron contains SAME organelles found in all animal cells
nucleus
produces proteins
spherical, centrally located part of the neuron cell
nucleus is contained within a…
DOUBLE MEMBRANE called the nuclear envelope
nuclear envelope
the double membrane that contains the nucleus
it’s perforated by POREES
some hormones act within the neuron nucleus and…
have TRANSCRIPTIONAL properties
nucleus and hormones
some hormones affect the nucleus
bind to receptors
GENE TRANSCRIPTION to create proteins
gene transcription
- RNA molecules are synthesized by RNA polymerase
- then processed into mRNA to carry genetic instructions for protein assembly from nucleus to cytoplasm
- transcription is initiated at promoter region of gene and stopped at terminator region
- initial RNA must be spliced to remove the introns that don’t code for protein
- then are exported from nucleus
membrane
- serves as BARRIER to enclose the cytoplasm inside the neuron
- excludes certain substances that float in the fluid that bathes the neuron
- various types of proteins are embedded within the neuron cell membrane
4 proteins embedded within the membrane of neurons
- ion channels
- transporters
- receptors
- enzymes
ion channels
allow the passage of ions (ie. sodium, potassium, calcium) in and out of the cell
facilitates nerve impulse transmission
transporters
assist in the movement of molecules and ions across the membrane
receptors
bind to specific neurotransmitters or signalling molecules, initiating cellular responses
enzymes
catalyze specific chemical reactions crucial for cellular functions
function of neurons cannot be understood without understanding structure and function of…
membrane and its associated proteins
membrane is important in maintaining difference in…
electrical charge
resting membrane potential
cell is at rest
not generating impulses
inside the cell = negative
outside the cell = positive
how do neurons maintain the difference in electrical charge?
ion channels
sodium, potassium and chloride
sodium and potassium = positively charged
chloride = negatively charged
sodium-potassium pump
uses energy (via ATP -> ADP)
to constantly pump 3 sodium ions OUT of the cell
and 2 potassium ions INTO the cell
uses energy to constantly maintain the difference in electrical charge
sodium-potassium pump helps to keep the electrical charge inside the cell…
less positive than the electrical charge outside the cell
so more negative inside the cell
what percentage of brain’s energy is used to maintain the electrical diff for neurons?
60-70%
very energy-demanding
what happens when neuron receives a stimulus?
rapid reversal of the resting state
the membrane becomes positively charged
the _______ and _______ of action potentials constitute the code used by neurons to transfer info from one location to another
FREQUENCY and PATTERN
in resting state, charge of inside of cell is
-70 mV
while the outside is positive
when neuron receives stimulus, what channels open?
sodium channels
because sodium ions are more concentrated OUTSIDE the cell, they flow inside the cell (concentration gradient)
because sodium is positive, the membrane DEPOLARIZES
repolarization
cell depolarizes with the stimulus
sodium ions enter the cell and make the inside positively charged
reaches a peak of 50 mV
then there is a process of REPOLARIZATION where the neuron returns to resting state
during depolarization, what peak charge does the inside of the cell reach?
50 mV
hyperpolarization
when neuron is too negative
cannot depolarize/fire in this state
refractory period
what channels open during repolarization?
potassium channels
potassium leaves the cell (because of concentration gradient) until it reaches its resting state of -70 mV once more
how does neuron return to resting potential after firing?
sodium-potassium pump
brings 2 potassiums into cell
brings 3 sodiums out of cell
now ready for another action potential
unmyelinated versus myelinated axons
myelinated with nodes of ranvier = faster transmission
signal/action potential jumps from node to node
5 action potential properties
- all same size
- all same duration
- don’t diminish as they move down the axon
- frequency and pattern of action potentials constitute the code used by neurons to transfer info from one location to another
- very rapid - 100 times faster than blink of an eye
how long do action potentials last?
