Cellular and molecular neurobiology

09.25 (kézi jegyzet gépelve)

Outlines:

 neurotransmission systems

 signal transduction

 plastic at any time: different levels of synaptic plasticity

 neurodevelopment

 neurodegeneration (eg. autism is a neurodevelopment disease + psychiatric diseases eg.

schizophrenia)

mariateresa.fiorenza@uniroma1.it

Synaptic transmission

Calcium-dependent neurotransmitter

Exocytosis: quantic neurotransmitter release (kiss and run connection)

Ionic channel activation

Targeting different aspects of synaptic transmission

Neurotransmitters:

 receptors, and signal transduction

 synthesis, activity and inactivation of the brain, under the point of view of chemical
neurotransmission is largely a glutamatergic …

Monoamines:

 catecholamines (epinephrine, norepinephrine)

 dopamine

 acetylcholine

 serotonin

09.27

Similarities among diffuse modulatory systems

 each system has a small core of neurons (only a fe thousands)

 most of the cores are found in the central core of the brain and brain stem

 each neuron from the core can influence more than 100.000 postsynaptic neurons spread all
over the brain

 the synapses are not terminal but rather run along axons (called boutons en passant)

 each system only modulates the actions of other neurons and does not turn them on or off
  activity: like adjusting the volume on a radio insteead of the power

Boutons en passant

 neurons from the core meander through the brain, and when activated, release
neurotransmitter from thousands of sites along the axon

 any neuron in close proximity is affected

fundamentals for a worthwhile attainment of this course

 amino acids: different side chains, R determine the properties of 20 amino acids (amino
group, carboxylic acid group, H, R, C in the middle with 4 kovalens bounds) -> each protein
has a unique structure (fehérjeszerkezetek 4 típusa!) - hidrofil és hidrofób fehérjék; a-helix
és ß-sheet

Protein folding

 occurs in the cytosol

 involves localized spatial interaction among primary structure elements, i.e. the amino acids

 yields secondary structure

Secondary structure:

 non-linear

 3 dimensional

 localized to regions of an amino acid chain

 formed and stabilized by hydrogen bonding, electrostatic and van der waals interactions

Tertiary structure

Quaternary structure

 non-linear

 3 dimensional

Two main types of neurotransmitter receptors

 ion-channel receptors: ligand-gated or voltage-gated

 G-protein-coupled receptors (GPCR), also named metabotropic receptors

 GCPR exists for each neurotransmitter

 acetylcholine

 biogenic amines (dopamine, etc)

GABA receptor

 pentameric structure (ie. made of five subunits)

  each subunit is an amino acid channel -> each subunit is a protein, and each subunit has 4
transmembrane unit

 plasma membrane (extra- and intracellular)

 the GABA-A receptor is subjected to phosphorylation by PKA e PKC, as a means to modulate

its activity

The nicotinic receptor

 agonist: nicotin

 antagonist: d-tubocurarin

 conformational change of the channel gate, channel opening leads to the movement of
positively charged ions -> exci

Typical structure of a metabotropic (GPCR – G-protein caplin receptor) receptor

 seven transmembrane domains

10.02 (kézi jegyzet gépelve)

GPCR exists for each neurotransmitter

 acetylcholine

 biogenic amines

o dopamine, norepinephrine

o epinephrine, serotonin

o histamine

 amino acids

o glutamate

o glycine: no GPCR

o GABA

 ATP, adenine

Ionic channel activation

the acetylcholine nicotinic receptor

GABA-A receptor

the nicotinic receptor

The neuron and the action potential

The resting potential

Contribution of Active Transport

Contribution of facilitation diffusion

Action Potential

Myelinated axons

G-protein coupled receptors

Metabotropic receptor

(indirect mechanism of activity of a neurotransmitter: the metabotropic receptor)

2012: Nobel prize, Lefkowitz and Kobilka

Metabotropic receptors: the cascade of events

10.04 és 10.09 - ?

10.11

Classical neurotransmitters

10. 16 – nem voltam

10. 18 – nem voltam

10. 23

November 22 – midterm

The noradrenergic system

 neurons from this system release norepinephrine (NE)

 the core of this system is called the locus coeruleus (meaning “blue spot”) and is located in
the pons

o each locus coeruleus (there is one on each side) contains ~12.500 neurons

 destination: nearly everywhere in brain (celebral cortex, cerebellum, thalamus,

hypothalamus, spinal cord, etc.)

 noradrenalin synthesis, clearance

 all these receptor types are expressed at the level of the prefrontal cortex, but their
activation depends on the level of the noradrenalin

 working memory -> on the network of the prefrontal cortex

Adrenergic receptors

 belong to family of G-protein coupled-receptors

 endogenous agonists: epinephrine (E), norepinephrine (NE)

 major source of E: adrenal medulla

 major source of NE:

We keep stressful events in our memories, this help us recalling it and it’s important for our survival
behavioral stress adaptation is modulated by the autonomic nervous system, which releases
noradrenaline

CANNON FÉLE VÉSZREAKCIÓT ÁTNÉZNI!!!

Negative feedback -

Serotonergic System

serotonin is an enigma. it is at once implicated in virtually everything but responsible for nothing.

 although serotonin has been studied extensively, its precise role in the brain remains poorly
understood

 at least in part because selective pharmacologic tool for each subtype of serotonin receptor

The system

 neurons from this system release serotonin (5-HT)

 this system cores are located in the nine Raphe nuclei (located in the brain stem)

 destination: everywhere in the central nervous system

 superior Raphe project up to the cortex

 Inferior Raphe project down the spinal cord

 tryptophan AA and hidroxiláz enzim -> production of serotonin (5-HT)

Serotonin receptors

 Cell-body autoreceptor (reduces neutronal fining)

 Terminal autoreceptor (reduces 5HT release)

 postsynaptic receptors

 each family includes various subtype, ie. 5-HT1 subtypes are: 5HT1a, 5HT1b, 5HT1c and
5HT1d

 5-HT3 is unique in that it does not couple to G protein receptors, instead, it is a ligand-gated
cation channel

Serotoninergic synapse

 the same drugs that disrupt catecholamine storage by inhibiting VMAT (i.e. reseprin) also
disrupt serotonin storage and cause profound serotonin depletion

 the reuptake of synaptic serotonin into serotoninergic nerve terminals represents the most
important mechanism by which serotonin signal is terminated

 it is mediated by SERT, a large transmembrane

Ecstasy

 NMDA is a substrate for the SERT and also has a high affinity for 5-HT2 receptors. These two
actions cause a mdoarate elevation of the mood and mild perceptual alterations
  side effect of its use includes: tachycardia, agiation, hyperthermia, and panic attacks

 NMDA has been proven to selectively destroy serotoninergic axons and cell bodies in rodents
and nonhuman primates. After it is transported into serotoninergic neurons, by means of
SERT, NMDA produces oxidative stress

 Therefore, NMDA leads to a loss of serotoninergic neurons, which explains the tolerance to
some NMDA euphorigenic effect that occurs with repeated use

SERT

 a unique feature of SERT is its large number of potential phosphorylation sites; it has eight
such sites, whereas the NET has only one

 these phosphorylation sites tightly regulate the activity of SERT by producing short-term
changes of the serotonin transport into neurons. SERT is highly phosphorylated by both
protein kinase A and protein kinase C;

 Upon phosphorylation by PKC, the SERT gets internalized, leading to a reduction of serotonin
reuptake

 SSRI require several weeks of repeated administration to produce therapeutic benefits.

Why?

Details on 5-HT receptors

5-HT1A, 5-HT1B (Gi/O); 5-HT2A, 5-HT2C (Gq); 5-HT4 (Gs); and 5-HT3 (Na+ and Ca2+ conducting ligand
gated ion channel)

10.25

 besides the large receptor family, there is additional means to increase functional variability
of receptors

 RNA editing (epigenetic molecular mechanism)

 it depends on enzymatic activities that modify some nucleotides of RNA molecules

 epigenetics: a set of regulatory mechanisms that influence game expression above the level
of genome without changes in gene sequence

RNA editing

 nukleotid (pl. adenozin) can be modified adenozin -> guanozin RÁKERESNI HOGY VAN
ANGOLUL

 the de-amination of adenosine, catalyzed by adenosine de-aminase acting of RNA, ADAR,

transforms adenosine into inosine. Inosine is read as a Guanosine, determining a change in a
given codon, which is reflected on the aminoacid specified by the codon