about 2 milliseconds
at the axon hillock…
action potentials undergo summation
potentially is enough to generate the release of neurotransmitters
neural firing chain of 6 events
- neurotransmitter release
- receptor binding
- ion channels open/close
- conductance change causes current flow
- postsynaptic potential changes
- postynaptic ions excited or inhibited
ionotropic vs metabotropic
ionotropic: ligand-gated ion channels
metabotropic: G-protein coupled receptors
4 NT pathways
DANS
- dopamine
- acetylcholine
- norepinephrine
- serotonin
NTs occur in specific pathways, aren’t just randomly distributed across the cortex
what creates the potential in a cell?
the -70 mV difference between the inside and the outside of the cell
this difference in charge creates the potential for action and for rest as well
communication between neurons and glands is very _____, while communication between glands is very _____
fast
slow (because the hormone must travel through the bloodstream to reach the target)
behavioural endocrinology is the study of…
scientific study of the BIDIRECTIONAL interactions between hormones and behaviour
hormones can affect behaviour, and behaviour can influence hormones
3 things that hormones affect which indirectly impact behaviour
they affect individuals’:
- sensory systems
- integrators
- effectors (output systems)
^all three of these are important for behaviour, and hormones affect all of them
hormone
organic chemical messenger
released from endocrine cells
travels through blood system to interact with cells via very specific receptors
endocrine glands
ductless glands from which hormones are released into the blood stream
release hormones in response to specific physiological signals
hormones do what to influence probability of behaviour
- they change gene expression
- or they change rate of cellular function
these two things affect behaviour by increasing the probability that a given behaviour will occur in the presence of a specific stimulus
endocrine cells versus endocrine glands
endocrine cells: ie. fat
- fat releases hormones
endocrine glands:
- produce and release hormones
- can be released by direct innervations or if something binds to the gland (cascade)
the effects of hormones are notable not only in behaviour…
but also in several human ATTRIBUTES and CHARACTERISTICS
3 examples of how hormones affect other human attributes and characteristics
- Lance Armstrong admittedly used EPO, blood transfusions, testosterone and corticosteroids when competing
- Testosterone treatment
- Messi had Growth Hormone Deficiency (GHD) and received GHD injections
testosterone causes…
gain of body mass
higher levels of aggression/paranoia
messes with feedback mechanisms between communication between brain and testes
historical roots of behavioural endocrinology
remarkably multidisciplinary from the very beginning
relationships among endocrine glands, their hormone products and behaviour have been implicitly recognized for centuries
example of relationship between endocrine glands, their hormone products and behaviour
male sex organs (testes) produce and secrete a hormone called testosterone that influences sexual behaviour, aggression, territoriality, as well as other behaviours that differentiate males from females
Alessandro Moreschi
the last known castrato
removing testes causes behavioural changes
Europe in times past - if you were a young boy with a good voice, the Church might castrate you to keep your voice high (castratos)
childlike voice
first method used to determine if a gland affects a behaviour
ablation and replacement
steps of ablation and replacement
- gland that is suspected to be the source of the hormone affecting behaviour is surgically removed
- effects on behaviour are observed
- hormone is replaced by re-implantation, injection of an extract from the gland, or injecting a purified hormone
- determination is made whether the observed consequences of removal are reversed by the hormonal replacement therapy
Berthold’s experiment
first ever hormones and behaviour experiment
had three groups of chicks
group 1: castrated
group 2: castrated and re-implanted
group 3: castrated and transplanted
Berthold’s experiment: group 1
castrated
caponization
- small comb and wattles
- no interest in hens
- no aggression towards other males
Berthold’s experiment: group 2
castrated and reimplantation of testes
normal male development
- normal comb and wattles
- normal male behaviour
Berthold’s experiment: group 3
castration and transplantation of testes
normal male development
- normal comb and wattles
- normal male behaviour
Berthold’s 3 major conclusions
- testes are transplantable organs
- transplanted testes can function and produce sperm
- because testes functioned normally after all nerves were severed, there are no specific nerves directing testicular function
- so certain glands receive info from innervations, but not all
- can conclude that something in the blood is functioning to affect behaviour
most research in behavioural endocrinology involves only a few types of _______ _______. why?