 ADAR e-code mRNA-encoding for serotoninergic, glutamatergic receprors, as well as K+

channels. ADAR are abundantly expressed in our CNS? thus regulating the generation of
electrical signals and signal transduction

 nuclear pre-mRNA editing by selective adenosine deamination is catalyzed by a family of

enzymes known as Adenosine Deaminases Acting on RNA
Serotonin 2C receptor

 the 2c subtype of serotonin receptor, a member of the G protein-coupled receptor

superfamily, undergoes A-toI RNA editing at five editing sites

 all five editing sites reside closely in the second intracellular loop of the receptor

 non-coding RNA: do not encode for any protein

 five adenosine (A) of the o 5-HT2C may undergo RNA editing

 as a consequence, the level of activation upon serotonin binding to the receptor varies

 when all five “A” are edited, the activation of the receptor displays a 20-fold reduction

 RNS és DNS vegyületeket átnézni!!!

RNA editing: summary

 nuclear pre-mRNA editing by selective adenosine deamination is catalyzed by a family

Epigenetic layers at work: mechanisms controlling the expression of SERT (serotonin transporter)

 polymorphism, DNA sequences

Glutamatergic system

 functional classes (ionotropic/metapotropic)

 gene families

 ligand-gated ion channels, second messenger system

 NMDA receptor: 4 diff subunits (Nr1, Nr2A, NR2B)

 group, receptor, subunit, gene, locus

 starting point: aminoacid

10.30

Glutamate Receptors

Structure of glutamate receptors

 even a small aminoacid change will affect the sequence

Ion selectivity of Glutatamte Receptor Channels

AMPA receptors containing GluA2 are Ca2+ impermeable due to editing at the Q/Rsite

NMDA receptors: physiology and pharmacology

 NMDA receptors have several properties that set them apart from other ligand-gated
receptors. At membrane potentials, more negative than approximately -50 mV, the
concentraton of Mg2+ ions in the extracellular fluid of the brain is sufficient to virtually
 abolish ion flux through NMDA receptor channels, even in the presence of glutamate. Thus,
at resting membrane potentials of about -70 mV, the activation of these receptors result in
little current flow, even when glutamate or asparate is bound to the receptor, because the
entry of Mg2+ ions into the channel pore blocks the movement of monovalent ions across
the channel

 the activation of NMDA receptors, like that of AMPA receptors, produces a nonspecific
increase in permeability

 permeable to K+, NA+ and Ca+

 (upon entrance into the neuron, Ca2+ ions act as second messenger, influencing the cellular
metabolism)

 the NMDA receptor channel is typically blocked by Mg2+ ions, even after the glutamate
binding

 too much Ca2+ is dangerous

 this mechanism lies on the NMDA receptor’s work

 NMDA receptor is unique among all neurotransmitter receptors in that its activation requires
the simultaneous binding of two different agonists: glutamate and glycine

 cycloserine important in the treatment of schizophrenia

Dendritic spines

Synapse morphological changes induced by sustained NMDA stimulation

Spine morphology changes over time

the spine is an electrical and biochemical micro compartment

11.06

The GABAergic synapse

The GABA-a receptor (ion channel reception type)

Benzodiazepines

Barbiturates

Ethanol – no specific side for GABA-type receptors

synthesis of acetylcholine

Neuropeptides

11.08

Neuropeptides

 chain of 5 to 35 aminoacids

 act al lower concentrations than other neurotransmitters

 longer lasting effects

  stored in axon as terminal as larger secretory granules (called debse-core vesicles)

 some functions as hormones or neuromodulators

 some also released from digestive tract (gut-brain peptides cause food cravings)

Neuropeptides vs. classic neurotransmitters

 neuropeptides are packaged separately, inside secretory granules and released by different
mechanisms from many parts of a neuron

 they evoke specific effects on behavior

Neuropeptide synthesis and transport

Release of neuropeptides

 Dense core vesicles are transported down the axon to terminals

 DSVs are docked outside the active zone

 Released upon large, sustained calcium entry into the cell

 high frequency action potentials, burst firing

 recent hypotheses suggest that classical neurotransmitters convey information between

pairs of neurons whereas neuropeptides convey information and coordinate activity across
broader networks of neurons

Neuropeptides co-localize with classic neurotransmitters

(diát bevágni)