simple behaviour
this narrow focus on only a few behavioural measures is partially a response to the enormous variation in complex behaviours
this approach has advantages and disadvantages
categorize behaviour into two main classes
- description of action
- how questions: allow us to infer causation and role of experience - description of consequence
- why questions: evolution and function
4 things that flow out of “description of action” and “description of consequence”
mechanisms (how question)
development (how question)
evolution (why question)
function (why question)
2 areas of “how questions”
- immediate causation: the physiological mechanisms underlying behaviour
- development: the role of experience in individual behaviour
2 areas of “why questions”
- evolution: the perspective(s) adopted by biologists who assume that evolutionary processes are central to issues in ecology, systematics, and behaviour
- adaptive function: the role of any structural, physiological, or behavioural process that increases an individual’s fitness to survive and reproduce
3 pieces of evidence needed to determine that a particular hormone affected a specific behaviour OR that a specific behaviour changed hormone concentration
- a hormonally-dependent behaviour should disappear when the source of the hormone is removed, or the actions of the hormone are blocked
- after the behaviour stops, restoration of the missing hormonal source or its hormone should reinstate the absent behaviour
- hormone concentrations and the behaviours in question should be covariant: the behaviour should be observed only when hormone concentrations are relatively high and never/rarely when hormone concentrations are low
latency of action of hormones
have a long latency of action
manner in which many hormones are released (and an example)
pusatile manner
ie. secretion of cortisol occurs approximately every 90 minutes
5 classes of evidence for determining hormone-behaviour interactions
- quasi-experimental designs
- comparison groups
- natural observations
- non-random assignment
- use of statistical controls
quasi-experimental designs
inferring causation in a research setting in cases where experimenting would be unethical/difficult
this approach falls between experimental and non-experimental designs
aims to investigate cause-and-effect like experimental design, but often lacks some key elements (like random assignment)
comparison groups
researchers might compare groups that naturally differ in hormone levels (such as individuals with naturally high vs low testosterone) or groups influenced by external factors (such as individuals using hormone replacemeent therapy versus those not using it)
natural observations
these designs often rely on naturally occurring variations in hormone levels rather than manipulating them artificially
try not to influence/bias the quantification
non-random assignment
participants aren’t randomly assigned to groups due to limitations in controlling or manipulating hormone levels in a controlled manner
instead, researchers observe and compare existing groups
try to account for confounders in the analysis - lack of random assignment makes it harder to establish causality
use of statistical controls
quasi-experimental designs employ statistical techniques to control for confounding variables to strengthen the validity of the findings
lack of random assignment makes it harder to establish…
a clear cause-and-effect relationship between hormones and behaviour
correlational designs
examination of relationships between variables without intervening or manipulating them
instead of seeking causation, they aim to IDENTIFY ASSOCIATIONS/CORRELATIONS between variables to understand how they relate to each other
4 traits of correlational designs
- measurement of variables
- assessment of relationships
- no manipulation of variables
- identification of associations
correlational designs: measurement of variables
researchers measure 2+ variables to determine if they’re related
these variables can be anything measurable (height, age, test scores, hormone levels…)
correlational designs: assessment of relationships
statistical analyses, likely correlation coefficients, are used to assess the STRENGTH and DIRECTION of relationships between variables
correlation coefficients range from -1 to 1 (positive, negative or no relationship)
correlational designs: no manipulation of variables
in contrast to experimental designs, correlational studies don’t manipulate variables or control experimental conditions
instead, they observe and analyze existing data
correlational designs: identifying associations
these studies can reveal is changes in one variable = associated with changes in another variable
ie. correlational study might examine whether there’s a relationship between testosterone levels and aggression
longitudinal designs
study changes in variables over an EXTENDED PERIOD within the SAME INDIVIDUALS/GROUPS
unlike cross-sectional studies that capture data at a single point in time, longitudinal studies follow participants over time
6 key aspects of longitudinal designs
- data collection over time
- tracking changes
- identification of trends and patterns
- analysis of stability and change
- challenges and attrition
- resource-intensive
example of longitudinal study: cortisol in pregnant women
measured cortisol during 1st, 2nd and 3rd trimesters
cortisol release in 1st semester = more similar to non-pregnant women
but in 2nd and 3rd trimester, cortisol is produced more during the day
this expression has an association with depressive symptoms
immunoassays
very important technique in behavioural endocrinology
analytical techniques used for the quantification of an analyte (hormone) based on the antigen-antibody reaction
antigen
a molecule that can bind to a specific antibody
antibodies for a given hormone are produced how?