Classical neurotransmission vs. peptide transmitters

Classic transmitters:

 released at lower firing rates

 mediate

Neuropeptides:

 released at higher firing rates and particularly under burst-firing patterns

 mediate slower neurotransmission (metabotropic receptor activation = slower)

 degradation after release

 must be synthesized in cell body and transported to the terminal

 stored in large dense core vesicles (100 nm)

Hypothalamo-hypophyseal connection

 some hypothalamic neurons release the content that neuropeptides local blood circulation
connects, they sitmulate neurons in the anterior part

Hypothalamus
  most important structure of homeostasis

 contains neutral center for hunger, thirst, sexual drive

Ventral Tegmental Area (VTA)

Endocannabinoid Signaling in the Control of Social Behavior

 Anandamide

Medical use of cannabis and its derivatives

 when your endocannabinoid system is not working properly, you may have an imbalance,
which can manifest as a medical condition

 there are no pharmaceutical medications that directly address an endocannabinoid

dysfunction

 the cannabis plant is a natural medicine that can help balance the endocannabinoid system
for a number of

Phytocannabinoids

 delta-9-tetrahydrocannabinol (THC)

 marijuana THC 0,5-14%

 hashish THC 2-20%

 hashish oil THC 15-50%

 they are lipid, not neurotransmitters

The endocannabinoid system

 in the early 1990s research scientists discussed and identified the endocannabinoid system

 THC, causes a mental state described as a combination of enhanced sociability, quickened

mental associations, increased appetite for sweets and fatty food, alterations in the
perception of time and space, and heightened sensitivity to certain sensory stimuli

 important for stress release

 cannabis sativa receptor

 the word endocannabinoid is used in medical literature

 ECS modulates energy balance and metabolism through central and peripheral mechanisms

Endocannabinoids and their receptors

 CB1 (1990) and CB2 =1993) receptors were discovered as “orphan receptors”

 CB1 are highly expressed in: hippocampus, amygdala, basal ganglia, cortex, cerebellum

 CB2 are mostly expressed at microglia and outside the brain

 the e.c. system consists of lipid-derived messengers that act on G-protein-coupled

cannabinoid receptors
  e.c. transmitters have a unique set of properties:

o they act as retrograde synaptic signals or local modulators to control presynaptic

firing receptors are localized pre-synaptically on the surface of both excitatory and
inhibitory neurons

o of-note, 2-AG may primarly serve as a point-to-point retrograde signal, whereas

anandamide may act as a local modulator

o they act as lipid mediators: endocannabinoids are not stored in vesicles but are
instead ‘demobilized’ in phospholipid membranes under baseline conditions to
become ‘mobilized’ on demand during signaling activity

 they are neuropeptides, but they are lipids

 when anandamide released and activate the Cb1 receptor, the CB1 sitting on glutamatergic
terminal, and the glutamate release is inhibited

 CB1 receptor is the most anandubed repector in our brain

Brain areas expressing CB1 receptors

Synthesis and degradation

Endocannabinoid system

 ligands (endocannabinoids) + receptors (cannabinoid receptors) CB1 and CB2

 ligands are ‘neurotransmitters’: anandamide and

 the e.c. system modulates rewarding properties of food by acting at specific mesolimbic
areas in the brain

 controlling appetite and food intake

 obesity seems to be a condition, in which an overactivation of the endocannabinoid system

occurs, and therefore drugs interfering with this overactivation of the endocannabinoid
system

Endocannabinoid control of food intake and energy balance

The endocannabinoid system in neurons

 Leptin: neuropeptide, produced by adezitides

 leptin signaling can influence 2-AG biosynthesis in the hypothalamus and anandamide
hydrolysis in T-lymphocytes

 signaling energy storage, important perifering singlaing blocking the activity of

endocannabidoids

11.13

Retrogade signaling by endocannabinoids negatively regulates the neurotransmitter release in

Gluergic and GABAergic synapses

 transporter release at the level of synapses

Endocannabinoid signaling induces synaptic depression at excitatory synapses

neurotransmitters (endocannabinoids) are synthesized at the presynaptic side, we start at the

postsynaptic site

Depolarization-induced suppression of inhibition

 we are considering that the post synaptic side, producing endocannabinoids and on the
presynaptic side (GABAergic interneuron, releases GABA) the production are modulated by
ligand-gated channels

 Endocannabinoids act on the cb1 receptor, which will inhibit further GABA release

 GABA-release is suppressed, which means inhibition is suppressed

 they play an important role in depolarization induced suppression of excitation

 CB1 receptor is the most diffuse metabotropic receptor in our brain

 act in the level of presynaptic terminals

 always find depolarization in use

Glyal neurotransmission

Astrocytic CB1Rs modulation of synaptic transmission

Short-term plasticity

Opioid system

 opium, extracted from poppy seeds (papaver somniferum), has poweful pain-relieving
properties and produces euphoria

 They used to be used as painkillers

 Morphine, named after the god Morpheus, is the most active ingredtient of opium

 Heroin was synthesized chemically by morphine deacetylation, commercialized to treat

asthma and coughing

 opiates have long ranked among the most important drugs in the pharmacopoeia.

 Commonly used today, morphine is one of a small number that were available in the
nineteenth century

 the great need for opiate analgesics, combined with the serious risks of developing
dependence and addiction that they pose, has produced a continuing search for nnon
addictive form of these drugs

 to date, it has not been possible to separate the most effective aspects of opiate analgesia
from the addictive properties of these drugs

 opiates, a subclass of opioids that are natural derivates of the opium plant, papaver
somniferum, include morphine and codeine, which are the two major metabolites of herion

 they can motivate repeated self-administration, produce tolerance, dependence, and

ultimately opioid addiction
  the missuse of opioids causes withdrawal symptoms, which is a serious side effect, tipically
manifest as a use of a drug

Opioid-synthesizing

 opioid cereptors with a capled alfa

 most poweful modulatory system

Opioids

 1973: discovery of morphine receptors in CNS

 1975: Hughes and Kosterlitz have isolated and identified two opioids provided with
morphine-like activity

 Met-enkephalin <- enkephalins -> leu-enkephalin

 endorphins

 dynorphins

 3 molecule families differ for their biosynthetic pathway and anatomical localization

 they share similar amino-acid sequence

 portion recognized by the receptor, the part of the molecule interacts with the same
receptor (same binding site)

Structures of the three opioid precursors

Anatomical distribution of endorphins

Anterior hypophysis

They are stored in cells that also secrete ACTH: hence, ACTH and endorphins are released together

Anatomical distribution of Enkephalins

are stores in neurons, provided with short processes, interneurons, hence, their soma and terminals
localize withn the same brain neuculus

Dynorphins

they are generated from pro-dynorphin, which contains the sequence of three peptides:
neoendorphin, dynorphin A (17 AA) and B (13 AA)

Opioid system: overview

 enkephalinergic neurons

o produce pro-enkephalin

o are provided with short processes

 endorphinergic neurons

o produce POMC
 o provided with very long processes

 dynorphinergic

Receptors

Agonist Receptor type

Enkephalin delta
Endorphin mu
Dynorphin kappa

11.15

Dynorphins

They are generated from pro-dynorphin, which contains the sequence of 3 peptides

Opioid system

 mu and delta receptors have similar activity, kappa receptors are mostly activated by
dynorphin

 mu and delta receptors are associated with mood elevation (neuron activity), kappa is
associated with delevation

 kappa receptors are located in limbic system

 kappa activation leads to dysphoria and anhedonia in our species and anxiety in rodents

 limbic system w amygdala may largely involved in the symptoms of eg. depression

 by interfering with the reuptake of serotonin, might lead to receptor desensivitation

 mainly enzymes in our cells are also regulated by ions

 they act on a sertain number of enzymatic activity, the result is the deasease

 bipolar disorder is tricky, kind of a depression, but has up- and down phrases, you can’t treat
the up and down symptoms with the same drugs

Receptors

 all of them are capled to an inhibitory alfa

 mu and delta mediate opposite effects

 gabaergic interneuron targeting the receptor, the gaba released will be inhibited

 all receptors are involved in mood regulation

 their activation leads to increased domapine activation, they are located in gabaergic
neurons

 kappa receptors are new, focus of interest, with particular reference to the treatment of
depression (especially when they don’t respond to classic antidepressants)

 targeting this system, evolutionary point of view,

Positive reinforcement

 mu and delta, versus kappa

 mu and delta agonists produce positive reinforcement, whereas kappa agonists induce
aversion, hallucination and malaise

 positive emotions, such as pleasure, hedonism or reward, when associated with the ability to
learn from experience, can act to increase the probability of the occuerence of a particular
behavior, a phenomenon called p. r.