by injecting the hormone of interest (antigen) into an animal to raise an antibody against the hormone
body produces very specific antibodies, and the given antigen will only bind to a certain antibody
bioassay
assays that use LIVING TISSUES/ANIMALS to test effects of hormones/other chemical compounds
useful historically - measured biological response to hormone in question
sometimes determined PRESENCE/ABSENCE of hormone in question
sometimes allowed QUANTITATIVE MEASUREMENT of specific hormones
test the effects of the hormone and measure its biological activity on a living animal
but require much time and labour
bioassay example: pregnant rabbits
relies on human chorionic gonadotropin (hCG)
^a hormone produced during pregnancy
- take pregnant woman’s urine
- inject that urine into the mice
- if the hCG hormone was present in the mice, the animal’s ovaries would enlarge and show follicular maturation
level of precision in the bioassay rabbit example
98%
bioassays: evolution of pregnancy testing
1927-1960: used the animal testing method (urine injection)
- carried out in lab
1960-1988: antibody testing method (blood tests, pregnancy test sticks)
- carried out at the doctor’s office/at home
radioimmunoassay (RIA) is based on the principle of…
competitive binding of an antibody to its antigen (hormone) that results in a change in radioactivity
a radioactive antigen competes with a non-radioactive antigen for a fixed number of antibody binding sites
an antibody produced in response to any antigen (in this case a hormone) has a binding site that is specific for that antigen
antibodies have a given number of binding sites for its antigen
- antigen molecules can be “labeled” with radioactivity
- antibody cannot discriminate between radiolabeled (hot) and normal (cold) antigen
radiolabeled hormone and cold hormone compete for binding sites on antibody
so the more cold hormone there is present in the tube, the less hot hormone will bind to the antibody
quantity of hot hormone that binds can be determined by precipitating the antibody and measuring the associated radioactivity resulting from the radiolabeled hormone that remains bound
concentration of hormone in sample can then be determined by subjecting sample to same procedure and comparing results with standard curve
competitive binding process in radioimmunoassay
several test tubes with different known concentrations of the hormone (antigens that are unlabeled and radioactive labeled) and the antibody are used to BENCHMARK hormone concentration
cold antigen
in radioimmunoassay, the known hormone concentration
hot antigen
in radioimmunoassay, the known RADIOLABELED hormone concentration
what do radioimmunoassays do?