11.22 - midterm

11.27

Development of the Central Nervous System

Gene expression

Embryonal and fetal development of human brain: up to 8 weeks

Evolution: increase the surface and the cell numbers, but keep them in a fixed room

We have sulcus and gyrus on the surface, it’s not smooth

Stages of human development: embryonal and fetal development

Every neural precursor undergoes height subsequence phases

 mitosis/proliferation

 migration

 differentiation

 aggregation/integration

 synaptogenesis

 apoptosis

 synapse reorganization

 myelinization

Stages of Development

(KÉPET BEVÁGNI)

purkinje neurons/cells -> the full maturation of this neuron is accomplished by 9 years of age, the
most relevant cells in our cerebellum

The ability to judge the risk and the opportunity to take some behaviors are related to the
functionality of the prefrontal cortex, it is typically an acquisition of an adult -> adolescents usually
take risky behavior because the PFC is not mature yet

Early development

Fertilized eggs: totipotent cells -> can become any type of cell in the body or placenta
Totipotent stem cells -> fate decision

Embryo: blastocyst: pluripotent stem cells (pluripotent cells also can become any type of cells in the
body), we don’t call them totipotent anymore because they are forming the placenta, they are in
charge of providing the proper cells in the embryo

The embryo is already planted in the uterus -> the mother’s blood circular stem is in charge

fate decision 2 -> multipotent stem cells/ starting of gastrulation -> at the end of the gastrulation, the
embryo has 3 layers of cells (ENDODERMA-MEZODERMA-EKTODERMA) -> multipotent cells develop
different type of tissues from the 3 layers

The Embryonal stem cells

The blastocysts cells contains up to 128 and formed after ~5 days of fertilization

Gastrulation: a morphogenetic process which supposed to provide shape, an inward folding

morphogenetic process, produce 3 cell layers

Neural induction:

 At 18 days the embryo begins to implant in the uterine wall, in 3 layers of cells, thickening of
the ectoderm leads to the development of the neural plate (process called neurulation)

 prior to the induction of the neural plate, the cells are undifferentiated, these are the stem
cells -> after the induction, cells are destined to become a neuron

 at 20 days, the neural groove begins to develop -> neural tube develops which is a primer in
the neural system

 by 40 days, the anterior end of the tube develops 3 swelling that become the forebrain,
midbrain and hindbrain

Development of the Nervous System

 gastrulation, neural induction, neurulation, neural tube (at day 21), which contains
neuroblasts (generates neurons and glial cells)

 gastrulation also forms the notochord (a rod-shaped structure of mesodermal cells)

 above the notochord lies the ectoderm that gives rise to the nervous system ->
neuroectoderm

 the notochord also sends signals to make certain neuroectodermal cells become neural
precursor cells, a process called neurulation

 the notochord is only active on a group of cells that are close to the notochord itself

 the cells at the dorsal part of the neural tube are called the neural crest cells

 these cells form the basis for sensory relay neurons to the thalamus

 neural proliferation is the first step of the development of the system

 cell division occurs in the ventricular zone of the neural tube, when they leave the cell
division cycle, cells migrate into their layers
  in adult brain we have neurogenesis of cells which is near the cavity

 neurodevelopment has several stages, and at some points, some cells leave the cycle (POUR
TOUJOURS) -> migration

 cell division -> they split the chromosome -> they split cytoplasm

 cell division can be symmetric with the same cytoplasm and DNA

 also can be asymmetric, the genetic material is the same but the cytoplasmic construction
might be different (the list of proteins)

 transcription factors: they might activate a genetic program which makes it different (since
they sit on the promoter region of the genes)

 the neural tube is uniform, it is going to fold

12.04

Prosencephalon

 telencephalon

 diencephalon (thalamus, hypothalamus, optic cups)

Mesencephalon (superior colliculus

Rhombencephalon

 metencephalon (cerebellum, pons)

 myencelencephalon (medulla)

This patterning of the neural tube is the result of the activity of the morphogenes

 anterior-posterior rostrocaudal patterning

 along the neural tube, there are several regions identified by diff. names

 regulated by homeotic genes

How do these regions know what to become?

 concentration gradient

 gradient of FGF and retinoic acid, affect the expression of homeobox (hox) transcription
factors

 the gradient of the FGF is in the anterior part, the retinoic acid in the posterior part

 rostrocaudal patterning

 the expression of these homeotic genes varies along the body parts, they have a very precise
way of expression, the very precise manner depends on the activity of these molecules

 embryologist did a very elegant experiment – they transplanted parts of the embryo’s tissue
to the head -> they were able to obtain the development

 master transcription factor: they regulate the expression of 150 genes

  homeotic gene: control the pattern of body formation during early embryonic development
of organisms

 they encode proteins called transcription factors, that direct cells to form various parts of the
body

 homeotic gene can active or repress genes, producing effects that are complementary and
necessary for the ordered development of an organism

 each subregion of the body axis will develop a region specific for that area

 humans process 39 HOX genes, divided into A, B, C, D

 changing the expression of even just one homeobox transcription factor can have a huge
effect

Summary

 the rostral neural tube froms the bais of the brain’s subdivisions

 concentration gradeient of FGF and retinoic acid help direct the development of these
subdivisions

 this gradient affect the expression of homeobox (HOX) transcription factors, as know
rostrocaudal patterning

Neurodevelopment – Migration (second step)

Migration and Aggregation

 cells migrate away from the ventricular zone along the temporary network of radial glial
cells, which are present in only the developing neural tube

 the cells of the neocortex migrate in an inside-out pattern: the deepest layers form first, so
the cells of the superficial layers must migrate to them

 only a soma and immature axon at this point: undifferentiated neurons start migration

 differentiation begins as the neurons migrate: they develop neurotransmitter and action
potential

 radial glia

o radial glia cells act as guide wires for the migration of neurons

o migrating cells are immature, lacking dendrites

o cells that are done migrating align themselves with other cells and form structures
(aggregation)

 development of the cerebral cortex

o ventricular zone (VZ) contains the progenitors of neurons and glia

o the first neuron generated establish the preplate (PP), their axons as well as
ingrowing axons from the thalamus, establish the intermediate zone (IZ)
 o neurons of cortical layers II-VI establish the cortical plate (CO) which splits the
preplate into the marginal zone (MZ), or future layer I, and the subplate (SP)

o six cortical layers are visibly overlying the white matter (WM) and the subplate

o neural precursors in the subventicular zone (SVZ)

 role of the reelin protein in cortical development

o reelin is expressed by Cajal Retzius cells, in the layer of the developing cortex

o reelin binds to a receptor, VLDLR or ApoER2 in the surface membrane

o which leads to downstream signaling via Dab1

o gene in coding for reelin are in candidate of autism

Key concept in neurodevelopment:

 spatio-temporal schedule

 activating the right genes at the right place

Axon growth/synaptogenesis

 once migration is complete and structures have formed (aggregation), axon and dendrites
begint to grow to the mautre size/shape

 growth of cones and chemo-attancants are critical

Cytoskeletal elements

 acting molecules

12.11

Semaphorins: stop signals

Synaptogenesis

 formation of new synapses

 depends on the presence of glia cells, especially

 chemical signal exchange

Initiation of synapse

These molecules stick together on the pre- and the postsynaptic side

They mediate between cells

Assembly of active site and postsynaptic density

they are proteins that are sticking together

Genes that encode the proteins neurexin, neuroligin are among the candidates for autism disease

They are relevant for proper synaptic functioning

Autism together with number of neurodevelopment diseases are due to synapse degeneration

Synapse formation itself is quite complex, part of a genetic program

Neuronal death

 between 40-75% neurons made, will die after migration – death is normal and necessary

 part of the genetic program, genetically determined -> genes that trigger cell death needs to
be activated in order to carry this out

 neurons die due to failure to compete for chemicals provided by the targets

The neurotrophic hypothesis

Targets of innervation secrete limiting amounts of survival factors to generate a balance between the
size of the target organ and the number of innervating neurons

The discovery

 frog embryo (vertebrate, has symmetry) -> spinal cord section: elaborated limb: missing
neuron, the other side is healthy

 the removal of a limb bard resulted in a number of reduced neurons in the spinal cord

 they release a trophic factor that influences the survival of neurons

The discovery 2

 transplantation of supernumerary limb bud

 extra limb, extra neurons

Based on the results, Hamburger hypothesized that the targets of innervating neurons provide
signals that recruit undifferentiated cells to develop into sensory or motor neurons -> he was wrong