use antibodies to detect and quantitate the amount of antigen (hormone) in a sample
increase precision with which hormone concentrations can be measured
RIA process
in the well, place antibodies
introduce the corresponding antigen
- labelled antigen
- this will bind to the antibodies
then add the unlabelled antigen (ie. from patient serum)
we want to see if the serum has the unlabelled antigen or not
if the sample has a bunch of unlabeled antigens (radioactive), then radioactivity would DECREASE progressively with the increase in antigen concentration
if the sample has a bunch of unlabeled antigens, then radioactivity…
radioactivity would decrease progressively with the increase of the antigen concentration
if there’s a low level of concentration in the sample, radioactivity…
won’t fall that much
because there’s less competition for binding (less labeled antigens are being replaced by unlabeled antigens)
3 main steps of radioimmunoassay
step 1: immune reaction
- radiolabeled antigen
- competition between labeled and unlabeled antigens
step 2: competitive displacement
- radiolabeled antigen released
- unlabeled antigen (hormone) binds
step 3: radioactivity count
- measure radioactivity
enzyme immunoassay (EIA) or enzyme linked immunosorbent assay (ELISA)
most common way to quantify hormones
uses an enzyme instead of radioactivity
ie. Rapid Covid Test
4 main steps in EIA/ELISA
step 1: have a well of captured antibody
- add the antigen that we want to measure (antigen binds to antibody)
step 2: add labelled antibody
- labelled antibody will attached to first set of antibody/antigen
step 3: add the substrate and enzyme
- substrate will attach to the labelled antibody
- enzyme will cause a change in the substrate that will produce a SIGNAL (ie. change in colour)
detect the signal and/or quantify the product
EIA/ELISA can do what two things…
- detect a hormone (ie. through colour change)
- quantify a hormone using a standard curve (gradient of colour)
immunocytochemistry (ICC) versus immunohystochemistry (IHC)
immunocytochemistry: applied to CELLS
immunohystochemistry: applied to TISSUES
both use antibodies to determine the LOCATION of a hormone in a specific tissue/cell
antibodies linked to marker molecules, such as those in fluorescent dye, are usually introduced into dissected tissue from an animal, where they bind with the hormone/neurochemical of interest
immunofluorescence
use fluorescence to determine the location of a hormone in a specific tissue/cell
what does autoradiography do?
determines the location of hormonal action
(location of hormonal uptake and receptor location)
autoradiography basic idea
radiolabeled hormones are injected into a tissue/animal to determine hormonal uptake and indicate receptor location
4 steps of autoradiography
step 1:
- tissues = sliced into several very thin sections
- adjacent sections = subjected to different treatments
step 2:
- one section of target tissue = stained in usual way to highlight various cellular structures
step 3:
- next section = placed in contact with photographic film
- emission of radiation from radiolabeled hormone develops an image on the film
step 4:
- areas of high radioactivity on the film can then be compared with stained section to see how areas of highest hormone concentration correlate with structures
autoradiography basic process
- determine cellular structures
- use radiation emission to determine location of radiolabeled hormone
- overlay the structure with the hormone locations
(dark spots on photo of structures = where radiolabeled hormone has undergone binding)
blot test - western blot
quantifyies several proteins at the same time
technique used to fractionate mixtures of proteins, DNAs or RNAs so they can hybridize with markers that travel different distances in an electrophorectic gel based on their size
electrophoresis: application of an electric current through a matrix/gel that results in a gradient of molecules separating out along the current basis of size (smaller molecules move farther than larger molecules during a set time period)
steps: blot test - Western blot
step 1: get the sample
- tissue of interest is homogenized and the cells are lysed
- makes it so that all the cells’ contents are loose in the fluid
step 2: homogenate is placed in gel and subjected to electrophoresis
- the small electric current applied to the gel separates the proteins based on their electric charges
step 3: transfer gel to a membrane
- protein bands will have formed
step 4: transfer gel to an incubator with a specific antibody
- antibody is specific to the antigen that is being searched for
step 5: detect bound antibody by chemiluminescence
in blot test/Western blot, how do we detect the bound antibody?
chemiluminescence
can quantify the substrate that emits the colourful reaction
in situ hybridization
determining whether a particular substance is produced in a specific tissue
figuring out WHERE the protein/hormone is produced
in situ hybridization follows the principles of…
immunohystochemistry
technique is used to identify cells/tissues in which messenger RNA (mRNA) molecules encoding a specific protein (hormone/neurotransmitter) are being produced
why does in situ hybridization look for mRNA?
it looks for specific sequences of mRNA - because if that mRNA is present in a tissue it means that gene transcription is happening for the protein of interest
because mRNA is what carries the genetic info needed to produce proteins
4 steps of in situ hybridization
step 1: tissue is fixed, sliced very thin, mounted on slides, and either dipped into an emulsion or placed over film
- developed with photographic chemicals
step 2: tissue is also counterstained to identify specific cellular structures
step 3: radiolabeled cDNA probe is introduced into the tissue. if the mRNA of interest is present in the tissue, the cDNA will form a tight association (hybridize) with it
step 4: the tightly bound cDNA, and hence the mRNA, will appear as dark spots
in situ hybridization: the hybridization part
if the mRNA of interest is present in the tissue, the cDNA will form a tight association (hybridize) with it
hybridization will appear as dark spots
immunohistochemistry versus in situ hybridization
IHC: marks protein
ISH: marks RNA associated with gene expression for the same protein
PCR stands for
polymerase chain reaction
applying immunoassays and genetic marking for COVID - PCRs
PCRs use fluorescence
PCRs amplify and replicate a specific segment of DNA and RNA
this process creates NUMEROUS copies of the targeted DNA/RNA
this enables detection and analysis
basically, saliva provides nucleic acids and RNA or DNA - any viral contents will be marked and become fluorescent
the viral mRNA will bind to complementary DNA of the test
then we’ll create a reaction to amplify the signal and we can quantify the presence of the hormone
applying immunoassays and genetic marking for COVID - lateral flow immunoassay technology
in a RAPID ANTIGEN TEST
patient test sample is mixed with chemicals that react with specific proteins on the surface of the SARS-Cov-2 virus
saliva swabs - if mRNA from virus is present, the immune response will occur and colour will change
7 types of immunoassays
- bioassays
- radioimmunoassays
- enzyme immunoassays (EIA) or enzyme linked immunosorbent assay (ELISA)
- autoradiography
- blot tests
- in situ hybridization
bioassay in a nutshell
biological assays assessing the effects of substances
uses living animal tissues
radioummunoassay in a nutshell
measures concentrations of antigens using radioactive labels
concentrations of a given hormone is inversely related to the radioactive labels
measures concentrations using standard curves
enzyme immunoassay (EIA) or enzyme linked immunosorbent assay (ELISA)
detects antigens or antibodies using enzyme-based reactions
immunocytochemistry (ICC)/immunohystochemistry
visualizes proteins in cells (ICC) or tissues (IHC)
does this using immmunohystochemistry
labels antibodies with dyes or enzymes
autoradiography in a nutshell
captures and visualizes radioactively labeled molecules in the tissue
photographic film is overlayed with an adjacent stained tissue
blot wests in a nutshell
techniques (ie. Western blot) for transferring and detecting biomolecules using specific probes
detects several proteins at same time
in situ hybridization in a nutshell
locates and visualizes specific nucleic acid sequences (ie. mRNA) within cells or tissues
pharmacological techniques
use of synthetic chemical agents can alter the action/function of a hormone
two major groups of synthetic chemical agents
2 major groups of synthetic chemical agents
- agonists: simulate endocrine function by affecting hormonal release
- antagonists: inhibit endocrine function by affecting hormonal release (blocking hormones)
example of pharmacological technique
cyproterone acetate -> anti testosterone
cyproterone acetate binds to the testosterone reception
inactivates it
testosterone levels fall
brain imaging 2 techniques
- Positron Emission Tomography (PET)
- Functional Magnetic Resonance Imaging (fMRI)
positron emission tomography
PET
injection of radiotracers - specific molecules (hormone)
PET scanner detects the radioactivity as the compound accumulates in different regions
can infer the locus of the hormone’s action
functional magnetic resonance imaging
fMRI
involves exposing brain to multiple magnetic fields
high spatial and temporal resolution
detects changes in brain activity during specific tasks/conditions
can be task-based or resting-state
gene
discrete region of DNA within a chromosome that when expressed (transcribed), leads to the production of ribonucleic acid (RNA)
transgenic
relating to an animal in which a gene has been INSERTED, ALTERED, or DELETED