1942: new hypothesis by Levi-Montalcini and Levi -> they repeated the exp. together in 1949, and
found the results supported the neurotrophic hypothesis

1960: NGF purified

1969: NGF purified to homogeneity

Various neurotrophic factors exist

 neurotrofine

NGF-mediated signal transduction

 NGF acts as a binder

 once the lingand binds to the x, it gets phosphorylated -> this indicates the phosphorylation
of that part
  after phosphorylation, it gets activated

TRK receptor singanling

 when a neurotrophin binds to a trk receptor, the kinase domain is activated resulting in
autophosphorylation

 autophosphorylation

Intracellular signals

 PL 3 kinase

 ras

 PLC activation is associated with the calcium pathway

Neurotrophic receptors

 ligand-dependent receptors

 when the ligand is missing, they act in an opposite manner (they active as well, but they
activate a death program)

 APOPTÓZIS VÉGRE

Apoptotic cell death

 cell shrinks away from neighbours -> chromatin gets condensed -> nuclear and cellular
fragmentation

CNS myelinization

PNS myelinization

Myelinization last up to thirty years

Postnatal Cerebral Development in human infants

 postnatal growth is a consequence of synaptogenesis, increased dendritic branches,

myelination (prefrontal cortex development)

12.13 – nem volt óra

12.18

Epigenetic mechanisms from gene to protein

 DNA transcription to various classes of RNA

 messenger RNA – RNA editing

 mRNA – translation/protein folding

 protein: post-translational modification

Non-coding RNA: regulatory role

DNA assembly in cromatins

  DNA in chromatins

 histone octamers: DNA wraps around it 2 times

 Chromosomes: single molecules of DNA

 Histone octamer: 8 subunits

 nucleosomes: histone + DNA

 2 filaments needs to be separated in order to get access to the nucleotide

Transcription

 DNA has NET- charge; sugar; phosphor;

 refers to the transcription of DNA into messenger RNA

 mRNA has 4 nucleotide bases: purin, guanin, uracil, citozin

 transfer RNA (szállító RNS)

Translation

 covalent group modification

Epigenetic modification

Histone modifications: the histone code

 DNA methylation: pluripotent cell transforms to unipotent cells with a methyl group

 once the cytosine is mutilated, specific molecules bind to it, making the DNA accessible for
the transcription binder -> they will be inactive for RNA polymerase ensime

Gene programming by methylation

 genome -> methyl group -> DNA metylation -> tissue-specific developmental genes

 genome -> development

 genome -> methylation-regulated -> regulated over the time -> inducble genes

 gene expression is fundamental in our life

These are in charge of DNA transcription

The promoter region regulates gene transcription

 cg eyelance (?) is located in the promoter region

 4 nucleotides: adenine, cytosine, uracil, guanine

Novel epigenetic mechanisms

 they stand on the RNA ward (?)

 non-coding RNAs and RNA editing

 coding RNA -> mRNA

  non-coding RNA -> transcriptions RNAs -> tRNA (transfer) and rRNA (ribosome) -> both
participates in the protein synthesis, but not codes proteins

 small RNAs -> sRNA; mRNA (stRNA) ; snoRNA; snRNA

Non-coding RNA

 does not encode for protein

 80% of the genomes of complex organisms in fact transcribed into ncRNAs, many of which
are alternatively spliced and/or processed into smaller products

 3’ and 5’ primus, filaments C-D and A-T

Further reasoning on Epigenetics concept

 epigenetic factors greatly affect phenotype: a gene dosage issue

 genes that are free of mutation may become harmful if they are not expressed at the
appropriate time and at the required level

Lessons from genomic imprinting

 genomic imprinting refers to the epigenetic phenomenon by which a few gene are expressed
in a parent-of-origin specific manner

 this means that

o if the allele inherited from the father is imprinted, it is silenced, and only the
maternal allele is expressed

o if the allele inherited from the mother is imprinted, it is silenced, and only the
paternal allele is expressed

Deletions on chromosome 15 can result in

 Prader-Willi syndrome

 Angelman syndrome

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